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THE UNIVERSITY OF TEXAS 
PUBLICATION NUMBER 5914 
— JULY 15, 1959 



BIOLOGICAL I 
CONTRIBUTlONa: 

- ■ ' ' ■" * -' ^^^ :■ '^^^' •'■■' .■-. ',': f *■■ ' 

A CoUedidn df Essays 'cmd\:.-'-'t^'y'^ / 
Research Articles Dediccdedtof^^^^^^^ . 

JOHN T H O MA S PA XT E R S O N 

on the Occasion of his Eightieth Bi/rthdky. - ^ 



Edited bv Marshall R. meeler 



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575. 1 W 
T365b 



j: UNIVERSITY OF TEXAS 
AUSTIN 12, TEXAi 




UNIVERSITY 
OF FLORIDA 
LIBRARIES 




Copies of this publication may be procured for $2.00 each from 

the University Press, The University of Texas, 

Austin, Texas 



BIOLOGICAL 
CONTRIBUTIONS 



The benefits of education and of useful knowledge, generally 
diffused through a community, are essential to the preserva- 
tion of a free government. 

Sam Houston 



Cultivated mind is the guardian genius of Democracy, and 
while guided and controlled by virtue, the noblest attribute of 
man. It is the only dictator that freemen acknowledge, and 
the only security which freemen desire. 

MiRABEAu B. Lamar 



PUBLISHED BY THE UNIVERSITY TWICE A MONTH. ENTERED AS SECOND-CLASS 

MATTER ON MARCH 12, I Q 1 3, AT THE POST OFFICE AT AUSTIN, 

TEXAS, UNDER THE ACT OF AUGUST 24, I912 



DITED BY MARSHALL R. WHEELER 



BIOLOGICAL 
CONTRIBUTIONS 

A Collection of Essays and 
Research Articles Dedicated to 
JOHN THOMAS PATTERSON 

on the Occasion of his 
Eightieth Birthday 



THE UNIVERSITY OF TEXAS : AUSTIN 



i^ 



1^ 



73 6^6-J- 



J. T. P. : An Abstract 

Knowing you 

As teacher^ colleague, friend. 
Has been to drop the plumbline 
Of integrity beside the words 
And find them true. 

That just acclaim 

The world of science has given unstintingly 

Rests well on you 

To whom the day's work means more 

Than fame. 

The years will tell 

Your best reward — a growing company 

Who teach and work more knowingly 

Because you worked and taught 

So well. 

.... Linda Wharton McDanald 
Lamarque, Texas 




JOHN THOMAS PATTERSON 



Contents 

Introduction: John Thomas Patterson - - 9 

C. P. Oliver, Austin, Texas 

The Making of a Scientist - - 1^ 

Carl G. Hartman, North Plainfield, New Jersey 

High Islands and Low - - 25 

Warren P. Spencer, Wooster, Ohio 

Darwin's Influence on the Study of Genetics and the Origin of Life.. 49 

James F. Crow, Madison, Wisconsin 

Tumors in Drosophila .- - - ^' 

Walter J. Burdette, Salt Lake City, Utah 

Sex Balance in Drosophila melanogaster: Aneuploidy of Short Regions of 

Chromosome 3, using the Triploid Method - 69 

Sarah Bedichek Pipkin, Washington, D.C. 

^It^ene-Environment Interaction in Relation to the Nutrition and Growth of 

Drosophila - "^ 

Forbes W. Robertson, Edinburgh, Scotland 

Observations on Variegated Position Effects in Drosophila melanogaster 99 

George H. Mickey, Baton Rouge, Louisiana 

Telomeres and Terminal Chiasmata — A Reinterpretation 107 

M. J. D. White, Melbourne, Australia 

Dominant Lethal Mutation in Irradiated Oocytes 113 

D. R. Parker, Riverside, California 

Mammalian Chromosomes in vitro. XI. Variability among Progenies of a 

Single Cell - - - 1^9 

T. C- Hsu, Houston, Texas 

Heterochromatic Control of Position-Effect Variegation in Drosophila 135 

William K. Baker and Janice B. Spofford, Chicago, Illinois 

Genetic Studies on the Cardini Group of Drosophila in the West Indies 155 

William B. Heed and Nikam B. Krishnamurthy, Tucson, Arizona 
and Bangalore, India 

A Nomenclatural Study of the Genus Drosophila - - 181 

Marshall R. Wheeler, Austin, Texas 

Genetic Studies of Irradiated Natural Populations of Drosophila. III. Ex- 
perimental Populations of Drosophila ananassae derived from 

Irradiated Natural Populations - 207 

Thomas G. Gregg, Madison, Wisconsin 



Contents ' 

Genetic Studies of Irradiated Natural Populations of Drosophila. IV. 1968 

Tests ^^^ 

Wilson S. Stone and Florence D. Wilson, Austin, Texas 

Some Values of Endomitosis - - - 

Theophilus S. Painter, Austin, Texas 

Alexander von Humboldt and the Science of the Nineteenth Century tt... 241 

Adolf Meyer-Abich, Hamburg, Germany 
Molecular Configuration, Synthesis and Gene Action - 261 

R. P. Wagner, Austin, Texas 



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Introduction 



JOHN THOMAS PATTERSON 

A Tribute on His 80th Birthday 
(November 3, 1958) 



C. P. OLIVER 

Chairman, Department of Zoology, 
The University of Texas, Austin 

Very few scientists have the opportunity to recall a half of a century of past 
experiences and activities during which they played a part in developing a field 
of science. Any man must have a feeling of satisfaction when he can see how 
much his own work has meant to a science. John Thomas Patterson is one ot 
those few, fortunate scientists. i • i 

Fifty years ago J. T. Patterson began his scientific career at the relatively 
new University of Texas. The biological research at the school was not, at that 
time influencing scientific opinions to any great extent. Actually, the Depart- 
ment of Zoology (known as a School in those days) was only about ten years 
old- and its predecessor, the School of Biology, had been organized less than ten 
years earlier. For fifty years. Professor Patterson continued his activity as a 
member of the faculty of the school. By his own Herculean efforts as an investi- 
gator and by following a sound pohcy as an administrator, Patterson has been 
responsible for developing a department which has gained international recog- 
nition for its research in biology. 

Soon after he came to the University of Texas, Professor Patterson began a 
teaching program that attracted the attention of the more alert advanced stu- 
dents They were attracted to him by his friendliness, scholarly attitude, and 
enthusiasm. Science was truly a big part of his everyday life. He came to the 
laboratory early and he stayed late. His students developed a devotion for their 
teacher and the devotion was lasting. Those students are justly proud of their 
associations with their teacher. They have a right to their pnde. They cannot, 
though, keep all of the feeling and elation for themselves. The man's heart was 
big enough to take in more than his own students. We who merely worked m the 
same laboratory with Professor Patterson, as students or colleagues or visitors 
have also enjoyed the man's warmth of friendliness, his spirit of cooperation, and 
his words of encouragement in our work. Either by his spoken word or m his 
thoughts every one of us refers to the likeable "bundle of enthusiasm" as Pat. 
On November 3, 1878, John Thomas Patterson was born near Piqua, Ohio. 



10 The University of Texas Publication 

He received the Bachelor of Science degree from the College of Wooster, Ohio, 
in 1903. Patterson then joined the faculty of Buena Vista College in Storm Lake, 
Iowa, where he was Professor of Biology during 1903-1905. He then entered 
the University of Chicago to complete his university education. In 1908, the 
University of Chicago awarded Patterson the Doctor of Philosophy in Zoology, 
summa cum laude. 

In 1908, the new Dr. J. T. Patterson accepted an invitation of the young 
University of Texas to join its staff as an Instructor of Zoology. He was one- 
half of the departmental faculty, the other being Dr. H. H. Newman who 
came to the department as Head also in 1908. Dr. Patterson was promoted to 
Adjunct Professor in 1911 and to Professor in 1913. He was elevated to the 
position of Distinguished Professor when that rank was established by the 
University in 1937. With his retirement in 1955, he was named Emeritus 
Professor. 

Two predecessors (William M. Wheeler and T. A. Montgomery) transr 
mitted to Patterson and Newman an atmosphere for research in the institution. 
The new staff, though, began to develop experimental research in the laboratory. 
They set out purposely to develop strong undergraduate courses, few in number, 
but selected to meet the needs of a general education as well as to interest stu- 
dents in and to prepare them for graduate work. 

In 1911, Professor Newman resigned and Professor Patterson became Chair- 
man of the School of Zoology. He remained Chairman for the term 1911-1913 
and again for the period 191 6-1 925 . 

This appointment had a strong and lasting beneficial effect on Zoology at the 
University of Texas. He established the policy of electing new staff members 
who showed promise and who were especially trained to teach and direct some 
particular phase of Zoology. Courses were kept to a minimum number to give 
the new teacher an opportunity to carry on productive research. Facilities as 
well as time were made available for scholarly productivity. Professor Patterson 
became Director of Research in Zoology in 1928 and retained the position until 
his retirement in 1955. In this position he was able to help promote active 
research programs of his colleagues. 

As a teacher and as a researcher. Professor Patterson has had a profound 
influence upon the development of genetics and cytology. When he first accepted 
appointment with the University of Texas, Patterson taught cytology, physi- 
ology, and comparative vertebrate anatomy. In 1911, following Newman's 
resignation, he assumed the duties of teaching heredity and genetics. As new 
staff members were added to handle each his own specialty, Patterson gave up 
courses to them and took over other duties. During his period as a teacher in 
the department, Patterson has taught twenty-six different courses. 

The influence Patterson has had on research in genetics and cytology is the 
result of his own productivity and the effect he has had on the research of his 
colleagues. His intellectual curiosity led him to ponder over unknown recesses 
in the exoskeleton of biology. Then experiments were designed to explore the 
unknown. Once that had been done, his whole being was poured into the 
investigations, regardless of the subject, until an answer appeared. Collaborators 
in the department permitted a broader scope of the research. No pressure was 



Oliver: Introduction 



exerted though, to get the staff to work in any particular field. Each was per^ 
Lu d to do what he'preferred to do. But the spint of the department was one of 
Tooperation, an example set by Patterson. Research reports came rapidly from 
the Department of Zoology and represented m many instances a community 
effort to open up new avenues for the study of genetics and cytology. 

Professor Patlrson's early research was m embryology and polyembryony^ 
He gave us the story of the monozygous origin of the quadruplets produced by 
fhe frmadmo. After'the discovery by his colleague, H. J. Muller, that mutanon^ 
ran be induced by irradiation, Patterson began an active research program m 
gene^l problems. This included studies in the effects of irradiation and he use 
of material so produced to investigate other biological problems He determined 
iTmutations'can be induced m somatic cells of Drosophila. W.th .« e t a" 
colleagues, he established the relationship between chromosome parts and sex 
determination m Drosophila, and showed that mutations can be reversed. 

An interest in evolution and genetics led Patterson to studies of the specie 
relationships in the genus Drosophila. Intensive collecting excursions m the field 
became necessary. This study began in 1933 and sample collections were mad 
of wild types of Drosophila in Texas and surrounding areas. Results were so 
promising that in 1938 a large and definite program was organized^ Many trips 
were made. The interest was great. Any one who made an automobile trip with 
Patterson to collect specimens might wonder whether or not he would ge 
around the next curve. But he was confident that the trip would end with a mass 
of material on hand and enough work to last the laboratory for some t ime^ 
Because of the success in the near areas, collecting trips were made^to other 
parts of the United States and to areas to the south. The aboratory became a 
cooperative caldron for studies of taxonomy and ^Pf '^^/^.l^^^""^*^^?" '" "'t^ 
to report the investigations in more detail, a series of bulletins was published by 
the University of Texas Press. These "Studies in the Genetics of Drosophila 
were edited by Patterson until his retirement from the facu ty. In the bulletin 
are descriptions of species, reports on hybridization tests, relationships between 
species as shown by genetical and cytological tests isolating ™-hanisms, and 
other genetical studies. In 1952, a book entitled "Evolution in the Genus 
Drosophila" was authored in collaboration with W. S. Stone. The If^t paper by 
Patterson was published in 1957 in the University of Texas Publication 5721. 
Patterson has gained the recognition of and has been honored by his colleagues 
in the United States and m foreign countries. He was elected to membership m 
Sigma Xi by the Chapter at the University of Chicago. That school and, later, 
the College of Wooster elected him to Phi Beta Kappa. He received an honorary 
Doctor of Science from the College of Wooster. In 1913. he was elected to mem^ 
bership in the Town and Gown Club in Austin, Texas. A number of professiona 
societies have elected him to be their president; they are, Ae Texas Chapter of 
the Society of Sigma Xi (first president) , American Society of Zoologists (1939), 
Society for the Study of Evolution (1947), and Genetics Society of America 
(1954) In 1956, he was made honorary member of the Genetics Society of 
Japan In 1941, Patterson was elected to membership in the National Academy 
of Sciences. He received the Daniel Giraud Elliot Medal in 1951 from the 



12 The University of Texas Publication 

National Academy of Sciences for his study of isolating mechanisms reported 
in 1947. 

Students who received their training under Professor Patterson's direction 
remain loyal to him. Evidence for this is definitely shown by the number who 
attended the eightieth birthday celebration. As one who was not Patterson's 
student, I am permitted to say that the scientific success of his students shows the 
soundness of his training in the ways of science. The enthusiasm, devotion to 
science, and cooperativeness rather than strict competition and individualism 
leached down to them and became permanently incorporated. 

From 1913 through 1952 Patterson was major adviser for 31 successful M.A. 
candidates. The first student to be granted the Ph.D. degree under Patterson 
(and by the University of Texas) received the degree in 1915. He has had 
twenty-nine of his candidates receive the Ph.D. degree from 1915 through 1955. 

Professor Patterson's eightieth birthday offered a happy occasion for his 
colleagues, former colleagues, and students to express their gratitude and esteem. 
The contributions in this anniversary volume are offered as a small tribute to 
this remarkable man. 



The Making of a Scientist 

CARL G. HARTMAN^ 
North Plainfield, New Jersey 

It was Professor Patterson (hereinafter called "Pat") who invited me in 1912 
to join his Department as Instructor. As a result he was "stuck" with me for 
thirteen years, when I joined the Department of Embryology of the Carnegie 
Institution of Washington in Baltimore. In those thirteen years I learned much 
that influenced my later career. Incidentally, I also enrolled as Pat's student, and 
being successful in winning a Ph.D. degree, I have the honor of holding the first 
doctorate conferred by the University of Texas. 

In an attempt to explain how I came to be a scientist, I must furnish a some- 
what detailed background. I may, at the outset, state categorically that I never 
consciously said to myself: "I am going to be a scientist." I have sympathy for 
the High School youth who cannot make up his mind, in view of the fact that I 
was 33 before I reached this stage myself! I have, in fact, been a drifter, partly 
for reasons beyond my control (or so I imagined), always following the next 
line of endeavor that interested me at the time and which was often quite remote 
from natural science. However, it will be apparent from the following that any 
young man with ambition, energy, a fair IQ, and good health can rise well up 
the ladder of accomplishment in science if he has a mind to do so. 

My tendency to take up various Hues of employment was perhaps a matter of 
expediency, but it shows that I had not set my goal early, certainly not before I 
had reached the age of 32 when I was lucky enough to be able to join Pat's compe- 
tent staff at the University of Texas. Rumor has it that Pat, before he offered me 
the job, wrote my former Professor, Dr. W. M. Wheeler, asking whether he 
thought that I would "stick" this time. I stuck. 

It all began back in my early days in Iowa, my native state. Not getting along 
with the male teachers in the German-American parochial school, my mother 
and my preacher father (D.D., M.D.) sent me to the public school where I 
entered the eighth grade. In the first two years of the Clinton, Iowa, High School 
I had the only pre-university science course, physics. This was well taught, I 
think, for it interested me greatly and for some years I kept my notebook con- 
taining drawings of all the apparatus the school possessed. 

Since my father beheved that a classical education was the proper preparation 
for any career, I was naturally required to begin Latin early. My High School 
studies were no challenge, as I remember them, which gave me the opportunity 
to read most of the fiction in the small school library. I also had a bout with 
several hundred dime novels, the reading or even possession of which was strictly 
prohibited. I recall Horatio Alger, Oliver Optic, and Henty, wept at the death of 
the Last of the Mohicans and rejoiced with Tiny Tim. From fiction I moved into 
history. No popular science books such as the youth of our day have available 
had been written — unless one excepts those of Jules Verne. 

1 B.A. 1902: M.A.. 1904; Ph.D., 1916, The University of Texas. 



14 The University of Texas Publication 

My most profitable and awakening year was spent at Wartburg College, a 
German-Lutheran pre-theological preparatory school. I had not the slightest 
thought of becoming a minister, but the cost of tuition and board ($250 a year) 
was in keeping with my father's ministerial income (even when supplemented 
with some fees from his small medical practice) and a family of seven children 
plus two adopted ones! At Wartburg, Greek and Latin were stressed — to be trans- 
lated into German. In fact, all of my studies were in German: German literature 
and rhetoric, church and secular history, and weekly essays. There was no Math, 
and no study at all in English. The professors were graduates of German Uni- 
versities and Gymnasia and were "tough hombres." For the first time in my life 
I really learned to study. Something like their demands, which obtain generally 
in nations with which America is in competition, is now needed in this country. 
The year at Wartburg was most valuable to me, as it gave me a command of 
German almost equal to that of English. This later proved a great asset in my 
future research work. 

At the end of the Wartburg session I took the entrance examination at the 
University of Iowa and was admitted, probably because of my fluency with the 
languages. Of my year as a green Freshman I recall only the courses in English 
and mathematics. But the University, where my father had taken his M.D. 
degree ten years previously, seemed to me an imposing institution. The summer 
between Wartburg and the University I spent as a carpenter's assistant in 
LaPorte City, Iowa, my parents having moved to Texas and there being no funds 
for my traveling back and forth between Iowa and Texas just for a vacation. 

At the end of the 1896-7 session at the University of Iowa it became necessary 
to have my eyes looked after. The famous ophthalmologist, Dr. Schneider of 
Milwaukee, diagnosed the trouble and referred me to Dr. Hilgartner in Austin, 
Texas, near which city I was to live for the next three years. As will be noted 
below, my eye trouble greatly affected my career, at least for a number of years. 

Eye rest was prescribed. What better place for this than a farm? The next 
three years I spent on a farm at Pflugerville, 13 miles from Austin and 10 miles 
from Round Rock, and from the nearest railroad — i.e.^ before the M.K. & T. came 
through Pflugerville and destroyed its rusticity. My apprenticeship was soon over 
and I became a full-fledged farm hand at $3.50 a week plus room and board. A 
city boy suddenly found himself at a rugged calling. All the activities, including 
five meals a day, were new to me, from work in the cotton and corn fields to 
driving a few thousand head of sheep across the prairie. My first initiation to 
surgery and endocrinology was participating in the castration of mature boars, 
and sterilizing the wound with a handful of mud from the nearby pond or "tank" 
with its coating of algae — and its content of antibiotics. I should explain, perhaps, 
that a boar has little market value until his scent glands atrophy as a result of 
castration. 

One day the local school board urged me to get on my horse, ride to Austin, and 
take the examination for a teacher's certificate. This I did, bringing back a 
temporary certificate (which was later made permanent). When, many years 
later, I was head of the Department of Zoology at the University of Illinois, and 
since there was certificate reciprocity between Texas and Illinois, I was the only 
member of that large and competent faculty who was "certificated" to teach in 



Hartman: The Making of a Scientist 15 

the public school of that state! But for two winters at Pflugerville I taught the 
local school of 50 pupils in eight grades for $50 a month. 

I did pick up a little science on the farm. The Texas prairie produced many 
handsome flowers in gorgeous profusion-roadways lined with the pmk evening 
primrose, the prairie blanketed with gaillardias for a thousand miles. I wished 
to get better acquainted with them, so I secured a copy of Gray's Botany and 
learned the technical terms necessary to run the keys to the flowering plants 
("repand margin," "mucronate pointed," "parietal placenta"). Then with the 
further aid of Coulter's Botany of Western Texas I proceeded to determine many 
species and to press them in amateurish fashion, but well enough to have them 
accepted by the University Herbarium. r i j tt^ i i 

At this time I also succumbed to another hobby, photography. I had a Kodak 
box camera of universal focus and I took a lot of pictures, developmg the plates 
at night and printing the pictures on "Solio" paper in direct sunlight. Many ot 
these I hardly surpassed later with a fine Leica camera; the negatives and plates 
are still in perfect condition after 60 years. My somewhat crude skill came m 
good stead in my researches later. Photography is a useful art, one which 1 
recommend to all young scientists. It would save time, headaches and money, 
however, to get started under the tutelage of an expert. 

Our country homes were heated with wood stoves, the kitchen being the center 
of attraction. For the winter's wood supply we had to drive to the woods about 
Round Rock in the heat of summer and chop the necessary supply. This is a 
region in which bat caves abound and I saw for the first time clouds of bats 
emerge from a cave exit like smoke from a smoke stack. I was to visit these caves 
frequently in later years. The local community came to these woods for posts 
with which to build our own telephone line. We all cooperated m this project 
setting the poles and stringing the wire. I must have dug 30 postholes myself! 
Among the first messages that came over the wire was news that the <-oiorado 
River Dam at Austin had burst. I at once saddled my horse and rode through the 
mud knee-deep in places, to the city. With my box camera I took pictures which 
are now of some historical interest, since with the series of new dams up the 
river such a flood is not likely to occur again. 

In the spring of the same year I decided to visit the University to consult with 
Dr William Bray, Professor of Botany, concerning the purchase of a microscope. 
His advice was: "Don't; come instead to the University and we will lend you a 
microscope free." It was good advice and I took it. The office in which I was mter^ 
viewed by Professor Bray I later occupied as professor myself. The room suited 
me in spite of the odor of bats that occupied the spaces in the window frames. 
When I wanted a supply of bats, all I had to do was roll my office chair over to 
the window and, holding a net over the exit, capture all the bats I needed at the 
time Studies of bat embryology and habits occupied several of my students. Few 
papers resulted from the extensive embryological collection, however, since, as 
will be described later, my attention was largely absorbed by the opossum. 

I must digress here to relate my activity during the summer preceding my 
enrollment in the University in the fall of 1900. In a preponderantly German- 
speaking farm community southeast of Austin there were many immigrants who 
wanted their children to learn to read and write their native language and at the 



1 6 The University of Texas Publication 

same time to learn Luther's Catechism. I rode out into the community and 
drummed up pupils for a two-month's summer school — at $10 per pupil (in 
advance). School "kept" from 7:00 to 12:00 a.m., and pupils rode as far as seven 
miles to the school. I made no extra charge for picking out tunes on the reed 
organ to accompany the singing of Sunday School and other German songs for 
the children. I had been raised on these songs and I still know^ many of them 
"by heart." 

A more profitable phase of that summer's activities was my pursuit of Latin. 
I wrote out in English four books of the Aeneid, wrote out in Latin all of the 
exercises in Collar and Daniel's Prose Composition. Professor Penick refused to 
wade through these documents, but he made a bargain with me: if I made an A 
in the next advanced course in Latin, he would give me college credit for my 
summer's work. This I did, thanks in part to the basic training under the Wart- 
burg professors, and in part to the inspiring teaching of Professors Penick and 
Fay. 

After two years I secured the B.A. degree, still majoring in languages, and two 
years later I received an M.A. in Zoology and German. I "worked my way 
through" the University, but it was easier then than now. One year I was a 
student assistant; the last year I had the next higher rank, tutor. Earning one's 
expenses while going to school was, and is now, at the expense of social and 
recreational activities. However, I did keep my church affiliation, served as Sun- 
day School superintendent, and for several years conducted church services once 
a month while the minister took his turn for a morning service in a country 
congregation. At that time all of the services were still conducted in German. 

I suppose that the making of a scientist depends more on the influence of some 
one teacher than on any other factor. In my case it certainly was William 
Morton Wheeler, Professor of Zoology at the University of Texas during 1899- 
1902. He was ably assisted by Augusta Rucker who later took a degree in medi- 
cine at the Johns Hopkins University and then began practicing medicine in 
New York City. Perhaps it was Dr. Wheeler's reputation among the students 
which induced me to register for Zoology immediately upon my arrival. The 
move was a lucky break for me. 

Dr. Wheeler was considered by many to be the most brilliant biologist in the 
country. He was also a great linguist, a voluminous reader in belles lettres as well 
as science, particularly the philosophical aspects of the subject. He was a close 
friend of Bergsson whom he visited occasionally in Paris. 

Wheeler's lectures were fascinating. I took his entomology course and was 
permitted to accompany him on tramps in the field in those years when he was 
writing his famous "An?5." I like to claim that I helped him write the book by 
carrying his vasculum and eating his milk chocolate! I did, however, earn the 
chocolate by making some photographs for the book and by helping with the 
digging into ant burrows, notably the extensive nests of the leaf-cutting Atta 
fervens. Dr. Wheeler was also somewhat of an ecologist, and the rich fauna of 
the Austin region was spread out before me with considerable system and order. 
The distribution of plants and animals was interpreted for me by a master with 
all of nature as a laboratory. For example, it was apparent that the Carolinian 
i auna of the Atlantic and Gulf region followed up the flood plains of rivers, inter- 



Hartman: The Making of a Scientist 1 7 

digitating with the Sonoran or Western fauna of the uplands between river 
valleys. A stone's throw would make a difference in the habitat of the agricul- 
tural ant, Pogonomyrmex barbatus, and the fungus-growing Trachymyrmex. 

Dr. Wheeler left Texas to accept the post of Curator of Insects at the American 
Museum of Natural History in New York, inviting me to go with him. When 
he soon thereafter went to head the Bussy Institute of Harvard he again offered 
me an assistantship, which I also refused. My brilhant fellow student, Charles 
Brues, accepted and later succeeded his mentor as Professor of Economic Ento- 
mology at Harvard. Brues' boon companion, A. L. Melander, became a prom- 
inent entomologist out on the west coast. Other graduate students who came 
under Wheeler's influence in Texas at the turn of the century were: Walter 
Hunter, the psychologist; the gynecologist Harvey Matthews; and Jesse Mc- 
Clendon, the biochemist. As for myself, I applied at Columbia University for a 
fellowship in Germanic Languages and was, fortunately I think, turned down. 

Dr. Wheeler was at his best with graduate students; he was bored with under- 
graduates. Witness the following three instructions I received in the summer of 
1901 from his summer haunts at the Woods Hole Marine Biological Station. The 
diction is mine. 

June 15. "For the fall class in entomology lay in a supply of well-fixed hell- 
gramites and cockroaches." Both were easy to secure; the former abounded under 
rocks in the river and were easy to get at low water; the haunts of the latter I 
knew very well for I had previously worked in a local bakery. 

July 15. "As I shall be very busy, I want you to take over the laboratory in 
Entomology." 

August 15. "I will be too busy to fuss with the Entomology course at all; you 
are to take full charge of the course." That was my initiation in the teaching of 
science! The experience in entomology brought me an appointment for the sum- 
mer of 1904 as Field Entomologist to study the cotton boll worm. Unfortunately, 
a bout with malarial fever prevented me from getting this desirable experience. 

As someone perhaps truthfully put it, when Dr. Wheeler was through with 
Texas ants, he was through with Texas. He was succeeded by another great 
Zoologist, Dr. Thomas H. Montgomery, an authority on sex chromosomes. (Not- 
ing that the X-chromosome was superior to the Y-chromosome, he wrote an 
interesting paper on "-The Superiority of the Female Sex'). He remained at the 
University for only a few years, and was succeeded by H. H. Newman, an 
authority on twinning, who came in 1908 bringing Dr. John T. Patterson with 
him. 

As I had made little progress in one year on the subject I had selected for my 
master's thesis, the embryology of the bat, I decided to spend the summer of 
1903 studying solitary wasps which I had noticed frequently on my tramps 
through the woods with Professor Wheeler. In two months of roaming along the 
sandy, open post-oak woods on the banks of the Colorado River, I filled a dozen 
notebooks with my observations and experiments and made many photographs. 
Result: my Master's Thesis on ''The Habits of Some Solitary Wasps of Texas''; 
it was later published as a Bulletin of the University of Texas, and I occasionally 
see it listed by dealers in old books even today, 50 years after publication. I still 



18 The University of Texas Publication 

have solitary wasps as my hobby, making snapshots and motion pictures and 
pubhshing papers for technical and popular journals. 

Dr. Newman returned to the University of Chicago in 1911, from which time 
on Pat guided the destiny of the Zoology Department, bringing it to the high 
status which it has now enjoyed for many years. Under Wheeler and Mont- 
gomery the department was a one-man show; Pat, on the contrary, built up a 
well-rounded departmental staff of which the University may well be proud. 
Not only was he later elected to the National Academy of Sciences, but three of 
the men whom he added to his staff have similarly been honored. 

As I was about to obtain the Master's Degree under Montgomery in 1904, I 
applied for and was accepted as a research fellow to Dr. Edwin Conklin at the 
University of Pennsylvania. This was the year that our late colleague and friend, 
Dana CasteeL finished the work for the doctorate under Conklin. After reflecting 
on the condition of my eyes, I decided against accepting this $750 fellowship, one 
of the best in the country. Professor Montgomery was furious, and told me so in 
unmistakable terms. This was another turning point in my career — away from 
science. While in general I was attracted to scientific investigation and felt that 
I had had some success as a teacher, I had no set goal; I was still drifting. 

This time I entered politics. I ran for the office of County Superintendent of 
Public Instruction in Travis County and, fortunately or unfortunately, was 
elected — three times in fact, the last two times without opposition. For the period 
of my service I traveled, mostly on horseback, over the county's 1037 square 
miles, visiting its 1 70 schools, several of which were 40 miles away in the hills. 
In this day of anti- segregation I recall with some satisfaction that I was the first 
to pay some attention to the Negro schools of the county. I attended Negro com- 
munity gatherings, held Negro Teacher's Institutes the same as for the whites. 
I was told that this was quite an innovation. That the people approved was 
demonstrated by my being twice re-elected. 

What about Science during the five years of my running over the county? 

There was much of interest in my environment. Travis County, Texas, is one 
of the most interesting areas imaginable; its geology determines the fertility of 
the soil and thus the economics as well as the ethnology of the population. A 
gigantic fault line, a thousand miles long, divides the county into two vastly 
different parts: a fertile two-fifths of black land to the east of the line, where 
thrift and prosperity are evident in the large painted houses and the big red 
barns; and a cedar-clad western three- fifths consisting of limestone hills among 
which the Colorado River meanders in great bends. 

The hill country is sparsely inhabited. In conversation with some old-timers 
I learned what ecologists had overlooked — that before the days of barbed wire, 
when the Indians used to burn off the grass of the prairie, these hills were covered 
with a luxuriant growth of grass. When the salutary annual burning no longer 
kept down the cedar trees, these sprang up and choked out the grass, and the 
rains washed the top soil into the Gulf of Mexico. 

Cedar and liveoak are the predominant trees in this section. The question once 
came up at the University as to the rate of growth in these two species. This was 
determined in the following manner. In looking over back numbers of the Austin 
Statesman while collecting material for my book, ''The History of County Super- 



Hartman: The Making of a Scientist 1 9 



vision in 



Texas;' I learned that the Capitol of Texas was built of limestone. This 
splendid structure, contrary to popular belief, is merely veneered with granite, 
in response to the insistence of the owners of Granite Mountain at Llano. The 
limestone was quarried at Oak Hill to which the contractor built a railroad. Any 
trees now standing in the railroad right-of-way, I reasoned, must have grown up 
since the railroad was abandoned in 1886. So about 1923 a group of us (H. J. 
MuUer, George Finlay Simmons and I and our wives) held a picnic on the old 
right-of-way, sawed down two of the largest trees of both species and counted the 
tree rings. Result: cedar and liveoak grow at about the same rate, in ^^ to 40 
years to a diameter of about 12 inches, if I remember correctly. 

After resigning as County Superintendent of Schools I was fortunate in receiv- 
ing an appointment as Professor of Biology at the Sam Houston State Normal 
Institute, now the Sam Houston State Teachers College, the oldest institution of 
college level in the state. Again I thought that this would be my life work. No- 
where have I encountered deeper and more sincere devotion to teaching and more 
earnestness on the part of the student as at Huntsville. In retrospect I feel that 
the three-year period spent there was among the most satisfactory of my life. But 
the question before me at this point is: did these three years of science teaching 
and study lead me closer to a scientific career? 

In spite of having to teach thirty classes a week and to serve on several faculty 
committees, I am able to recall much interesting study on my part, especially in 
Botany in which I was poorly prepared. Every pleasant Saturday found me out 
in the piney woods botanizing. I knew where to find illustrative material for the 
class: Marchantia, fruiting mosses, and fern prothallia. In small lakes and ponds 
I found Chara and Nitella, the latter, along with stamen hairs of T radescantia, 
useful in observing the flowing of protoplasm. I also learned to induce conju- 
gation in Spirogyra when wanted. In one lake, I am now convinced. I discovered 
Chaos chaos, the giant ameba, which I later saw in considerable numbers in Dr. 
Kudo's laboratory in Illinois. However, at the time I made nothing of the dis- 
covery because a well-known protozoologist insisted that what I had seen was not 
an ameba but a slime mold. But, slime mold or ameba, my students were en- 
thralled at the sight, as I was also. 

Having had some experience with systematic botany of the flowering plants, 
even though this was self-taught, I prepared and published in the Bulletin of the 
University of Texas (1913) a ''List of the Trees and Shrubs Occurring in the 
Vicinity of Huntsville, Texas,'' with notes on the ecology of the region. Solitary 
wasps abounded everywhere and my old friends were not passed by unnoticed. 
I was the first to record the manner in which Eumenes belfragei builds and 
"decorates" her beautifully symmetrical jug-shaped nest {Animal Behavior, Vol. 
I) ; I sent two undescribed species of ants to Dr. Wheeler, with ecological notes— 
these species, which raise fungus gardens on caterpillar droppings, were Atta 
hartmanni and Trachymyrmex caroli. 

The various materials that I gathered on my forays were shared with my 
students. I was learning along with them. I feel that, while I was too immature 
scientifically to impart profound generalizations, I was able to awaken a warm 
interest in the study of plant and animal life. I have ever since held that engen- 
dering enthusiasm is more important than handing out facts, and I have found 



20 The University of Texas Publication 

some graduate assistants better at this than some permanent members of the 
staff. I have myself taught things that afterward turned out to be wrong. What of 
it? The wide-awake student will find out these mistakes for himself if he has been 
taught not to believe everything he reads or even hears from his professors. 

Many years ago, long before the University building boom, Pat invited Pro- 
fessor Conklin to deliver a lecture on Evolution. This was delivered in the Meth- 
odist Church, the auditorium of the old main building having been declared 
unsafe — structurally unsafe, that is, not "unsafe" for a lecture on evolution. This 
was before the Scopes trial! On this occasion Pat asked me to show Conklin a live 
tarantula out in nature, and we had no trouble doing so. Facetiously, the pro- 
fessor asked if we had "planted" the spider for his special benefit. I also had the 
pleasure of piloting Dr. H. J. Comstock of Cornell around among the hills and 
into the bat caves. In one cave he found a louse which proved to be a new genus, 
and which he promised to name after me! 

A chance remark by my father at about this time probably changed my life's 
work. Noting that I made up the entire biological faculty of the Huntsville 
college, he remarked, "Now you are the one-eyed king among the blind," quoting 
a German expression. I got the point, and thus after three years at the Sam Hous- 
ton Institute I decided to accept Professor Patterson's offer of an instructorship 
in his department. My motive in studying at a large center of learning was 
genuine; I recall that at Huntsville my salary was $2500 (good for that day) and 
I left it to accept a job at less than half that. Besides, at Huntsville I had a cow, 
300 chickens, a large garden and plenty of credit at the grocery store in exchange 
for produce! What a bonanza to leave behind! 

This discrepancy was, however, cancelled out by a streak of good fortune 
following a burst of unusual hard work. The last year at Huntsville, Dr. Louis 
Bibb, a young Austin physician, and I wrote two books for Texas schools: the 
First Book of Health for the third grade, and The Human Body and Its Enemies 
for the fifth grade. In September we defeated all of our "Yankee" competitors by 
securing the state adoption. How could we lose? Louis Bibb was secretary of the 
State Medical Society and I had made a reputation as an "Educator." Besides, 
did we not mention Texas a hundred times in the books? Our first year's royalties 
amounted to $7,000 for each of us, truly a magnificent sum. 

I was now embarking, with Pat's encouragement, on some researches that were 
to occupy my waking hours for the rest of my life. I had at last, thanks to my 
new environment, found my calling, and the road before me was clearly out- 
lined. Pat's offer to join his department included one "perquisite of office": the 
privilege of studying the embryology of the opossum — at my own expense! At 
that time the department was poor and there were no facilities in 1913-1915 for 
keeping animals on the campus. Even when the opossum collecting season was at 
its height in February and March, 1917, when we collected several thousand eggs 
and embryos, we had only primitive facilities — wooden cages built by myself 
under one end of an old army barracks which was used for laboratory instruction 
before the discovery of oil on University land. In 1923, however, Pat secured 
legislative appropriation for a new Biology building. 

On my arrival in Austin I had found Pat engaged in what to me were sensa- 
tionally interesting research projects. It was in his laboratory that I had the thrill 



Hartman: The Making of a Scientist 21 

of seeing my first mammalian embryonic vesicle in the open uterus of the arma- 
dillo. Even after handling several thousand vesicles of the opossum and more 
than a hundred from the monkey I still recall the impression which that first 
armadillo vesicle made on me. 

A further most interesting series of observations to which Pat introduced me 
involved the life history of Paracopidosomopsis, the tiny wasp parasite with the 
big name. This study was but an expansion of Pat's interest in polyembryony 
which the wasp exhibits in an astounding manner, for a thousand offspring often 
arise from a single egg. The wasplet is a potent parasite of the cabbage cater- 
pillar. These and other basic studies which greeted me in the Department of 
Zoology were eye-openers and inspiring activities. I did not myself "go to the 
ant, thou sluggard" as Professor Wheeler would have ordered, but, following 
Pat's example, I selected the opossum for my researches for the next twelve 
years. 

Why the opossum? For the same reason that Pat selected the armadillo: the 
prairie and the creek bottoms were full of both species and nobody before us had 
yet bothered to study the development of these, the most bizarre of all American 
mammals. It was at the University of Texas that the essential characteristics of 
their mode of reproduction were cleared up. 

For several years, with the aid of students (notably Kenneth Cuyler, now at 
Duke University, and George Hamlett, now at Louisiana Medical School) I 
hunted the animals in the field. I also took care of them, operated on them, se- 
cured, fixed, sectioned the eggs, stained and photographed the sections myself. 
The source of hght for photomicrophotography was a home-made illuminator: a 
bull's-eye lens set into a tin can which slipped easily into a slightly larger can for 
focusing, the light bulb being in the upper can. I have a little essay in the bottom 
of my barrel entitled "In Praise of Poverty." I realize, of course, that the poverty 
situation can be overdone, but I wonder if your young graduate students don't 
have too much done for them at the large, well-equipped centers. After all, they 
will likely get jobs at small colleges where they will be more or less on their own. 

As our method of procedure in collecting embryonic material required surgery, 
a score of pre-medical students volunteered their help, working far into the night 
during the height of the collecting season. There was no luxurious operating 
room available; we worked in the physiology laboratory when it was not other- 
wise in use. Thinking back on the enthusiasm which our young helpers exhibited 
40 years ago, I would suggest that research laboratories be not far removed from 
the teaching laboratories. It is good even for Freshmen to sense that there is some- 
thing new, even mysterious, going on around them. They will snoop to get an 
idea of what it is and if the researching professor will stop and explain the matter 
he may be making research scientists out of some of the choice pupils under his 
influence. 

When Pat assigned me to develop physiology in the department I again felt 
like that "one-eyed king among the blind," since I knew only too little about the 
subject. Using my book royalties I spent four summer quarters at the University 
of Chicago, studying physiology under the great A. J. Carlson, neurology under 
two masters, Herrick and CoghilL and biochemistry under Fred Koch. These 
studies were supplemented at Columbia University in the summer of 1917 by 



22 The University of Texas Publication 

clinical pathology and clinical bacteriology, the latter under the famous Hans 
Zinsser. A year's leave of absence (1917-1918) was spent at the Wistar Institute 
and the University of Pennsylvania, which enabled me to make rapid progress 
in working up the rich collection of eggs so that I could complete a monograph on 
the subject in 1919. 

In the ten years following 1915 I published several papers on the early embry- 
ology of the opossum, passing gradually to a consideration of the physiology of 
reproduction. Later I studied the embryology of the monkey but again I 
relegated the embryology to second place, preferring to pursue physiological 
problems. 

To the legitimate question: of what use are those opossum studies, I have a 
ready answer. Not, however, that their possible applicability ever constituted a 
motive on my part. Knowledge of physiological processes carries over from one 
species to another, including man. Some specific examples may be given. ( 1 ) The 
newborn opossum, a mere embryo, has been used by Burns to solve problems of 
sex reversal and the genesis of hermaphroditism. (2) The blood of the newborn 
opossum contains globulin but no plasma cells, the supposed origin of globulin 
according to the books. There are likewise no small lymphocytes, the supposed 
origin of fibroblasts in inflammation; inflammation can be produced in the new- 
born opossum, however. Therefore, according to Block, hematological theory may 
require basic revision. (3) It was on the opossum material that Painter dis- 
covered the X-Y condition in spermatogenesis, later proving this for man also. 
(4) The opossum taught me two basic principles, now generally accepted, which 
have had a great influence on gynecological thought: I clinched the principle of 
the short functional life of the unfertilized egg, and the unexpectedly rapid ascent 
of the spermatozoa in the female genital tract. I had to demonstrate the latter by 
rather spectacular experiments in the rat; now, after 30 years, the subject is 
receiving serious attention from gynecologists as I have shown in my recent 
paper, ''How do sperms get into the uterus?'' (1957. Fert. & Steril. 8:403-437). 

These examples serve to illustrate the fact that basic studies sooner or later 
find a practical application. I never bothered to ask: is there any use in this 
study? The main thing was that I was satisfying my own curiosity and having 
fun doing the experiments. 

The work on the opossum proved to be a preliminary training for my studies 
on the monkey, for in 1925 I wrote up, with Pat's blessing, a prospectus for an 
expedition to the Philippine Islands to gather monkey embryos since I felt that 
such a study would constitute a necessary preliminary to the securing of com- 
parable human stages. With the aid of Arthur LeFevre, formerly a Professor of 
Latin at the University of Texas and an intimate friend of Will and Mike Hogg, 
members of the board of the Texas Oil Company, these loyal University alumni 
were induced to provide the larger portion of the expenses for the proposed two- 
year "Texas" expedition. The National Research Council also made a grant to 
the amount of five digits. I sent my proposal to Dr. George Street er, Director of 
the Department of Embryology of the Carnegie Institution of Washington, 
whose laboratory was a part of the Johns Hopkins Medical School in Baltimore. 
Dr. Streeter's reaction was an invitation to me to join his laboratory and study 
monkey embryology the "safe and sane way," bringing the monkeys to the labo- 



Hartman: The Making of a Scientist -^ 

ratory and thereby avoiding the dangers of tropical infections for myself and 
family. George Corner had already demonstrated that the Rhesus monkey made 
a good laboratory animal once one learned how to house and handle them. So the 
"Texas" expedition, and my trip around the world, went by the board and m 
1925 I resigned and joined the Carnegie laboratory. Sixteen years of research m 
primate embryology and reproductive physiology ended after I had secured over 
a hundred monkey eggs and embryos, completing the story of the first thirty days 
of monkey development. This was published in a de luxe edition in the Contribu- 
tions to Embryology by Heuser and Streeter, and in which my name was omitted 
from the title by the editor, perhaps unwittingly. I was myself more concerned 
with the physiology of the female monkey, contributing my share to the unravel- 
ing of the cause of menstruation and the sequence of events in the menstrual 
cycle, especially the time of ovulation. These studies, supplementing those of 
P. E.' Smith, George Corner. George Bartelmez and others have had a profound 
effect on gynecological thought the world over. 

As I saw my obhgatory retirement from the Carnegie Institution approachmg, 
I decided to accept a six-year appointment as head of the Departments of Zoology 
and Physiology in the University of Illinois. These were the war years. Again I 
retired, and this time, having a broad knowledge of reproductive physiology, I 
accepted a position with the Ortho Research Foundation at Raritan, New Jersey, 
becoming Director Emeritus in 1958. 

The Department of Zoology of the University of Texas, under Pat's versatile 
direction, has of late years become a renowned center for research, with emphasis 
on genetics. Pat's interest in genetics began many years ago with his modest but 
thorough course in the subject (and which my wife Eva took, and made an A!). 
Genetics was given a boost by his bringing in T. S. Painter and H. J. Muller. I 
took Muller's lectures myself and, incidentally now that he has attained the 
Nobel Prize, I am not a little puffed up by recalling that I urged him to lose no 
time in studying the effects of x-rays on the mutation rate of Drosophila. To my 
mind Mavor, at Union College, had at least demonstrated that something signifi- 
cant lay in that direction. Muller's flies were radiated by my friend, Dr. Richard- 
son, an accommodating local radiologist with a feeling for basic research. This 
proved a lucky break for Muller and for the Department, for it opened up a new 
approach to Genetics. I am happy to have had a small part in the first successful 

study. 

In view of Pat's consuming interest in the genetics, physiology, and ecology of 
Drosophila one is apt to forget his contributions to embryology, his first interest. 
In my day I always thought that two basic courses of the department were the 
most outstanding: beginning Zoology, conducted by the fine scholar and gentle- 
man, Dana Casteel, and the course in embryology given by Pat. Each student in 
the embryology class was equipped with both compound microscope and the 
newly invented and improved binocular stereomagnifier, the former for studying 
the slides, the latter to aid in the dissection, with needles and fine-pointed forceps, 
of the 12 mm. pig embryo. Pat always maintained a high standard for his stu- 
dents. There was no pampering; his recommendations for admittance to medical 
schools meant a lot to the Deans of the top schools of the country. 

To my young friends who happen to read this condensed sketch of nearly 60 



24 



The University of Texas Publication 



years of teaching and research I would say that hard work and perseverance 
always bring results. Consistent prosecution of well-planned researches may, 
with meditation, lead to generalizations which are the key to progress in science. 
I used the following quotation as my motto in my 1932 monograph on the repro- 
duction of the Rhesus monkey: "It is certainly not the obligation of science to 
explain everything. We must collect facts. Once facts have been assembled, the 
explanation emerges of itself." It doesn't often happen that a natural law looms 
up in the serendipity of the scientist, but it can happen to any consistent re- 
searcher who stays on the job. Certainly it does not happen without a back- 
ground of a multitude of personal observations. To acquire such a background 
takes hard work, often long hours, the productive use of long academic vacations, 
and "guts." The IQ of a genius is, fortunately for most of us, not essential to 
making substantial contributions to knowledge and the satisfaction of having 
made them. 

I do not recommend my own devious course toward a scientific career; instead 
I recommend acting upon the spirit of the saying, "The world stands aside for 
the man who knows where he is going." It is best to make up one's mind when 
young and get going toward a goal — not omitting those amenities of the well- 
rounded life that make for a whole, completely social person which, I may add, 
describes almost every professor in my rather wide acquaintanceship. 




Left to right: H. J. Muller, the author, G. H. Parker, and J. T. Patterson. Taken on the occasion 
of Dr. Parker's visit to Austin about 1923. 



High Islands and Low' 

WARREN p. SPENCER 
College of Wooster, Wooster, Ohio 

This evening I am inviting you to travel westward, far westward, with me as a 
member of Task Force 7, under the auspices of the Atomic Energy Commission, 
to the Pacific Proving Grounds in the Marshall Islands and beyond. Our trip, 
and the special work we are doing, will take us at times far from the Proving 
Ground We will not discuss the work, but by no interpretation of our orders is i 
forbidden that we reminisce about what we saw in the Southern Marshalls and 
the Eastern Carolines, now under the United States, administered through the 
Department of the Interior (I should think that this phase of operations should 
be designated the Department of the Exterior) as a part of the United Nations 

Trust Territory 

We are at the Hopkins Airport in Cleveland on a late July morning. 1956. We 
are scheduled to take this trip with three scientists from the Genetics Foundation 
at the University of Texas, and so our seeming diversion to the Lone Star State 
rnakes some sense. 

Fasten your seat belts. No smoking. tt i • 

We will see that sign flash forty times before we set down again at Hopkins 
Airport on September 4th. Cleveland, Chicago, Dallas, Austin, San Francisco 
commercial airport, Travis Air Force Base near San Francisco, Honolulu, Kwaja- 
lein Eniwetok, Bikini Atoll Eniwetok, Majuro Atoll, Kwa]alem, Emwetok. 
Ponape in the Carolines, Eniwetok, Kwajalein, Honolulu, Travis Air Force 
Base San Francisco, Chicago, Cleveland— forty take-offs and landings. You 
only counted thirty-eight. But you forgot those two runs on the Hickam air- 
strip in Honolulu several hours apart and then the orders back to bed and up 
the next morning for another try. No wonder MATS (Military Air Transport 
Service) has such a fantastic safety record. If all four of the big engmes aren t 
just right you don't take off for Kwajalein over 2300 miles of water until they are. 
Of course we haven't counted the trips island hopping in a hehcopter, where you 
make a round-trip tour of the atoll and land on perhaps half a dozen islands 
between 9: 00 a.m. and noon. 

We will travel on the two-motored planes of Braniff , and the big four-motored 
transcontinental planes with good-looking hostesses and good food, on the big 
MATS planes across the Pacific, each one capable of carrying sixty tons of cargo 
but now carrying mostly military personnel, sergeants instead of pretty host- 
esses cold box lunches instead of hot meals, and hquid ration of coffee and water 
only' We will fly to Ponape by sea-plane and land on the lagoon, and take off 
with JATO (rockets which blast the plane off the water) . And in the islands we'll 
travel by fast water-taxi from one small island to the next; by EST, an ocean- 
going vessel over a hundred yards long, without a keel and capable of landing 
at an open beach, between Bikini, Rongelap, Rongerik and Eniwetok Atolls, a 

1 This is a modified version of a talk delivered to the Century Club of Wooster, Ohio, m 1956. 



26 The University of Texas Publication 

sea cruise of a few hundred miles; by Duck (an amphibious craft) off the LST 
to certain islands and over the islands in this strange mechanical monster, equally 
at home powering its way over the lagoon, rolling up the beach between coral 
heads, or plowing through the jungle, wreaking havoc on unsuspecting palm 
trees in its path. We will sometimes go by LCM boat, generally referred to as an 
M-boat, several miles from one island of the atoll to another. And if you wish 
you may take a trip in an outrigger as some of my friends did, and as I regretted 
not doing. 

On land we will be travelling by truck, by jeep, or on foot, but not likely by 
car. By these several and sundry means we'll travel some 20,000 miles in less 
than six weeks. But more significantly, perhaps, we might describe it as "flying 
through history." For we will have flown from the great centers of western 
civilization to the sites of ancient primitive cultures, and quite possibly we will 
have visited the symboHc center of the final quotation mark after that human 
autobiography which we call history. 

In 1846 Wiley and Putnam published Herman Melville's Typee, with the sub- 
title "A Peep at Polynesian Life during a Four Month's Residence in a Valley 
of the Marquesas." This served as a sizable part of my literary fare on the trip 
out from San Francisco to Honolulu and on to Kwajalein in the Marshalls. I 
recommend it as required reading for anyone making a trip to the Pacific Islands. 
Over a hundred years later it can still serve as a guide to much of what will be 
seen in native villages today, and of course it has the style and charm of the 
master craftsman. 

The first paragraph of Typee presents a soliloquy on the hardships and rigors 
of a six-months cruise on a whaling vessel to reach the Marquesas Islands and 
ends in these words: "Oh! ye state-room sailors, who make so much ado about a 
fourteen days' passage across the Atlantic; who so pathetically relate the priva- 
tions and hardships of the sea, where, after a day of breakfasting, lunching, din- 
ing off five courses, chatting, playing whist, and drinking champagne punch, it 
was your hard lot to be shut up in little cabinets of mahogany and maple, and 
sleep for ten hours, with nothing to disturb you but 'those good-for-nothing tars, 
shouting and tramping overhead,' — what would ye say to our six months out of 
sight of land?" 

As I read that paragraph I couldn't help wondering what Melville would have 
thought and written of those air travelers of today who grumble about slight 
inconveniences as they travel in hours literally the distances covered by those 
mid-nineteenth century travelers in days. We left Kwajalein on Sunday evening, 
September 2nd, at 6:30 p.m. Cleveland time and were in Cleveland Tuesday 
morning, September 4th, at 10:00 a.m., a distance of approximately 7500 miles, 
and this included several hours on the ground in Honolulu, a seventy-mile taxi 
ride between the two airports in the environs of San Francisco with several hours 
wait there, and a couple of hours in Chicago. And again Melville came to mind 
when, at the Officers Club on Eniwetok, the evening before we left for home, an 
air force captain told of recently flying a jet the 2500 miles to Honolulu in less 
than four hours. More than once I thought of Melville and his valley of Typee in 
the Marquesas Islands of 1850 during our visits to the Kapingamarangi Village 
on the island of Ponape in the Eastern Carolines in the summer of 1956. Much 



Spencer: High Islands and Low 



27 



that we saw there could have come right out of the pages of Me -he and fo 
that matter Typee might just as well have beer, wrxtten t'^Captam Cook m the 
late 18th century or by some bold manner had he chanced upon the islands a 

thousand years earlier. , , i 

Between latitudes 30 North and 30 South, the eastern ^wo-ftfthso the broad 

Pacific IS almost devoid of islands except for the G^^^P^^f ^ 5°"^ f *;^°' *;„ 
northern South America. But a look at a good map will show that the western 
three-fifths of the Pacific between these latitudes is liberally sprinkled wi h 
thousands of islands. These may be roughly divided by two lines runnmg north- 
west-southeast into three great island groups, an eastern Polynesian group, a 
middle Micronesian group, and a western Melanesian group. ^. . , ,, 

Polynesia includes the Hawaiian Islands, the Line Islands including Christma 
Island directly to the south of Hawaii, the Marquesas, the Society Islands ot 
which the most famous is Tahiti, and the Tuamotu group lying between the 
Marquesas and Society Islands but stretching further to the south and east to 

Melanesia forms a chain of island groups in order from northwest to southeast: 
New Guinea and New Britain, the Solomons, New Hebrides, and the Biiis. 

Micronesia (Fig. 1) lies roughly between Melanesia and Polynesia stretching 
from approximately latitude 20 North, longitude 140 West to latitude 10 South 



ENIWETOK ATOLL 



BIKINI ATOLL rqNGELAP 



^JL: 



ATOLL RONGERIK ATOLL 






^'-1. 



\ 



\. 



<l MAJURO 

ATOLL ^-^^ 



» .i\PONAPE 



<£3l 



I70° 



on the international date line, that is from northwest to southeast. While most 
of the islands of Polynesia and Melanesia are small the Micronesian islands are 
still smaller. To the north lie the Marianas, a north-south chain about six 
hundred miles in length and including Guam, Rota and Saipan. South of the 
Marianas are the Carohnes, stretching for two thousand miles m an east-west 
direction. In the Palau group of the western Carolines is found Babelthuap 
Island approximately 150 square miles in area and the largest of the Micro- 



28 The University of Texas Publication 

nesian islands (but less than a fourth the size of Wayne County, Ohio). Yap is 
another of the larger islands in the western Carolines. In the eastern Carohnes 
the largest islands are Truk, Kusaie and Ponape. the latter being the second 
largest island in Micronesia with an area of approximately 100 square miles. A 
few hundred miles to the east He the Marshalls, in about the same latitude. South 
and a httle east of the Marshalls are the Gilberts, close to the equator, and still 
further south and east lie the Ellice Islands. 

In his ''The Fortress Islands of the Pacific'' published in 1945 Wilham H. 
Hobbs gives an intriguing account of the geology and geography of the islands 
of Pacific Oceania and of their value as military bases. In brief his thesis is as 
follows. The islands of western Oceania, Melanesia, the Aleutians, the Marian- 
as, and the western Carolines are what he terms arcuate islands formed by cres- 
centic folds and upthrusts from the floor of the Pacific. Such folds form moun- 
tain ranges rising from the Pacific floor with their peaks forming a chain of 
islands. The folds tend to be steep on their eastern face and with gentle slopes on 
the western face. Eastward of each fold lies a deep trough in the sea. Thus the 
north-south chain of the Marianas may be taken as an example with the Nero 
Deep just to the east almost six miles in depth, the second deepest sea trough in 
the world. East of these arcuate chains lie what he terms the strewn islands, the 
islands of most of Micronesia and all of Polynesia. These are not mountain 
chains, but rather have been formed by groups of volcanoes at one time active on 
the sea floor, eventually rising above the surface. Some of these have become 
inactive, eroded, and formed atolls, rings of coral island motus or islets from the 
reef originally surrounding the volcano, now eroded and submerged. He recog- 
nizes several types of island groups forming transitions between these stages, 
the volcanic island, with its surrounding coral reefs; the almost-atoll, where parts 
of the crater still stick above the surface in the lagoon surrounded by coral reef 
and developing coral islands; the typical atoll, with coral reef and islets sur- 
rounding a lagoon, and still later stages in island evolution, partly raised atolls 
with shallow lagoon, and raised atolls with the lagoon repaced by flat-topped 
phosphate rock. It will be seen that Hobbs accepts the Darwin-Dana theory of 
the origin of atolls. 

However they were formed geologically it would be obvious to even the most 
casual observer traveling by boat or plane through Polynesia and western 
Micronesia that the islands are of two very distinct types, a relatively few 
larger mountainous islands and a myriad of tiny islets, the latter typically ar- 
ranged in circles, quadrangles, or less even geometric figures, but generally each 
island forming a long narrow strip, obviously arranged like long narrow beads on 
an encircling coral reef surrounding a lagoon. This is what we mean by high 
islands and low. 

In 1896 Robert Louis Stevenson published a treatise entitled ''In the South 
Seas'' and here he describes vividly this contrast as follows: 

No distinction is so continually dwelt upon in South Sea talk as that 
between the "low" and the "high" island, and there is none more 
broadly marked in nature. . . . On the one hand, and chiefly in 
groups of from eight to a dozen, volcanic islands rise above the sea. . . . 



29 



Spencer: High Islands and Low 

their tops are often obscured in cloud, they are all clothed with various 
forests, all abound in food, and are all remarkable for picturesque and 
solemn scenery. On the other hand, we have the atoll; . . . rudely 
annular in shape; rarely extending beyond a quarter of a mile at its 
chief width; often rising at its highest point to less than the stature of a 
man— man himself, the rat and the land crab, its chief inhabitants; not 
more variously supplied with plants; and offering to the eye, even when 
perfect, only a ring of glittering beach and verdant foliage, enclosing 
and enclosed by the blue sea. 

The Hawaiian Islands, which some of you have visited, are of course "high 
islands" of Polynesia. Our party spent several days on Oahu in the summers of 
'55 and '56 Through the hospitality of members of the Zoology faculty at 
the University and of Dr. Mitchell of the U. S. D. A. we saw a good deal of the 
island downtown Honolulu, Waikiki Beach and its fine hotels with their 
gardens, and curio shops. We drove out past Pearl Harbor and Scofield Barracks, 
past sugar cane and pineapple plantations, over the Pali drive up through the 
mountains and along the beach past Diamond Head. We made a tour of the 
University and of the famous Bishop Museum, and took a hike on a mountain 
trail along the shoulder of Mt. Tantalus far above the city of Honolulu. My most 
vivid impressions of Oahu: the air-conditioned climate, almost every home sur- 
rounded by gardens with a rainbow array of flowering trees, shrubs and herbs 
including orchids; steep serrated cliffs in the mountains, not bare rock but 
covered by dark green verdure; the polyglot of racial types seen on the streets 
and all talking good mid- Western American; the irony of Hawaii not having 
been given statehood long ago. But Hawaii would serve as subject for several 
papers. Let us go for examples of low and high islands to the Marshalls and 
Eastern Carolines 2500 miles southwest of Honolulu. 

We shall take the Marshalls in general as examples of low islands and the 
Majuro and Bongelap atolls in these islands for more specific reference as both 
lie outside the Pacific Proving Grounds test area, the former in the southern 
Marshalls and the latter in the northern Marshalls. We shall visit the island of 
Ponape in the Eastern Carolines as an example of a high island. 

In ''Fortress Islands of the Pacific'' Hobbs introduces the Marshalls as follows: 
"This double chain of atolls Kes to the northwest of the Gilberts and is separated 
from them by three degrees of latitude. Like the Gilberts, the Marshalls trend 
northwestward with their eastern chain known as the Badak (Sunrise) chain, 
and the western as the Balik (Sunset) chain. These chains contain thirty-six 
larger atolls with 876 motus (islets) and reefs which stretch from Ebon at the 
south in latitude 4 degrees, 34 minutes to Eniwetok at the extreme northwest 
in latitude 1 1 degrees, 30 minutes North." 

These almost nine hundred islets have a combined area of a httle less than 70 
square miles, approximately one-eighth the area of Wayne County. Kwajalein 
atoll about midway in the western chain is the largest in land area with 6.33 
square miles. It also has the largest lagoon area, about sixty miles long and with 
some 90 motus or islets. It is in fact one of the largest atolls known. We visited 
Eniwetok, Bikini, Bongelap and Bongerik atolls in the northern Marshalls and 



30 



The University of Texas Publication 



lying m the western chain, Kwajalein (about midway between northern and 
southern Marshalls), and Majuro in the southern Marshalls and in the eastern 
chain. We were on approximately twenty different motus or islets in these atolls. 
What does an atoll look like? This depends on the viewpoint, whether one is 
flying, say, at an altitude of 10,000 feet above it, coming in low for a landing or 
flying a few hundred feet up in a helicopter, approaching it by sea in an LST, 
riding over the lagoon in a small craft, or actually walking along the beach of a 
motu on the lagoon side or on the sea-side of the islet. This is what a typical atoll 
looked like to me from the window of a MATS plane flying at an elevation of 
9,000 feet. It was as though a modern artist had taken a giant canvas and first 
laid down a background of dark blue with tiny ripple marks to represent the 
waves of the sea. Next he had taken a brush, dipped it in light brown paint and 
traced a rather regular quadrangular line. This line was the coral reef, but 
submerged. Next he had traced a white line just outside the brown, the breakers 
in the shallows off the reef. Outside the white line he had laid down a broad light 
green strip merging gradually into the dark blue, the shallow water on the sea- 
ward side of the reef. Finally he had daubed dark green splotches, some forty or 
fifty of them, some tiny circles, but mostly long strips of green and irregularly 
spaced but always directly on the light brown line of the reef. These were the is- 
lets covered with vegetation, mostly cocoanut plams, but these indistinguishable 
as such at that height. At the corners of the quadrangle the green splotches were 
somewhat larger and roughly shaped like a boomerang. For some reason the islets 
at the angles of a reef are generally, though not invariably, larger. One more 
Item: a few daubs of dark brown paint interspersed with the green blotches along 
the light brown line; tiny islets sticking above the surface of the water, but with 
little or no vegetation, probably covered at high tide. Such is an atoll as I saw it 
from 9000 feet in the air and it ought to take a prize at a showing of modern 
art. But the canvas is large; the average atoll will be twenty or thirty miles from 
one end of the lagoon to the other. And on some of the atolls the light brown line 
will be interrupted at one or more points, a deep water passage into the lagoon, 
and in other atolls the islands will mostly be in a semi-circle on the eastward 
side of the lagoon. But I chose to describe a particular atoll which we flew over 
and I don't even know its name, or whether it had any. 

From the beach on the lagoon side of an islet one can see adjacent islets in the 
circle both to the right and the left, but those across the lagoon twenty or thirty 
miles away are invisible as the islands themselves have a maximum elevation of 
less than fifteen feet generally, and the cocoanut palms are not likely to be more 
than forty or at most fifty feet tall. We shall return to the atoll islands for closer 
inspection later, but for contrast let's view a typical high island from the air. 

On August 22nd our party of nine boarded an SA-16 Aircraft on the Eniwetok 
air strip, with Captain Stahley and crew of three in charge, bound for Ponape 
some seven hundred miles to the southwest in the Eastern Carolines. This was a 
sea-plane and several hours later we came in for a landing on the lagoon between 
the reefs and the main island. The view of Ponape as we came in for the landing 
(if that is what a sea-plane hitting the water is called) is one which I shall not 
forget. I'm not kidding myself; it will get fuzzy around the edges, but the main 
show will be there years hence along with my first view of Naples Harbor, the 



Spencer: High Islands and Low 



31 



Grand Canyon from the South Rim, the sweep of the Teton range m Wyommg, 
the unearthly shimmer of Death Valley in the late spring, the colormg of Bryce 
Canyon, the majesty of Yosemite valley, and the fall coloring on a valley road 
in Wayne or Holmes County in a good year. These are things you can't take 
from me and on which I pay no income tax. If I did I'd be broke. 

What is it that makes a view worthwhile? I think it is partly a matter of con- 
trast. If the trees here were always colored as they are in October we would 
probably all be color blind. And physical conditions may help. For one thmg 
we had good windows for viewing in that navy plane, which is more than can be 
said for the tiny windows spaced well apart in the big MATS planes. Then on 
those two-engined planes used in flying in the islands we always had to wear a 
"May West" on take off and landing, and the landing sign flashed long before 
the landing. The May West is a horse collar affair weighing too much for com- 
fort. Mae must have been a buxom lady in her day. But the navy plane was out- 
fitted with a much more modest affair, worn the same way but better described 
as a Katherine Hepburn or a Grace Kelley, and so one could observe the scenery 
in some comfort. Then after looking at all those tiny atoll islets and tramping 
around over many of them I could hardly believe that out there in the wide 
Pacific, in that region at least, there could be anything like Ponape, a real chunk 
of land where you could undoubtedly get lost and lose the impression that there 
was a sea around. Without piling on the adjectives I'll just say soberly that my 
first ghmpse of Ponape struck me as something out of this world or more properly 
something out of this sea. There it was and I could hardly believe it. You may 
smile when I tell you that it was an island only about thirteen miles m greatest 
diameter, but compared to those islets where you stand in the middle and see 
the sea a few hundred feet on one side and the lagoon a few hundred feet on the 
other side, it was something to wonder about. Perhaps it was a little of the same 
kind of exahation which came to the ancient mariner on sighting land after a few 
months on the open sea. 

As the plane came in from the north over Langor Harbor, circled and lost 
altitude one saw the main mass of the island, with mountain ranges, rather 
rugged, some of the peaks rising to around 2500 feet, dark green with luxurious 
tropical vegetation, this broken in spots with lighter green patches which as the 
plane approached closer were seen to be clumps of palms on the mountain sides, 
the tops of the peaks hidden in masses of white and gray rain clouds, deep valleys 
cutting in between mountain ranges, the shore line deeply cut by estuaries where 
the valleys and their streams came down to meet the sea; and on the west shore 
of the largest estuary the settlement of Colonia, with its metal roofs glinting m 
the brilliant sun, more of a town than we had seen since leaving Honolulu (I 
don't count military installations, however extensive, as towns). Encircling the 
main island a few miles off shore were the coral reefs, with their usual garnish- 
ings of white surf and Hght green shallows. Between the reef and the main island 
were several small islands, small repHcas of the main islands, rugged, not flat, 
though of course the hills were also in miniature. On one of these was the landing 
ramp and concrete strip for the plane to taxi up and come to rest. To the west of 
the harbor a huge mass of rock with a sheer face jutted out of the sea. A few tiny 
craft could be seen on the surface of the harbor. This is a poor description of a 



32 The University of Texas Publication 

sight of surpassing beauty to a landlubber far from his native environment, mid- 
western U. S. A. A few moments later after careful reconnoitering (it is not 
healthy to set a sea plane down on a submerged coral head) the pilot feathered the 
engines and the plane settled down in the water at perhaps forty or fifty miles 
an hour, with the pontoon on the right wing swishing along in the water and that 
on the left wing a few feet above the surface, water swishing up against the 
sealed windows, and the plane rapidly losing speed. As we approached the 
landing ramp, a gentle concrete incline, the landing wheels were let down out of 
the sides of the plane and we taxied up and along a concrete strip, turned and 
came to a halt. We debarked and deposited our luggage in the picket boat which 
serves also as a crash boat for the sea-planes. While waiting the arrival of a 
Transoceania plane, flying weekly the rounds from Guam to Truk to Ponape to 
Majuro and back, we were greeted by friendly officialdom with a welcome to 
Ponape. In a half hour or so the other plane had arrived and with its passengers 
we took the half-hour ride across the lagoon to the dock at Colonia on Ponape. 
After a short wait on the dock, surrounded by a large group of natives, most bare- 
foot and in rather tattered garb, apparently on hand to see the weekly arrivals 
on the plane, we were transported (or should I say carted) in very ancient jeeps, 
with springs long since broken out of the seats, over rather a rough street, yes, 
a very rough street, up to the crew's quarters, a quonset hut with open sides 
below and thatching above, where we were to stay for the next eight days. The 
accomodations were adequate, semi-private rooms, flush toilet, hot shower, 
electricity and a water cooler which ran only lukewarm water. On the way, the 
galley, where we were to take our meals, was pointed out, the fare adequate 
though somewhat monotonous with rice served as a base every day twice a day 
without fail. But there were other dishes including some native stuff such as 
yams, breadfruit, and baked bananas. 

Perhaps before we start the exploration of the town of Colonia and adjacent 
regions of Ponape Island it would be well to lay a very brief historical back- 
ground. Most of the information was gleaned from a pamphlet issued to all 
visitors and prepared by Henry Hedges, District Administrator of the Ponape 
District of the Trust Territory of the Pacific Islands, and in conversation with 
the administrator, his wife and other officials. 

Since the second world war the United States has acquired the administration 
of a Trust Territory under the United Nations which includes the Marshalls, 
Carolines, and Marianas. This territory is administered by the Department of 
the Interior with an annual operating budget of $5,000,000 divided about evenly 
among the six districts. These six districts are: the Marshall Islands, the Mari- 
anas, and four in the Carolines, namely, Ponape and Truk in the Eastern Caro- 
lines and Yap and Palau in the Western Carolines. There are approximately 
60,000 natives in the territory, about 12,000 in the Ponape District and 11,000 
in the Marshalls. There is a general administrator of the Trust Territory and 
under him district administrators for each of the six districts. Each administrator 
has associated with him a group of twenty to thirty Americans, most of whom 
have their wives and families in residence on the site of operations. The per- 
sonnel on Ponape is distributed among the following departments: administra- 
tion, education, public health, agriculture, construction and maintenance, land 



Spencer: High Islands and Low 33 

and claims office, weather bureau, and special projects, thirty-four members 
in all but that includes some of the wives. Much the same organization is to be 
found at the headquarters for the Marshall Islands District on Majuro Atoll. 
These people are paid comfortable salaries, of course scaled according to the po- 
sition; they sign up for a two-year period with three months furlough in the 
States, travel expenses paid. On Ponape, where most of the personnel are young 
people in their early thirties the turn-over in personnel has been high, due 
among other things to the fact that some wives simply could not take the re- 
strictions of hfe as it must be lived in such a place. There are inconveniences if 
not actual hardships. The average age of the personnel on the Marshalls was 
considerably older, and more of the people there were adapted through long ex- 
perience to life on a tropical Pacific island. Of course there are many natives 
hired as helpers in the project, some few of them in responsible positions. Thus 
the administrator of education in the Marshalls is a native trained at the Univer- 
sity of Hawaii in Honolulu. It is the business of the Trust Territory personnel to 
work with and for the natives on these islands to improve their health, to educate 
them, to supply them with a decent standard of living through improvements in 
agriculture and housing, and to help them in estabhshing a democratic and 
equitable form of self government. 

I was favorably impressed by the caliber of the men and women in this work. 
Of course there are a few inevitable exceptions and at the other end of the scale 
people who seem to stand out in their positions. I shall mention two at this point. 
I was told by my colleague. Dr. Marshall Wheeler, of the individual who flew 
with him and several others down to the Kapingamarangi Atoll to handle some 
legal matters. I shall call him La Feete (that was not his name). He carried a 
bottle with him and was in a drunken stupor on the plane. During the negoti- 
ations with the native chief and his aides he had his bottle out on the counsel table 
and would occasionally take a swig from it during the negotiations. On the trunk 
of a palm tree growing near the door of the galley where we ate on Ponape was 
this inscription carved by some discerning individual: "La Feete, Jerk, First 

Class." 

In contrast I would like to mention Mr. Byron Bender, assistant administrator 
of education for the Marshall Islands District. Bender is a graduate of Goshen 
College in Indiana, and has done graduate work in Anthropology. When he heard 
that I was from Wooster the evening of our get-together at the Club on Majuro. 
he asked me if I knew Howard Yoder. I assured him that I did and that we both 
belonged to this august organization. It seems that Bender roomed with Howard's 
son in Goshen and has been a guest in his home here in Wooster. After the fes- 
tivities at the Club, Wheeler and I went over to the Benders' home on Majuro 
for a midnight cup of coffee and some cake. With his young wife and two daugh- 
ters, they have spent most of their married life, several years, in Majuro working 
in the educational program. Their home is half of a Quonset hut, well supplied 
with books, quite comfortable in so far as comfort can be obtained in that tropical, 
humid climate, their children delivered by native Marshallese doctors, their 
hearts and energy in the work they are doing. I liked young Bender, his enthusi- 
asm for the work he was doing, his dedication to it, his wisdom for one so young. 
I happened to overhear a conversation the next day at table between Mr. May- 



34 The University of Texas Publication 

nard Neas, the district administrator, and some of the A. E. C. men who had 
flown down with us. He was speaking in highest terms of Bender's work among 
the natives. On his own initiative he has set up a hand-printing shop and is com- 
posing and printing textbooks for the schools. He is to be in charge of taking the 
Rongelap natives back to their home Atoll and reestablishing them there this fall. 
The Rongelap natives were evacuated from their home atoll in the summer of 
1954, after they had received radiation from the fall-out in the big blast that 
summer. They have in the interim been on Majuro Atoll, and rather unhappy 
there. To me, one atoll looks as God-forsaken as another, but to native Marshall- 
ese whose ancestors have lived for many generations on that 3.07 square miles 
of palm and pandanus bedecked coral strand I suppose the adage "Be it ever so 
humble, there's no place like home" has a meaning. At any rate Bender has the 
job, among many others, of taking those people back home to the very adequate 
housing, western style at their request, which will take the place of the beautiful 
pandanus huts which they formerly occupied; yes, of course the U. S. foots the 
bill, but could they do less for a group of natives burned out in more senses than 
one. Needless to say the tail end of the evening which Wheeler and I spent in the 
Bender home was a fitting climax to what had gone before. 

To be successful in the work they are doing, and in my opinion most of those 
whom we met were eminently successful, these people must have a sense of 
mission. They must be and they are dedicated to the job they are doing. They 
must be able to take it. By that I mean to live a life quite different from that 
which we live in the states; work in a climate with no change of seasons (the 
only difference on Ponape is that there is a wet season and a wetter season with 
an annual rainfall of around two hundred inches), a debilitating climate where 
the day temperatures are in the eighties and the humidity close to 100%. An 
outsider wonders what is to be gained by foisting western civilization onto a 
people whose culture seemed well adjusted to the environment. The U. N. work- 
ers see those problems even more clearly. This I know from very rewarding 
conversations with several of the personnel on Majuro and Ponape. There is 
also the constant frustration of wondering how long this project will be con- 
tinued. They know full well, and they're not kidding themselves, that the U. S. 
is over there for one major purpose. These islands constitute what Hobbs has 
called Fortress Islands of the Pacific. We're in there because the islands give us 
potential military and naval bases, and who knows, with the evolution of up-to- 
date means of war how long such islands will remain of importance. The Trust 
Territory people must act and build as though this is a long term project, but 
none realize better than they that the project may fold up tomorrow. And the 
budget, compared to the billions of dollars spent at the Pacific Proving Grounds for 
military purposes is certainly puny. There are so many things that could be done 
on Ponape if they only had the money and personnel to do them. But I heard no 
complaints from the Trust Territory people on this score. I only heard this point 
very strongly put by Tom Hardison of the A. E. C. Many others in the A. E. C. 
feel the same way. It is heartening to see how some of these men go out of their 
way to see that the district administrators and others get the scraps from the 
military table, scraps which would rot on coral beaches otherwise, and it is 
heartening to see what these administrators can do with such scraps. I can only 



Spencer: High Islands and Low 35 

hope that this Trust Territory project may be continued for many years, for it 
is my opinion that the goodwill invoked from the wise administration of such a 
microcosm experiment may well furnish a defending bulwark for democracy al 
out of proportion to the money invested. This, of course, is one man's opinion, and 
the opinion of one who feels strongly that the cold war will eventually be won 
by a constructive attack on the problems of providing a decent chance to live the 
good life for every human being on the planet. Incidentally this will involve an 
honest consideration of the greatest problem which faces us today, the problem of 
the limitation of world population. 

Of course the peoples of the Marshalls and CaroHnes have been subject to 
foreign influence and domination for a period long antedating the second world 
war, and it is against this background that the Trust Territory personnel must 
work today, knowing full well that what they do will be compared with what 
went before. Between the first and second world wars the Japanese held a man- 
date over the Marianas, Marshalls and Carolines. To this island empire, already 
overpopulated and feeling the need for ''lebensraum;' the mandated territory 
offered a new potential for immigration, and the estabhshment of industries and 
agriculture to supplement the precarious economy of the home islands. During 
this period there were over 3000 foreigners, mostly Japanese, living in Colonia, 
the principal town in the north of Ponape. On the other side of the island m 
Matalanim (Madolenihm) province there was a settlement of over 2000 Jap- 
anese and Okinawans. There were more Japanese living on the island than the 
native population of some 8000. The Japanese developed commercial farming 
in the interior valleys, introducing the culture of rice and sugar cane. They 
estabhshed industries. To quote from Raymond E. Murphy in an article on 
''High and Low Islands in the Eastern Carolines'': 

In the years before World War II certain non-agricultural activities 
had developed in Ponape and Kusaie (the other high island of this dis- 
trict), but particularly in Ponape, in which the natives had little part. 
Commercial fishing was in the hands of Japanese using Okinawan 
labor. There was some small-scale manufacturing in Ponape, including 
fish processing, paper making, the making of starch from cassava and 
of alcohol from sugar cane, lumbering, soap making, ice making, cot- 
ton weaving and cigarette making. On both the high islands electric 
power plants were in operation. But none of these activities was in 
native hands. 

However, the natives or at least some natives did enjoy some fringe benefits from 
the agricultural and industrial activities of the colonizers. Actually there was 
room and resources for both colonizers and natives. They were not stupid col- 
onizers. They estabhshed health service for the natives, some schools and intro- 
duced many products of commerce for sale or barter, goods which changed the 
economy and way of life of the natives, whether it bettered them or not. 

The earlier history of foreign influence in the islands can only be outhned 
briefly. While both the Marshalls and Carolines were visited sporadically much 
earlier by sea-faring explorers (they were first discovered by the Spanish ex- 



36 The University of Texas Publication 

plorer, Alvaro de Saavedra, in 1528-29) they were regularly visited by New 
England whalers during the period 1820-1850. No one seems to have a good 
word for the influence of the whalers. Hobbs writes of the Marshalls: "The 
whaling crews introduced the diseases of syphilis and tuberculosis and the vice 
of drunkenness with the usual results." Henry Hedges, district administrator 
for Ponape, writes of that island: "contact with western whalers left disastrous 
effects on the lives of the people in this district, as in other areas of the Pacific 
during the middle 1800's." 

About the middle of the nineteenth century Protestant missions, (the Boston 
missionaries) were established in both the Marshalls and Carolines. The islands 
were held by the Spanish during much of the latter part of the 1 9th century and 
their priests established Catholic Missions on both the Carolines and Marshalls. 
The Protestant missionaries reduced the local languages to writing, translated 
the Bible and taught many of the local populations to read and write. Today both 
Catholic and Protestant missions are flourishing on Ponape and in the Marshalls, 
and the natives are roughly evenly divided between the two churches. It is my 
impression that the civil administration maintains friendly cooperation with both 
religious groups. Thus on Ponape the school administrators have worked out a 
plan with the Protestant mission by which efforts will be supplemented and not 
duplicated in the education of the natives. We met the Catholic priest on Majuro 
and he seemed to be a person of tolerance and charm. 

In 1899, following the Spanish American war, the Marshalls, Carolines, Gil- 
berts and Marianas, with the exception of Guam in the latter group, were ceded 
to Germany for the small sum of 840,000 pounds (Hobbs). Germany lost the 
islands to the Japanese following World War I. Thus through the 19th and first 
half of the 20th century the natives of these islands have felt the impact of out- 
side influence: American whalers, American Protestant missionaries, Spanish 
Catholic missionaries, Spanish, German, Japanese, and now American adminis- 
trators. Whether one likes it or not the fact remains that they are engulfed in the 
tide of western culture and civilization, nor can one except the strong Japanese 
influence, which was essentially western in many respects. The Americans at 
Majuro and Ponape are committed to the task of superimposing upon and blend- 
ing and coordinating some of the values of western culture with a native culture 
and economy seemingly in most areas of living well adapted to the environment. 
It is a challenging and at times frustrating problem. The program, both on the 
low islands of the Marshalls and on the low and high islands of the Carolines is 
designed to help the natives help themselves and as fast as possible native leader- 
ship is being trained to take over positions of responsibility. In the Marshalls at 
Majuro a hospital of several large quonset huts is at present staffed entirely by 
native Marshallese doctors, aides and nurses. The Marshallese have a hospital 
ship, the crew and staff of which all are Marshallese and which make the rounds 
of inhabited atolls, visiting each at least once every three months. Serious emer- 
gency cases may be flown to the Naval Base on Kwajalein or to Honolulu. Miss 
Elizabeth Hirst, hospital administrative assistant on Ponape, showed several of 
us through the hospital there. She had formerly been in a well-equipped hospital 
financed by the Rockefeller Foundation in China, and kept apologizing for the 
inadequate facilities and poor equipment in the local hospital. However, to the 



Spencer: High Islands and Low 



37 



layman it appeared that the Ponape Hospital was doing excellent work with 
modest equipment. We visited the T. B. ward, the leprosarium, and the general 
wards Yaws, one of the worst scourges in these islands in the past, has been elimi- 
nated by the use of modern antibiotics. T. B. is now the worst disease among the 
natives, probably due to inadequate diet and the extremely humid climate. 
Serious cases in both Marshalls and Carolines are hospitahzed. There were 
thirteen cases of leprosy in the Ponape hospital, none in the advanced stages; 
this disease seems on the way out with new drugs developed in recent years. 
Venereal disease is at present relatively rare in both island groups, though for- 
merly much more prevalent. One cannot but feel that the introduction of western 
medicine has been a godsend to these people, whatever may be said of other 
phases of western civilization. 

At Majuro, Tony Cruz, a native of Guam of Chinese extraction, with graduate 
training at Oklahoma A. and M. is in charge of the Agricultural Experiment 
Station. This is an Ag. station in miniature. We saw a good deal of Tony Cruz 
and were impressed both by his energy and with the problems which he faces. 
Cocoanut palms grow luxuriantly on all the low islands. In some of them taro 
swamps are maintained quite successfully. Papaya and breadfruit grow quite 
well. Tony has the task of trying to find other food plants which can be grown 
successfully in the salt-spray laden air and the sandy soil of the atoll islands. 
While Robert Louis Stevenson makes his point in the quotation read above on 
the paucity of animal and plant life on the atolls by a gross understatement, 
nevertheless there are few plants which grow here in any abundance. William 
Randolph Taylor in ''Plants of Bikini and other Northern Marshall Islands'' lists 
38 dicots and 5 monocots in addition to grasses. To those of you who know some- 
thing of taxonomic botany this is a restricted flora indeed. Atolls of the Southern 
Marshalls and Carolines would add very few new species. In contrast Sydney 
Glassman in his ''The Flora of Ponape'' Hsts several hundred species of the higher 
plants. I can say that from some experience in tramping over the motus of several 
Marshall Island atolls about a dozen trees and shrubs make up the bulk of the 
dominant flora. I will not burden you with a list of these plants. In contrast on 
several long hikes on Ponape I was impressed by the variety and luxuriance of 
the flora. Some of the Americans on Majuro try to grow a vegetable garden but 
when it comes to the culture of tomatoes, lettuce, etc., the results are rather dis- 
couraging. Most food plants can't take the salt air. Tony Cruz is breeding 
chickens, ducks and pigs in some numbers. Improved breeds of these will be re- 
leased to' the natives who have for a long time raised pigs and chickens, but with 
no attention to improving the stock. In the past the economy of the atoll islands 
has been based largely on cocoanut, shell-fish and fish. Taro is also an important 
food crop and is really the only crop which they culture. Cocoanuts grow wild 
and with little cuhivation. Natives simply harvest the crop which is maturing the 
year round. I hope Tony Cruz is able to find some new crop plants with high 
yield on small land surface, but the task is a formidable one. Papaya, bananas, 
bread-fruit, and pandanus can be grown on the low islands, but all flourish better 
on the high islands. 

On Ponape according to Murphy "between 40 and 45 species of wild and culti- 
vated plants are used as food by the natives. The list for the atolls in this region 



38 The University of Texas Publication 

would probably not total more than fifteen." Furthermore the high islands are 
undoubtedly adapted to growing many plants not now found there. Thus an ex- 
periment station on Ponape has a real chance of developing a diversified plant 
and animal breeding program. We spent parts of several days at this station and 
were shown around by the head agriculturist, Edward Iwaniec. I can only 
mention briefly some of the things we saw: taro swamps, where many varieties 
from the several atolls and Ponape were being grown and crosses made to de- 
velop improved types adapted to certain local conditions, cocoanut palm groves 
where records were kept of yield and quality, planting of tea, coffee, cacao, and 
rubber as possible commercial crops for the island, pigs imported from Japan 
and being fattened on Ponape bananas, of which there are many species grown, 
a small herd of the new Brahma cross cattle from the King Ranch in I'exas, and 
of course many exotic plants grown for ornament. One had the impression that 
Iwaniec and his corps of workers had a wide-open field for investigation in con- 
trast to the limited future of agriculture in the Marshalls. However, at the present 
time, the atoll islands both in the Marshalls and Carolines produce much more 
per acre and support a much larger population per square mile than the high 
islands. Ponape is still mostly a jungle wilderness. This point can be documented 
best by figures: Ponape has a population of 120 per square mile of arable land. In 
contrast in the Eastern Carolines Mokil Atoll has a population of 878 per square 
mile of arable land, Pingelap of 1025 per square mile, Kapingamarangi of 817. 
The atoll islanders have reached and perhaps in cases exceeded the population 
which can be supported there. They are free, however, to migrate to the high 
islands, where life would be easier and more abundant for less effort. Some migra- 
tion has occurred. It is not unusual for individuals, families and even larger units 
to migrate from low to high islands; practically no migration in the opposite 
direction has occurred. However, in spite of the more rigorous and precarious 
existence on the low islands the natives there for the most part are loathe to leave 
their homes for the easier life on Ponape and Kusaie. They claim to like the 
atoll climate better, though from my experience I can see little difference. In both 
places it is hot and humid. 

It is my impression from observation, conversation and reading that the low 
islanders of the Marshalls and Carolines are somewhat more industrious than 
the high islanders of Ponape and Kusaie. But one must be careful about such 
generalizations. If this be true it is probably because with more limited potential 
resources on the low islands the natives have had to work harder and to chance 
more to survive. Thus I understand that atoll dwellers in general are better 
sailors. They do much of their fishing in the open sea and in the past traveled 
hundreds of miles in out-rigger boats from atoll to atoll, navigating by stick 
chart. They were formerly expert in knowledge of ocean currents as M. W. de 
Laubenfels has pointed out in an article on ''Ocean Currents in the Marshall 
Islands'' in the following words: 

Every time a native sailboat was becalmed, its crew paid careful at- 
tention to the rate and direction of drift. If they succeeded in reaching 
land, they reported their experiences with great exactness. The older 
navigators thus accumulated a dependable store of information. Many 



Spencer: High Islands and Low 39 

scientists who value only complex instruments may belittle the words 
of natives, but practical naval experts (for example Captain John Vest, 
former governor of the Marshall Islands) recommend serious reliance 
on them. Special mention may be made of Lokrap of Ebon Atoll, whom 
the present author regards as the greatest of the surviving wise old 
navigators. The younger Marshallese do not carry on the old skills 
but prefer rather to get a Diesel engine and forget about currents. 

It seems unfortunate that the Marshall Islander of today has largely given up 
fishing, a major factor in their earlier economy. Now they work for the Trust 
Territory projects, or for the navy at about 25^ an hour and buy canned fish at 
the grocery. It is one of the projects of the administration to encourage a return 
to fishing, but now with larger boats carrying reefers for refrigeration. The 
Ponapeans do some fishing but inside the reefs; in contrast the men of the Ka- 
pingamarangi village on Ponape (immigrants from the home atoll five hundred 
miles to the south) go fishing regularly both inside and outside the reefs with 
their fleet of outriggers. 

On the Marshalls by far the largest industry is the production of copra, the 
partially dried meat of the cocoanut, which is used in making cocoanut oil, soap 
and other products. They have a native company, Mieco, Marshall Island Export 
Company, supervised at their request by an American, but run by Marshallese 
for the Marshallese. Last year they exported 12,000 tons of copra. At market 
price of $175 per ton this runs into over $2,000,000 gross, no small business for 
11,000 natives hving on less than 70 square miles of land. In the summer of 1955 
we attended an outdoor movie on Majuro, a walk-in not a drive-in. We sat on the 
grass with the natives and used our rain-coats before the show was out. We saw 
a picture "Storm Over Tibet" featuring avalanches in the Himalayas and won- 
dered how much of the picture was getting across to island natives who had no 
experience with snow or mountains. But they were attentive. On our visit to 
Majuro this past summer we were shown through the fine new Majuro Club 
building, put up by Mieco, designed and constructed by Marshallese from lumber 
they had imported from Japan. We left on a Thursday and the formal opening 
with appropriate celebrations was scheduled for the following Saturday. On the 
first floor was a movie theater accommodating between 1500 and 2000 with wide 
screen and modern projecting equipment, a restaurant and soda-bar and store. 
On the second floor were recreation rooms and a large Council Room where the 
Marshall Island Congress, with its upper and lower house, would meet. Cer- 
tainly, the high island natives of Ponape have no such ambitious native com- 
pany. A convenient bread-fruit tree near his dwelling furnishes the bulk of the 
family food the year round. While the crop is seasonal, the bread-fruit is pre- 
served in various forms and used the year round as food. Wilham Finale (native 
of Cleveland and the Educational Administrator on Ponape) told me that he 
lived for two months with a native family and that the diet was almost wholly 
of bread-fruit in different forms. He had meat only once in the two months. This 
of course, is a subsistence diet and not a properly balanced diet. But mam 
Ponapeans live that way. 

While the Ponapeans may be more shiftless than the low islanders because of 



iMHi 



40 The University of Texas Publication 

their more easily secured food supply, I don't want to imply that they are not 
capable of hard work, at least some of them. In the eight days we were on Ponape 
we saw natives bring coral on barges to the landing; grind it up; haul it by truck, 
spread and roll it down and surface a road approximately a mile long from the 
town of Colonia out to the Agricultural Experiment Station. It might be remarked 
that this road had a strong, fishy smell. Undoubtedly this would disappear as 
the coral animals decayed and the rain leached the organic material out. The 
Ponapeans were also putting up a new movie house but on a much more modest 
scale than that on Majuro. 

Let us now consider very briefly the school system set up by the Trust Terri- 
tory Administration. In the Ponape District there is an Educational Administra- 
tor, William Finale, and three American Assistants. In the neighborhood of 
thirty elementary schools are maintained, at least one in each of the five prov- 
inces on Ponape Island, several on Kusaie and at least one on each of the in- 
habited atolls. Under these administrators native teachers, invariably men, gen- 
erally in their late twenties or thirties, conduct these elementary schools. Here 
children are taught to read and write in Ponapean, and in English. Arithmetic, 
geography and history are also taught. The better students from these schools 
are brought to the secondary school in Colonia on Ponape and as boarding stu- 
dents continue their studies. They may spend two or three years here with 
infrequent visits back to their homes particularly if they live on distant atolls. 
This school would correspond to the upper grades in our own school system. 
Some of the better students from this school then go to the Island of Truk to the 
PICS school (Pacific Island Central) where they get a high school education. 
From there a very few of the best students go to the University in Honolulu or to 
the British University in Suva in the Fiji's. At Suva the British maintain a first 
class Medical College and it is here that advanced laboratory technicians and 
doctors get their training. Plans are underway to move the PICS School to Ponape 
where it is to be located on a tract near the Experiment Station. Ground is prob- 
ably already broken by now for some of the buildings. As I was walking in the 
Experiment Station grounds one hot, humid afternoon (the only kind they have 
the year round) I couldn't help overhearing an argument between Iwaniec, Ag. 
Station Head, and the chief surveyor for the road to be cut through the station 
grounds to reach the school. The argument was over where the road should go. 
Tempers were a little short. One school of thought had it that the road should 
be as direct as possible, the other that certain exotic trees and shrubs, not easily 
transplanted or re-grown should be saved regardless of where the road had to go. 
I was for the second position but didn't wait to hear the outcome as it was none of 
my business. 

After thirty pages of introduction we haven't yet gotten around to what was to 
have been the main thesis of this paper: the natives of the Marshalls and of Pona- 
pe, who they are, where they came from, something of their customs and 
manners. According to Thrum's Hawaiian Annual and Standard Guide, the 
Recognized Book of Authentic Information on Hawaii, "The Polynesians were 
the descendants of a family or tribe of Caucasian people who left their original 
homeland somewhere in India and wandered through Malaya to the island 
groups of the Pacific. In their travels through Asia, they intermarried with Ma- 



Spencer: High Islands and Low 



41 



layans and Orientals. From the inhabitants of the southeastern islands of the 
Pacific (Melanisia) [sic^ they also acquired darker blood, so that by the time 
they reached the Society Islands, now considered the ancient homeland of the 
Polynesian, they were a light brown people." If the thesis of Thor Heyerdahl in 
Kon Tiki is correct they also received an infiltration of American Indian blood 
from Peru, but of course the American Indians were originally mig^nts from 
Asia, so this does not notably change the origin of their racial stock. The Mela- 
nesians are thought to have a strong admixture of primitive Australoid blood, m 
addition to Asiatic strains, and the Melanesian aborigines have kmky hair and 
darker skins, which favor this thesis. The Micronesians would seem to have a 
mixed ancestry of both Polynesian and Melanesian origin. And one does not 
have to be an anthropologist to see in individuals of both Ponape and Ma,uro 
atoll evidence of blood transfusions from their earlier Western and Japanese 
overlords I gathered from conversations with personnel both on Ma]uro and 
Ponape that extracurricular activities between natives and admmistration per- 
sonnel are at present neither condoned in the social rules of order "or even en- 
gaged in clandestinely. Of course, during the occupation of the islands m World 
War II things were different. , 

The people of Majuro Atoll in the Marshalls and of Ponape m the Carolines 
are on the average people of medium height, stout in body build, with straight, 
coal black hair, brown eyes, and broad flat noses. Superficially at least they are 
friendly greeting you with smiles and a friendly salutation either m town or on 
a trail far from town. When I mentioned this to the anthropologist on Ponape 
he suggested that this might be an act put on by natives to keep on the good side 
of the Americans. My answer to this was that I always spoke to all children I met. 
The American children on Ponape were likely to be reserved; they might return 
the salutation but they didn't often smile. In contrast the Ponapean children 
almost always smiled broadly. I don't know that I won the argument^ Possibly 
my bizarre appearance was sufficient to elicit the incredulous smile of a native 
youngster But I still hold to the hypothesis that among these people there is a 
larger proportion of pleasant extroverts. Mr. Neas, administrator on Majuro, 
told us that there were no crimes of violence, murder, rape or major thievery 
among the Marshallese. 

Perhaps rape is less common in the islands because of the somewhat more 
casual relationship of the sexes. It is significant to note that in both the Marshalls 
and the Ponape District there is a matrolineal system of land inheritance. Ihis 
was explained to us as being a system which fitted in well with social customs 
where it is always easy to determine the mother of a child but not so easy to know 
who the father was. In the Carolines I am told that each young man of mar- 
riageable age has a love-stick with unique carvings on it. If he fancies a certam 
maiden he goes around to the hut where she sleeps and pokes the love stick m. 
Apparently she can recognize the carvings on the stick in the dark and if she 
favors the suit she pulls on the love-stick and the suitor comes m. If he doesn t 
get the green light I suppose he goes off to the hut of No. 2 on his hst. I don t 
know how much of this is fact and how much fancy. But I do know that Chaplain 
Hoppe, from Eniwetok, a member of our party, had a native boy as a guide on 
one hike which he took. He questioned the boy on this matter and he replied that 






42 The University of Texas Publication 

he had the love-stick and the girl picked out but so far hadn't gotten up the nerve 
to try the experiment. On the eve of our departure from Ponape the very at- 
tractive waitress at the galley presented a love-stick (unfortunately not to me 
but to the one unmarried member of our party). She assured him that this was 
just a souvenir and had no further significance. It would seem that the love- 
stick custom has some phallic significance but I am no anthropologist. 

If a married man goes off to another atoll to work for a few months it is not 
unusual for him to take another consort on that atoll and for his wife to make an 
appropriate connection back home. I noted in Glassman's ''TJie Flora of Ponape'' 
that between 30 and 40 plants were listed as drug plants used by native doctors, 
and among these a surprising number were used as contraceptives. I don't know 
how successful these are but I do know that the anthropologist on Ponape, Mr. 
Richard Emerick, told me that native families there were likely to be three or four 
children. To him the low birth rate was a puzzle and a problem for future investi- 
gation. Native populations are now on the increase after a long period of de- 
pression, but this is due to introduction of western medicine and not to high 
birth rates. 

What kind of clothes do the natives wear? I was asked this question repeatedly 
after my return from the Marshalls. I could only say then that the women of 
Majuro were the most dressed women I saw anywhere on my travels, with more 
bare epidermis on display on any beach in America or for that matter on any 
downtown street or in any grocery store in mid-summer than one would see in the 
Marshalls. Both men and women wore cheap cotton clothes, the women with 
"mother hubbards" which certainly concealed the form quite well. We were told 
that this was a result of the Puritan, prudish influence of the Boston missionaries. 
On Majuro male children often went naked; I saw a group of these boys diving 
off the dock and it reminded me of the old swimming hole back in Southwestern 
Ohio where I grew up. But females of the same age were always clothed. 

What kind of homes do the natives live in? Both on Majuro and Ponape most 
of the natives live in shacks constructed of odds and ends of lumber and sheets 
of galvanized iron. They are ugly, not a redeeming feature except that I suppose 
they keep the rain off and there's lots of rain. But I could not help but contrast 
them with the beautifully and elaborately constructed native houses on Rongelap 
atoll, and the less elaborate thatched houses in the Kipingamarangi village on 
Ponape. The Rongelap houses, abandoned when the natives left in the summer 
of 1954, are being replaced by western style homes. Since they were abandoned 
we had the chance to inspect them closely. The thatching is of pandanus leaves, 
with mid-ribs from palm leaflets used as binding and supplemented with cord of 
palm fiber. The work is elaborate, beautiful and durable. From the inside the 
pattern is perfection. These houses with their slanting roofs, broad, overhanging 
eaves, and with side walls rising only about four feet and open above are ad- 
mirably adapted to the climate. In the Kapingamarangi village on Ponape the 
roof thatching is of palm leaves (pandanus is not so abundant here) and much 
less elaborate but quite adequate. A few of the houses in this village and the 
church are constructed of lumber and galvanized iron. I remarked to the native 
teacher in the village, a young man who spoke some English, that I Hked much 
better the looks of the native huts. He said in reply: "But we like the other build- 



Spencer: High Islands and Low 



43 



Ws better. It's easier to lay on a sheet of corrugated iron than to make the thatch- 
ing " He told me, however, that the thatch would last about five years and one of 
the construction men on Ponape told me that the corrugated iron would last about 
the same length of time. Native boats, native clothes, native houses, soon to be 
a thing of the past. But quite possibly when and if we pull out of this territory 
the natives will have to go back to their old ways. Perhaps, who knows, the natives 
of Yap and Kapingamarangis are the wise ones. They have best retained the 
old cultures and would have less adjustment to make in case the ships and planes 
no longer came to their shores. . 

In the Ponape Co-op run by Joe Riddle, who lives with his family close by m 
the old Japanese Geisha house, natives can and do buy all kinds of western goods^ 
Utensils of rubber, buckets and dishware are very popular. They have a much 
longer life than those of metal which soon rust out. It's easier to buy cord at the 
store than make it out of palm fibers. Nails are so much better than palm hber and 
wooden pegs when it comes to constructing houses. The standard wage for labor 
on Ponape is 15^ to 28^ per hour. No doubt if I were a native Ponapean on that 
wage I would be buying utensils and cotton goods in the co-op rather than making 
cloth out of native grasses, and utensils out of cocoa-nut shells and sea-shells. 
More than once I saw what seemed a symbohc picture, a native woman far back 
in the bush, industriously running a Singer sewing machine, no doubt construct- 
ing articles of clothing to cover her family. But why? She'd be more comfortable 
with fewer clothes. I sometimes wonder if the human rat-race which we call 
western civihzation is the final answer to the age old problem of the good life on 
the planet earth. Just think of buying a Singer sewing machine on a 25^ per 
hour wage. The Cadillacs aren't there yet but surely they're on their way. 

I want to close this rambling discourse with a few word pictures as substitutes 
for the kodachromes I didn't take because we were not allowed to carry photo- 
graphic equipment into the Proving Grounds and therefore to other places we 

visited. r t» i j 

A native luau (feast) on Ponape. Bill Finale and his charming wife, Betty, had 
invited us to dinner at their home on Ponape, Monday evening, August 27th. In 
the meantime Bill had discovered that his native teachers on Ponape were ar- 
ranging for a luau in honor of three young men of Ponape, returning from the 
University of Hawaii in Honolulu. So the two festivities were combined. About 
dusk Henry Takeshita, a Japanese American originally from Honolulu and ad- 
ministrative assistant (if you wanted to know anything ask Henry) called at 
the crew's quarters and picked us up in that ancient truck we had ridden on 
many times before. A few minutes later we were in the living room of the Finale 
home, when drinks were being served, and introductions made to native teachers 
and the guests of honor, the Honolulu boys back home. Soon we were out on the 
lawn, brilliantly hghted with arc lamps. On the lawn were spread big pandanus 
mats; in the shadows could be seen banana trees, palm trees and other luxuriant 
tropical vegetation; a buffet table had been prepared; a table covered with huge 
banana leaves as table cloth; on the table was an unbelievable array of native 
dishes. I am no gourmet, only an amateur in the arts of the delicatessen and the 
cuisine, but I can think of only one comparable spread in my experience, a 
wedding feast in Egypt in the home of a wealthy land-owner long before the days 



^g^^gmmmmmmmm 



44 The University of Texas Publication 

of Nasser. On that table lay a half-grown pig, roasted and bisected down the 
middle, a huge plate beside it containing the roasted extremities of pigs' legs, 
jaws and parts of heads; some of the jaws looked too much like dog to suit an 
amateur anatomist, so I settled for a leg; breadfruit in three different guises, 
yams, baked bananas, taro root, cocoanut in different forms including a pudding; 
rice in monumental heaps and several other items. We helped ourselves, buffet 
style, and sat on the pandanus mats. Before we ate, natives made the rounds and 
placed on the brow of every banqueter a wreath made of exotic tropic flowers. 
During the meal cocoanuts were passed and from them we drank the milk. Coffee 
and drinks were also served. As table companion I was fortunate in having Connie 
Hedges, wife of the administrator, and she regaled me with tales of their years on 
Bora Bora near Tahiti, and of the experiences in taking natives of Tahiti and 
Typee back to Chicago. Ponape skies were kind and it didn't rain that night. To 
me it was an experience out of the Arabian Nights. I could only wish that my wife 
had been there for I'm sure her capacity for the enjoyment of the romantic and 
the exotic is greater than mine. But such is life. What do I think of native food? 
It's flat, tasteless, and not sufficiently seasoned. I'd settle any time for the kind of 
food served us daily at the A. E. C. base on Parry Island. 

Saturday night. Party at Colonia Club on Ponape. What's better than an eve- 
ning of good conversation, particularly when the setting is a Clubhouse on a far 
off Pacific Isle, and the participants young intellectuals eager to match wits with 
outside visitors of which there are too few in that faraway place? I'll tell you 
what's better: a beautiful young Japanese American from Honolulu, dancing an 
authentic hula at 2:00 A.M. that "evening" at the club. It's the arms, the hands, 
and the fingers that tell the story and not the hips, believe it or not. Well, that's 
something to park away in the cortical folds along with the first view of Ponape 
from the air. 

Long Hike into the Hinterland on Ponape. On the morning of August 28th 
Marshall Wheeler and I took a long hike, about ten miles, from the province of 
Net into the province of Uh on Ponape. Afterwards Wheeler told the others that 
there was only one thing wrong with taking a hike with Spencer: he walked too 
fast. But that wasn't fair, for Wheeler had coral cuts on both feet, which I didn't 
know of at the time but which must have been painful. Then I had gotten into 
condition by tennis earlier in the summer. I had earlier made a trial run on this 
trail. Along the trail we passed two mountain streams where Ponapean women 
were busy beating the dirt out of the family washing with clubs. I suppose this 
treatment gets the clothes clean, but it must be hard on buttons, and fabric. I've 
seen the same technique used in Egypt and I suppose it is universal custom in 
the tropic belt around the world. A little later we crossed a meadow on the border 
of which we saw several species of tropic birds (a few species are found in Ponape 
and nowhere else). One is a brush-tongued honey-sucker, a specimen of which 
the Riddles have as a pet. They call him Einstein, and when I asked Riddle why 
he didn't take Einstein to the parties he said that he would be bored at the level 
of the conversation. A little further along the trail we saw huge clumps of bam- 
boo, and further along on higher ground between two mountain ranges huge 
tree ferns, some forty feet in height; still further a species of palm, the individual 
leaves of which were between thirty and forty feet long and the petioles of the 



Spencer: High Islands and Low 



45 



leaves at the base almost big enough to fashion into a dugout canoe; unfortu- 
nately not being a botanist I could not check the species from Glassman s check- 
list of the plants of Ponape. The trail led at one place across a very substantial 
bridge built by the Japanese over a lagoon; most of the trail found black volcanic 
rock underfoot, sometimes slippery, always rather rough and not too easy hiking. 
But the changing scenery as we hiked back along a valley trail between he 
mountain ranges, with rank tropical vegetation on either side, native huts tor the 
most part solitary and not in villages, natives on the trail and at work near their 
homesites, all this was an experience worth more than the primary object ot the 
iourney We decided later that this was the main-line trail from the north to the 
south of the island and a few hundred thousand dollars would turn it into a 

passable highway. , i • i • . „f 

A visit to the Kapingamarangi Village on Ponape. This was the high-point of 
my stay on Ponape. Actually I made three visits to this village, but we will merge 
them into one Let's make the trip together, but we'll take Chaplain Hoppe along, 
for he has been out there several times doing some oil painting and he knows the 
ropes As we leave the crew's quarters with the tropic sun shinmg brightly on a 
late August morning some will carry rain-coats and others will not. I belong to 
the latter school, for I have decided that it's easier to get wet from the ram and let 
my dacron pants dry out than to wear a rain-coat during a shower and get wet 
from sweat We'll take the road up past the administration building and the 
ball diamond, past the hospital, the garage and the filling station, all important 
landmarks in this community. Before we come to the secondary school and the 
experiment station we take the road oft to the right, past a very ftne planting ot 
pineapple and up onto the high ground overlooking the lagoon, beyond which 
can be seen Jokaj (Sokehs) Rock. We simply follow the trail and after passmg 
several Ponapean houses of the usual scrap lumber and galvanized iron construc- 
tion we come upon a few thatch roof houses. A few minutes later we are walking 
down Main Street, Kapingamarangi Village. These people are not Ponapeans. 
They have moved here from the home atoll five hundred miles to the south. 1 here 
are about 200 natives in the village and four hundred more on the atoll. That's 
all the Kapingamarangis there are. Yet they have their own distinct language, 
quite different from Ponapean, and they have their own culture also quite dif- 
ferent Hoppe knows proper protocol. He takes us first to the long-house where we 
meet the head-man of the village. The long-house sounds like a chapter out 
of Melville's Typee. This is the biggest thatched building m the village. The sides 
are open; there is a raised board floor. Here only men congregate, except on very 
special occasions. That day there are seventeen men, young and old, m the long- 
house. Most of them are busy making fish-nets. A few of them are lying around, 
evidently taking a rest. The nets vary in size from one which must be thirty or 
forty feet long with large mesh, two inches in diameter, to dehcate one-man 
throw nets, with a mesh about one-half inch diameter. Finished nets are also 
hung around the long-house. The chief does not speak English, but fortunately 
the young Ponapean teacher, who runs the school in the village, is there and he 
can answer some of our questions about fish nets and other things. I attempt to 
duplicate the tatting technique of one of the net-makers, and my awkwc:rd move- 
ments mildly amuse the onlookers. Either mystify a native by magic or make him 



■Mi 



46 The University of Texas Publication 

feel superior by obvious awkwardness in a skill in which he is expert. Soon 
cocoanuts are brought in and one of the young men makes a few deft slashes with 
a machete and has them opened at one end. We take one and drink the cocoanut 
milk; there must be a quart of it, mildly sweet and cool, a good refreshment for 
a parched palate on a hot day. The chaplain then suggests that we visit the 
church, which we do, walking through the back yards of villagers on the way and 
keeping our eyes open for the sights, including (as you have guessed) Kapinga- 
marangi women in their native costumes, often soon supplemented by a swath 
of cloth. The church is a wooden building, smaller than the long-house, open on 
the sides but with iron bars. There are wooden benches for the communicants; 
seating capacity is about 150. Having visited the church and the long-house, 
where village council meetings are held, and met the head-man of the village, 
we are free to wander about through back-yards and peek into native houses. At 
least this was the Hoppe theory and it certainly gave us a fine opportunity to 
view native life. In one house there was a young mother, swinging her infant 
tied in a hammock. At the backdoor of another house we saw a young woman 
grating cocoanut. To do this the native sits astride a stool shaped like a saddle. 
In place of the horn on the saddle there is a projection carrying either a jagged 
piece of shell or a piece of metal with jagged edge, purchased at the Co-op. A half 
cocoanut is rubbed over this rather deftly and fine grated cocoanut results. In one 
back-yard pandanus leaves were spread out to dry in preparation for weaving 
them into mats. Women were cooking over open fires. Water was drawn from 
a stone-lined well near the center of the village. Large fish were hanging up near 
some huts. Children were bathing by pouring water over themselves, dipped 
from empty gasoline drums, set under the eaves of thatched huts to catch rain- 
water for washing and bathing. A new thatched house was being constructed 
for a newly married couple. I actually helped to raise the framework of the roof 
onto the stout poles set in holes in the ground. My help was not needed as there 
were about fifteen natives there to do the job. The following day a revisit of the 
village found the young husband alone starting on the laborious process of con- 
structing the thatch. The wife was a pretty girl of about twenty years. Inci- 
dentally, the Kapingamarangis are Polynesians and somewhat lighter in com- 
plexion than their Ponapean neighbors. The day previous to this visit I had seen 
several natives sawing up the carcass of a beef in the village. I like to think this 
was in preparation for the wedding feast, and probably it was. We climbed down 
the steep path along the cliffs to the edge of the lagoon where we found the 
Kapmga boathouses. Perhaps a dozen big out-rigger canoes were housed there 
under thatched roofs, in preparation for trips both inside and outside the reefs to 
use the nets we had seen in construction. We were told that the big nets lasted 
only a few weeks, but the smaller throw nets might last for a couple of years. The 
nets were being constructed of cord, bought at the Co-op. That's easier than 
making the cord out of palm fiber. You'd probably do the same if there were a 
Co-op nearby. We were impressed with the neatness of Kapingamarangi yards; 
carefully kept sand and gravel paths, patches of grass with no weeds in them, 
quite in contrast to the rather slovenly appearance of the surroundings of a 
typical Ponapean home. But I am loathe to draw the obvious conclusion that low 
islanders are more thrifty and neat than high islanders, for certainly the yards 



Spencer: High Islands and Low 47 

of natives on Majuro were as ill-kept as those of the Ponapeans. Yet they were 
low islanders. When the villagers heard that Wheeler and I were flying down to 
their home atoll, five hundred miles to the south, on the following mornmg they 
were quite excited. Through the school-teacher as interpreter they asked if they 
might send a packet of letters to their relatives there. We, of course, said we 
would be glad to take the letters. The following morning one of their number was 
on the dock, from which the picket boat left, with a big stack of letters. Unfor- 
tunately I was bumped for a V.I.P. on this trip, so never got to see the atoll. But 
Wheeler delivered the letters and brought a big pack back, as well as models of 
outriggers and some fine basketware given him in appreciation by the chief of 
the Kapingamarangis. 

An Evening at the Club on Majuro. It is an evening in late August, 1955; our 
party is lounging in easy chairs in the Majuro Club. We are listening to a dozen 
or so of the Trust Territory boys discuss their several problems, including absent 
and incoming personnel, friendly gossip if you wish. The native bar-boys pad 
softly about, taking orders and mixing drinks. The surf from the open sea beats 
on the coral reefs just outside the screened but open siding at our backs. The 
temperature is down to the comfortable seventies with soft sea breezes wafting 
gently. Except for the lack of British accent, just like a chapter out of Kipling. 
Same setting, late August 1956, but the restful, out-of-this world atmosphere 
completely spoiled by the addition of one new environmental factor. Guess what 
it is? An American juke-box blaring jazz tunes all evening on a constant diet of 
nickels which you may be sure I did not supply. Let us consider it a symbol of 
western influence on a far-off tropical isle. 

I close with a quotation from John Wesley Coulter in an article entitled: ''The 
United States Trust Territory of the Pacific Islands'': 

The aspiration of native peoples for self rule is one of the factors of 
disquietude in the world today. It is not likely that any of them could 
maintain independence. They have to be educated in their own best 
interests and trained to look after themselves as far as possible. The situ- 
ation demands from the advisors to native authorities training, knowl- 
edge of the psychology and history of the people and constant skill and 
attention to determine when to interfere and when to leave well enough 
alone, when to hasten the development of more advanced methods of 
government and when to advise a slowing down of processes too rapid 
to be assimilated. There are no pecuniary rewards to be hoped for by 
those nations charged with trusteeships. A trustee is, in terms used 
thousands of years ago, his ''brother s keeper.'' 



■■HHB 



Darwin's Influence on the Study of Genetics and the 
Origin of Life' 

JAMES F. CROW 

University of Wisconsin 

It is customary for geneticists to point with pride to the fact that twentieth 
century genetics has supphed the crucial missing elements in Darwin's theory 
of evolution by natural selection. But what about the reverse problem? How has 
modern genetics been influenced by Darwin? It is this question that I propose 

to discuss. 

I suppose that among all the diversity of subjects on which Darwm wrote, his 
writings on genetics appear to be the most confused to the modern reader. 
Consider his works on coral reefs, on earth worms, on heterostyly in plants, on 
self sterility mechanisms in general, on barnacles, on the habits of climbing 
plants, on orchids; these are remarkably perceptive and their essential correct- 
ness has been confirmed by modern work. 

On the other hand, his vacillation about Lamarckism, his hypothesis of 
pangenesis, and his astonishing credulity in accepting folk lore and anecdotes 
makes his writings on heredity seem among the weakest and most puzzling of his 
work. Yet modern genetics owes a tremendous amount to Darwin. 

The reason, of course, is the notion of change by natural selection. Para- 
doxically Darwin's great influence on modern genetics has come, not from his 
writings on heredity and variation, but from his idea that natural selection, by 
multiplying variations, is sufficient to account for the diversity of living forms. 

INBREEDING AND HYBRIDIZATION 

With his characteristic patience and attention to detail, Darwin did a monu- 
mental study of the effects of inbreeding and what we now call heterosis. He 
published the resuUs, also characteristically, as a large book. 

After trials on many plant species, he noted that inevitably inbreeding led to 
a decline in height, weight, and vigor. He also realized that the effects of inbreed- 
ing were gradual and cumulative over several generations, but that there was 
immediate recovery on outcrossing. A similar increase in size was seen m 

variety crosses. 

All these results are famihar to modern geneticists and easily understood as 
consequences of mendelian theory. Darwin also noted two other effects of 
inbreeding that seemed very puzzling to him— the prepotency of inbred lines m 
outcrosses (now understood to be a consequence of homozygosity and domi- 
nance) and the uniformity of plants within inbred lines. He called attention to 
the fact that the flower colors in Petunia and Dianthus, which were ordinarily 
heterogeneous, became remarkably constant with inbreeding; so much so. that 

1 Paper Number 725 from the Department of Genetics. This is a slightly modified version of a 
talk given at the Symposium on "Darwin's Impact on Biology" sponsored by the Society for the 
Study of Evolution. 



50 The University of Texas Publication 

his gardener had no difficuky in recognizing inbred strains by this fact alone. 
Darwin says (1 ) : "As such cases of flowers becoming uniformly colored without 
any aid from selection seems to me curious, I will give full abstract of my 
observations," 

Darwin's results and conclusions on crossing still stand up as empirical ob- 
servations. In his own words the general conclusions can be summarized (1): 
"The most important conclusion at which I have arrived is that the mere act of 
crossing itself does no good. The good depends on the individuals which are 
crossed differing slightly in constitution, owing to their progenitors having been 
subjected during several generations to slightly different conditions, or to what 
we call in our ignorance spontaneous variation." 

All Darwin lacked was an underlying explanation. The results of inbreeding 
and crossing follow so directly from mendelian theory that we find it hard now 
to put ourselves in Darwin's place of not having this knowledge. He did a 
remarkable job in seeing the generalities of this important phenomenon which, 
because of hybrid corn, is probably genetics' greatest economic contribution. 

Realizing, as he did, that inbreeding leads to decline in vigor and is generally 
harmful, Darwin was fully prepared to find inbreeding ayoidance systems and 
his studies of self sterility mechanisms followed naturally as a consequence of 
his knowledge of inbreeding and his theory of natural selection. He also noted 
the sterility that is a barrier to species and variety crosses. Again, I shall quote a 
pertinent passage (1): 'Tt is an extraordinary fact that with many species, 
flowers fertilised with their own pollen are either absolutely or in some degree 
sterile; if fertilised with pollen from another flower on the same plant, they are 
sometimes, though rarely, a little more fertile; if fertilised with pollen from 
another individual or variety of the same species, they are fully fertile; but if 
with pollen from a distinct species, they are sterile in all possible degrees until 
utter sterihty is reached. We thus have a long series with absolute sterility at 
the two ends; — at one end due to the sexual elements not having been sufficiently 
differentiated, and at the other end to their having been differentiated in too 
great a degree, or in some peculiar manner." 

It is interesting to note that Darwin took the view, so ably argued in recent 
years by Muller, that hybrid sterility is incidental to evolutionary divergence, 
not ordinarily selected as an isolating mechanism per se. 

PANGENESIS 

In many ways Darwin's most unfortunate hypothesis was the notion of 
pangenesis, put forth in great detail in "The Variation of Animals and Plants 
Under Domestication." Yet, despite the general discrediting that it now receives 
the hypothesis is worth a closer look. It did account for a number of diverse 
facts. Throughout his life, Darwin was always looking for ways of bringing 
seemingly unrelated facts under a common viewpoint. 

According to this notion "gemmules" were produced in various parts of the 
body and eventually reached the germ cells where they carried information 
from various somatic tissues to the next generation. This provided a formal 
explanation for Lamarckian transmission of effects of use and disuse. 



Crow: Darwin and the Origin of Life 51 

Darwin apparently was never quite satisfied with this idea. He even had 
trouble with the name itself as the following quotation from a letter to Huxley 
attests (4) : "What I called pangenesis means that each cell throws off an atom of 
its contents or a gemmule, and that these aggregated form the true ovule or bud, 
etc. Now I want to know wether I could not invent a better word. Cyttarogenesis 

i.e. cell-genesis — is more true and expressive, but long. Atomogenesis sounds 

rather better, I think, but an 'atom' is an object which cannot be divided; and the 
term might refer to the origin of atoms of inorganic matter. I believe I like 
pangenesis best, though so indefinite; and though my wife says it sounds wicked, 
like pantheism; but I am so familiar now with this word that I cannot judge. I 
supplicate you to help me." 

Furthermore, Darwin was always careful to label the pangenesis hypothesis 
as provisional (2) : "I am aware that my view is merely a provisional hypothesis 
or speculation; but until a better one be advanced, it will serve to bring together 
a multitude of facts which are at present left disconnected by any efficient cause." 

One of the most definitive tests of the hypothesis was made by Galton who did 
several experiments that failed to show any inherited changes consequent to 
blood transfusions (5). Darwin did not regard this as a definite refutation 
however, pointing out that pangenesis applies to plants as well as animals and 
that, although he certainly would have expected to find gemmules in the blood, 
this is not a necessary part of the hypothesis (2) . 

The hypothesis of pangenesis deserves mention today for another reason. Cur- 
rently we hear a great deal about self reproducing particles. Darwin thought of 
his gemmules as such particles. He was aware of the multiphcation of the causa- 
tive agents of smallpox and rindepest and suggested that gemmules might 
multiply the same way. 

Another aspect of the pangenesis hypothesis is that it provided an "explana- 
tion" for the still unsolved problem of communication between different organs 
of the body. How, for example, is the right kidney instructed to grow when the 
left has been destroyed? Hypotheses not too dissimilar to pangenesis are cur- 
rently invoked. 

DARWIN'S VIEWS OF LAMARCK 

It is of interest to see how Darwin's views of Lamarck changed during his life. 
In 1863 he wrote to Lyell (3) referring to Lamarck's book as "what I consider, 
after two deliberate readings, as a wretched book, and one from which (I well 
remember my surprise) I gained nothing." But with successive editions of the 
"Origin of Species" more and more Lamarckian ideas came in. 

I think there are three principal reasons for this. First this was the prevailing 
notion at the time and Darwin was keenly aware of the thoughts of his 
contemporaries. 

A second reason is what seems to be an amazing credulity on Darwin's part. 
He accumulated large numbers of isolated examples in his notes and these 
included folk lore and anecdotes about the inheritance of environmentally in- 
duced effects. His magnificent ability to consider a great diversity of facts and 
ideas simuhaneously, and to bring them all under a common point of view here 
played him false— for too many of them were wrong. Darwin was caught in 



52 The University of Texas Publication 

what he himself reahzed was a serious problem, for he once wrote (6): 
"False facts are highly injurious to the progress of science, for they often endure 
long; but false views, if supported by some evidence, do little harm for every 
one takes a salutary pleasure in proving their falseness." 

The third reason is the most interesting. Because he subscribed to the blend- 
ing theory of inheritance current at the time, he greatly exaggerated the amount 
of new variability that would be required to maintain the population in a state 
sufficiently variable for selection to occur. Fisher (7) has shown that the variance 
is decreased by exactly half each generation with blending inheritance, whereas 
with mendelian inheritance a very low rate of mutation is enough to offset any 
effects of selection and random extinction. 

Although Darwin does not appear to have been aware of the quantitative 
aspects of the problem, he must have had at least a qualitative notion. I believe 
he would have been highly receptive to Mendel's work for, as Fisher has noted, 
he once speculated in a letter about the possibility of particulate inheritance (4) : 
"Approaching the subject from the side which attracts me most, viz., inheritance, 
I have lately been inclined to speculate, very crudely and indistinctly, that 
propagation by true fertilization will turn out to be a sort of mixture, and not 
true fusion, of two distinct individuals, or rather of innumerable individuals, as 
each parent has its parents and ancestors. I can understand on no other view the 
way in which crossed forms go back to so large an extent to ancestral forms. But 
all this, of course, is infinitely crude." 

Today the discussion has shifted to new grounds. Within the framework of 
mendelian theory, what maintains the variance? Is it largely recurrent mutation 
to deleterious alleles with occasional polymorphic loci? Or is the typical locus 
heterotic as Wallace and Dobzhansky have suggested? How much potential 
variability is tied up in linked combinations? What is the role of the population 
structure in maintaining variability? 

The queston of what it is that mantains variability is still with us. The differ- 
ence is that whereas Darwin had no explanation, we now have too many. 
Because variability is so easily conserved in a Mendelian system, there are 
several mechanisms any of which could be sufficient. 

NATURAL SELECTION 

The greatest contribution that Darwin made to genetics is, of course, the 
same as the great contribution that he made to biology in general — natural 
selection. The idea is at once so simple and self evident, so general and all 
pervasive, and so thoroughly taken for granted that there is very little for me to 
add. It has given a historical meaning, a perspective of depth to all biological 
properties. It provides a new reason for the study of mechanisms of inheritance 
and variation. 

Usually the person most willing to accept and use a hypothesis is its inventor. 
Not so in this case. While Darwin was cautious in applying his notion, modern 
geneticists have paid Darwin the honor of being more doctrinaire in accepting 
his theory and extending it to new situations than he was. We are much more 
likely to regard selection as the sufficient explanation, the first factor to be ruled 



Crow: Darwin and the Origin of Life 53 

out in any novel situation. For example, we no longer even stop to ask why 
domestic animals are more variable than their wild forbears— a subject to which 
Darwin devoted a two- volume treatise. Whereas he was unwilhng to attribute 
this to simple selection, and looked for other mechanisms, we now regard it as 
entirely due to man's preservation of variants that would not have survived m 

nature. i i n 

We now emphasize that selection operates at many levels— between cells, 
between organisms, between demes, between species. But, although the ideas are 
now much more refined and quantified, thanks to Fisher, Haldane, Wright and 
more recently Malecot and Kimura, Darwin was aware of many of the complexi- 
ties. For example, he considered the evolution of sterile castes in a bee colony as 
an example of inter-population selection. Strangely, he overlooked the important 
evidence this offers against Lamarckian inheritance. 

Current selection theory assumes, much more than Darwin did, that much 
selection is wasted in that it has very little permanent effect on the genetic 
makeup of future generations. Darwin realized that not all variants were 
hereditary, but he had no way of knowing how frequently even genetically 
determined variants would not respond to selection. 

DARWIN AND THE ORIGIN OF LIFE 

Darwin's name is not ordinarily associated with the question of the origin of 
life, and I think he would want it that way. He argued strongly against some of 
his opponents who were maintaining that it was pointless to discuss the evolution 
of hfe without knowing its origin. He maintained, as almost all contemporary 
scientists would, that one doesn't have to know final causes to study immediate 

causes. 

In fact, Darwin was quite inclined in his early years to ignore the subject ot 
the origin of life. In 1863 he wrote to Hooker (3) : "It is mere rubbish, thinking 
at present of the origin of Hfe; one might as well think of the origin of matter." 

Yet, Darwin has made a contribution to contemporary thinking about the 
origin of life in two quite different ways. The first of these is a remarkably 
perceptive statement made in one of his letters. 

Until recently biologists have largely held to what Hardin (8) has called the 
autotroph theory of life's origin. It seems at first glance to be most reasonable to 
assume that the earliest forms of life made their own organic matter and had 
simple nutritional requirements. For, where do organic chemicals come from? 
From living organisms, of course. Ergo, the earhest forms had no organic com- 
pounds to live on. Furthermore, as George Wald has said, Wohler's synthesis of 
urea didn't prove the contrary— it only showed that a human being can make 
urea externally as well as internally. 

But since Haldane's essay in 1925 the idea that the original forms were 
heterotrophs has gradually become accepted— thanks to Oparin, Horowitz and 
others who have added the needed details. But Hardin has noted that Darwin 
himself understood the essential point. Haldane's famous essay was anticipated 
in part by Darwin a half century earher, when he wrote (3) : "It is often said 
that all the conditions for the first production of a living organism are now 



54 The University of Texas Publication 

present, which could ever have been present. But if (and oh! what a big if!) we 
could conceive in some warm little pond, with all sorts of ammonia and phos- 
phoric salts, light, heat, electricity, etc. present, that a proteine compound was 
formed ready to undergo still more complex changes, at the present day such 
matter would be instantly devoured or absorbed, which would not have been 
the case before living creatures were formed." 

Compare this with the following extract from Haldane's essay (9): "In this 
present world such (organic) substances, if left about, decay — that is to say, 
they are destroyed by microorganisms. But before the origin of life they must 
have accumulated until the primitive oceans reached the consistency of hot 
dilute soup. Today an organism must trust to luck, skill, or strength to obtain 
food. The first precursors of life found food available in considerable quantities 
for existence." 

Both writers were ahead of their time, and their views were not until recently 
part of current biological thinking. 

What is the essential feature of life? Muller as early as 1921 (10, 11) argues 
most convincingly that life depends on self reproduction, of course, but more 
uniquely on the ability to multiply variants. That is, life must have both self 
copying and mutation, with the mutant types being capable of copying 
//z^777 selves. 

It is not necessary that the earliest forms be able to copy themselves ac- 
curately. In the beginning the probability of a correct copy need not have been 
much higher than that sufficient to offset purely thermal or thermodynamic 
degeneration. There may have been many more mistakes than correct copies. In 
this sense mutation may have been anterior to self reproduction in life's history. 
Natural selection would favor those whose ratio of correct to wrong copies was 
greatest — that is selection would lower the mutation rate. 

Contemporary implication of DNA as the genetic material means that a 
relatively simple substance may have self copying properties. 

I want to bring Darwin into this by suggesting that a meaningful, and perhaps 
useful, definition of life is that it is a system that permits the operation of natural 
selection. 

I want to push natural selection as far back as possible into what is generally 
regarded as the pre-hving, chemical realm. If it is generally agreed that macro- 
mutation is inferior to successive micromutation as an explanation of organic 
evolution, why not use the same thinking for pre-organic? An example of such 
thinking is Sagan's (12) discussion of systems that favor the accumulation of 
polymers in preference to monomers. 

Now that space science is beginning we can start to answer questions about the 
nature of life on other planets. Soon the Arrhenius hypothesis of inter-planetary 
travel of spores can at last be tested. Is life (i.e. systems undergoing natural 
selection) on other planets chemically entirely different from ours? Or is DNA 
the basis of all self reproducing systems, i.e. is the primacy of DNA a chemical 
necessity or a historical-biological accident? I should like to record here my 
personal hope that life on other planets will turn out to be of independent 
origin — this will make the coming science of space biology a lot more interesting. 



Crow: Darwin and the Origin of Life 55 

REFERENCES 

1. Darwin, Charles. 1877. The Effects of Cross and Self Fertilization in the Vegetable Kingdom. 
D. Appleton and Company: New York. 

2. Darwin, Charles. 1868. The Variation of Animals and Plants under Domestication. J. 
Murray: London. 

3. Darwin, Francis. 1887. Life and Letters of Charles Darwin. J. Murray: London. 

4. Darwin, Francis. 1903. More Letters of Charles Darwin. D. Appleton: New York. 

5. Galton, Francis. 1871. Experiments in Pangenesis, by Breeding from Rabbits of a Pure 
Variety, into Whose Circulation Blood Taken from Other Varieties had Previously been 
Largely Transfused. Proc. Royal Soc. 19:393-410. 

6. Darwin, Charles. 1871. The Descent of Man and Selection in Relation to Sex. J. Murray: 
London. 

7. Fisher, R. A. 1930. The Genetical Theory of Natural Selection. The Clarendon Press: Oxford. 

8. Hardin, Garrett. 1950. Darwin and the Heterotroph Hypothesis. Sci. Monthly 70:178-179. 

9. Haldane, J. B. S. 1933. The Origin of Life. In Science and Human Life. Harper: New York. 

10. Muller, H. J. 1921. Variation Due to Change in the Individual Gene. Amer. Natur. 56: 32^50. 

11. Muller, H. J. 1926. The Gene as the Basis of Life. Proc. Int. Cong. Plant Sci. 1:897-921. 

12. Sagan, Carl. 1957. Radiation and the Origin of the Gene. Evolution 1 1 :40-55. 



Tumors in Drosophila* 

WALTER J. BURDETTE 
Department of Surgery and Laboratory of Clinical Biology 
University of Utah College of Medicine, Salt Lake City, Utah 

Few students of genetics have failed to be intrigued by the hereditary mela- 
notic tumors in Drosophila, which have been known since the report of Bridges 
in 1916 (4) Stark (89-95) was responsible for the most extensive early work 
on them and drew an analogy between them and lymphosarcoma. Smce that 
time a number of workers have isolated and examined strains bearing tumors 
(2, 6, 10, 28-32, 38, 45, 51, 53, 56, 65, 83, 85-87, 89-96, 101, 104) which occur 
spontaneously and sporadically in diverse populations and species (8. 69, 70). 
Matings of individuals with these tumors often show them to be hereditary. Our 
investigations have utilized only those which are inherited, since induction of 
the tumors by injections (25-27, 50) is beset by the difficulty that in,ury m 
insects elicits a response which may be indistinguishable from the tumors (88). 

MORPHOLOGY OF THE TUMORS 

The melanotic tumors (55, 60, 77, 78, 97) appear very early in larval life, and 
those we have studied contain two types of cells. The first are large round, or 
polygonal cells, and the second are fusiform. The latter, elongated elements 
probably represent stroma of the tumor as Ghelelovitch (43, 44) has demon- 
strated in cultures. Melanin (54) is deposited quite soon, and the tumors are 
visible grossly as pigmented spots before the fifth instar. Complete dissolution ot 
cellular components leaves an inert pigmented residue to mark tumor-bearers 
during pupal and imaginal life. Sections indicate amelanotic tumors to be rare 
or non-existent, so that scoring in adults is an accurate assessment of tumor in- 
cidence in the absence of trauma. The tumors may appear m abdomen, thorax, 
or head but are most frequently encountered in the abdomen. Occasionally they 
are situated in the circulatory stream and present a mobility which is striking. 
Rizki (84) reports that lamellocytes may encapsulate other tissues to produce 
the appearance of tumors in certain strains. The m" (75, 76) and .r (47) tumors 
also have a different histologic appearance. King, Burnett, and Staley (62, bi) 
recently have described ovarian tumors in three strains of Drosophila hearing 
respective, female-sterile mutants. Although all tumors studied in our laboratory 
have been benign, Harnly, El Shatoury (28-30), and Ardashnikov (1) have 
called tumors malignant which have affected the viability of the individuals 
bearing them. Russell (85) demonstrated that the lethality of the lethal (1) / 
stock is due to obstruction of the alimentary tract rather than to the presence oi 
the tumor as originally suspected. 

♦ Aided by grants from the U.S. Public Health Service and the American Cancer Society. 



58 



The University of Texas Publication 



INFLUENCE OF HORMONES ON TUMOR GROWTH 

Since the unique regression which has been described is not the usual course 
of an atypical growth, two types of experiments were performed in order to 
determine the mechanism for this behavior. Genes which damage the ring gland 
were introduced into tumor stocks (19), and, in other experiments, larvae were 
hgated posterior to the ring gland so as to impede access of hormone to the pos- 
terior end of larvae (1 7). In both instances the numbers of tumors were greater 
when the amount of hormone was diminished. This evidence and that of others 
(64, 79) supports the hypothesis that the tumor cells are sensitive to meta- 
morphosis hormone and regress in a manner similar to other larval elements 
The opposing hormones, ecdysone and neotenin, can be assayed respectively 
and ecdysone has been crystallized both from Drosophila (20) and Bombyx (23' 
61). Regression of mammalian as well as insect tumors has been observed when 
ecdysone is administered, although results so far are neither uniform nor pro- 
portional to dosage (23) . 

INCIDENCE OF TUMORS 

A spectrum of tumor strains exists in Drosophila (10) with incidence varying 
from less than 1.0 per cent to more than 99.0 per cent (Table 1). The incidence 
of tumors in each strain is constant within limits, the variation being wider for 
a stram such as the tu g strain which has intermediate penetrance than for one 
such as the tu^'- strain which has low penetrance. The strain with the most con- 
stant mcidence under the usual conditions in our laboratory has been the 



Table 1 
Incidence of tumors in DrosoDhilc 



Strain 



Times 
Counted 



bw St tu 

tu36a* 
tu3*^a 

ed Su2-dx 

f 257-i9B/in AM 

tu^ps ■ 

lz3f 
Wbf f5 



bw tu 

se e^i tu^Qh 
tuh* 
tug* 
tu48j 



tuh 

vg mt^ bw 

y ^263-43 

tug 

tu48a yg |3^ 



7 

13 

9 

7 

7 

13 

13 

7 

15 

14 

13 

13 

12 

14 

13 

14 

7 

14 

14 



Months 
Counted 



Number with 
Tumors 



24 

222 

182 

385 

416 

1,423 

2,428 

715 

1,901 

2,434 

3,275 

2,421 

2,156 

2,833 

6,616 

5,944 

2,274 

9,113 

10,540 



Total No. 
Counted 



4,426 

7,473 

3,394 

4,022 

2,449 

8,077 

10,160 

2,827 

7,144 

8,614 

8,799 

5,464 

4,626 

5,865 

12,236 

10,069 

3,120 

11,967 

10,555 



Percentage with Tumors 



Males 



.26 
3.21 
5.10 
11.59 
15.53 
12.10 
28.53 
30.20 
29.60 
26.72 
38.11 
42.09 
49.19 
53.66 
50.98 
56.49 
69.92 
87.30 
99.77 



Females 



.84 
2.73 
5.66 
7.52 
17.87 
23.18 
18.59 
19.85 
23.31 
29,92 
36.41 
46.86 
43.81 
43.67 
57.69 
61.09 
75.77 
65.34 
99.94 



Total 



.54 

2.97 
5.36 
9.57 
16.99 
17.62 
23.88 
25.29 
26.61 
28.26 
37.22 
44.31 
46.61 
48.30 
54.07 
59.03 
72.88 
76.15 
99.86 



* Isogenic. 



Burdette: Tumors in Drosophila 59 

m^- vg bw strain obtained from Ghelelovitch (38). This inconstant expression 
of the tumor phenotype in genetically uniform strains is found m mammals as 
well as Drosophila (9) . 

EFFECT OF ENVIRONMENT ON GENE ACTION 

A series of tumor counts was made on eight strains of Drosophila over a period 
of six months under laboratory conditions as nearly uniform as possible with 
regard to temperature, culture medium, population size, and handling (24). 
Fluctuations of tumor incidence during this period apparently depended on the 
specific genes present in each individual stock since the fluctuation m numbers 
of tumors in the various stocks was neither similar nor predictable. In another 
series of experiments, isogenic tumor stocks were obtained by appropriate 
crosses with inversions to prevent crossing over (14). Even though each pair of 
alleles on the four chromosomes had a common origin, penetrance was not com- 
plete. The ambient milieu in conjunction with or without suppressor genes thus 
may prevent complete expression of the tumor phenotype. 

The effect of temperature on tumor incidence is quite striking in some stocks, 
whereas changes in temperature in others produces little or no effect (Table 1). 
For example, temperatures ranging from 15 to 30° C. fail to alter the mcidence 
in the tu''^- stock appreciably. On the other hand, flies of the tu^'^ vg bw strain 
raised at 30° C. had less than half the incidence of tumors found in the stock 
when raised at 20° C. and 25° C. respectively. The prolonged life cycle at lower 
temperatures did not necessarily raise the tumor incidence. Hartung (52) and 
Harnly, Friedman, Emery, and Glassman (49) found that response to tempera- 
ture is not uniform for all strains and varies with the tumor genes present. 
Ghelelovitch (40, 41, 45) extensively investigated the effect of temperature on 
the tu^'^ strain. Only the maternal effect was changed in the tu'^ strain when the 
temperature was varied by Gardner and Woolf ( 36 ) . 

An inverse relationship between the size of the population and tumor inci- 
dence has been reported by Herskowitz and Burdette (59) and Wilson (101). 
This effect is probably related to nutritional state of the larvae. In general, a 
reduction in the amount of yeast available has resulted in a concomitant dim- 
inution in tumor incidence, and certain stages in development are more sensi- 
tive to this effect than others, e.g. the third day of larval life in the tu'''^ strain. 

Table 2 
Effect of temperature on incidence of tumors 



15° C. 



20° C. 25° C. 30° C. 

^^. n 637 1.7 1.091 1.6 1,271 2.7 526 

■^^ 90.5 558 99.5 1,281 11.2 1,118 9.5 84 

i^ps 20 3 118 14.4 1,122 19.7 1,220 0.0 60 

^^^i^ 72.0 93 95.1 595 82.3 1,113 

^tAbw 74.5 47 93.2 1,347 82.3 1,073 14. 288 

tu^sa^gbw 98.7 1,142 99.9 1,263 42.2 64 



60 



The University of Texas Publication 



In contrast to the results with Saccharomyces, Briones and Brncic (5) found that 
increasing the content of Torulopsis utilis in the medium diminished the number 
of flies with tumors. The reason for divergent resuhs with different yeasts is not 
clear. Caloric restriction in mammals may also reduce the number of susceptible 
individuals with tumors as well as prolonging survival (98-100) . Extensive work 
on adding and deleting specific dietary constituents has been done by a number 
of investigators (33, 34, 39, 42, 49, 57, 72, 74). Prolongation of the larval stage 
by administration of 2, 4-dinitrophenol delayed the appearance of tumors in 
experiments of Wilson ( 1 02 ) , but contradictory results have been obtained when 
the effect of various amino acids on the incidence of tumors has been tested (46, 
58, 71, 73, 80, 81, 102, 103). The action of vitamins on tumor appearance was 
inhibited by the appropriate analogue according to Friedmann, Harnly, and 
Kopac (35). 

LOCALIZATION OF TUMOR GENES 

The pattern of inheritance of the melanotic tumors is easily followed if sus- 
ceptibility to tumors is scored rather than the actual incidence of tumors. Exten- 

Table 3 
Chromosomal distribution of genes associated with tumors or focal melanosis in Drosophila 



in IV 



1(1)7* 1(1)76^ 
1-t* 

l(l)ml* 
1(1)76* 



me* 
f^I*~ 





tus 








tu^8j* 








mtA* 








eiitu* 








tu50j* 








fes* 








nw* 


md* 






1 


tuwps 




tuh 


tuh 


tuh 






tui 


tui 






tu49h 


tu49h 






tu 


su-tu 






su-er 


er 




tu36a 


tu36a 


tu36a 




tu49k 


tu49k* 


tu-i9k 




tu^ 


tu^ 


tu^ 




tu^Sa yg ]3^ 


tu48a yg b^* 




tu48a yg |3^ 


be(3) 


be (3)* 


be(3)* 


be (3) 


tu2 


tu2 


tu2 


tu2 











Symbols underlined: Main genes for susceptibility. 
Symbols designated*: Localized within chromosome. 



Burdette: Tumors in Drosophila 61 

sive linkage studies have been reported for mammalian tumor genes, but locali- 
zation along the chromosome is much more precise in Drosophila (Table 3) . The 
number of tumor genes present in a strain may be one or several, and they may 
be present on one, two, three, or four chromosomes. The contribution of each 
gene also varies, and both enhancer and suppressor modifiers have been found. 
The suppressor-erupt-tumor system studied by Glass and Plaine (47) and the 
modifiers for tu"^ found in aUen strains by Gardner, Scott, and Dearden (37) 
are good examples. For reasons unknown, many of the main tumor genes are 
present on the second chromosome. However, only occasionally are alleles found 
when strains of different origin are compared. An example is the presence of 
at least one common gene in m^- vg bw and tu^^ (12) strains. More than one 
third of the stocks have known modifiers in addition to the mam genes, and more 
than one main gene has been reported for three of them. Both the benign tumor 
studied by Stark and Bridges (93) and the tu-2 stock studied by Wilson (101) 
have at least one susceptibility gene present on each of the four chromosomes. 
Although tumors arise in the presence of a single mutant on either chromosomes 
I and III, an impressive number of main genes are located on chromosome II. 
Several of the tumor genes have been localized within narrow limits (Table 4). 

Table 4 
Examples of localized genes for tumor susceptibility 



Chromosome I 


Chr 


3mosome 


II 


Chromosome III 


Tumor 


Locus 


Tumor 




Locus 


Tumor Locus 


1(1)7 


0.1 


fes 




5 


be3 25 


fu 


59.5 


tu48a vg 


bw 


29.5 








nw 

e^itu 

tu"j 

be3 




44.3-55.7 
82.0-82.7 
83. 
88. 
90. 
107. 





Recombination of tumor genes has been more difficult to follow than for many 
visible mutants, but no evidence has been found that tumor pseudoalleles exist 
Since the germinal tumor mutants could be either point mutation or associated 
with chromosomal aberration, it is of interest to determine in this favorable 
biological material which type of change has taken place in the chromosome 
leading to the occurrence of tumors. Salivary chromosomes have been examined 
under both light and electron microscopes, and no evidence of chromosomal 
aberrations have been found in the region of tumore genes which have been 
localized by crossing over (16). In certain instances, tumors have appeared m 
strains with duphcations and inversions, but these pre-existing chromosomal 
aberrations are the only ones which have been observed in the salivary gland 
preparations. Although this study has not been exhaustive, being confined only 
to the tu^'j and vg mt^ bw stocks, this evidence is sufficient to show that the tu- 
mors may appear without visible chromosomal rearrangement. 



^2 The University of Texas Publication 

Since the time of Boveri (3), great interest has centered around the possible 
relationship between aneuploidy and cancer. Therefore, the addition of euchro- 
atin and heterochromatin to the normal chromosomal complement to test the 
effect of changes in gross amounts of chromosomal material within the cell is 
of interest, and studies have been completed on the effect of additions and dele- 
ions of heterochromatic material to cells containing tumor genes in Drosophila. 
This was done by the use of the tandem XY chromosome. Through appropriate 
crosses, the effect of varying multiples of the Y chromosome, which contains little 
euchromatin in Drosophila, was studied. The results indicated that there was no 
consistent change in tumor incidence, the amount and direction of the change 
depending on the tumor genes present (24) . 

MUTATION AND THE ORIGIN OF SUSCEPTIBILITY TO TUMORS 

A recurrent idea about the origin of cancer is that a mutation occurs in a 
somatic cell. Since mutations to tumor susceptibility occur in germ cells, the 
possibility that this may lead to cancer when it occurs in a somatic cell cannot be 
denied. However, the event is uncommon in germ cells, and the frequency with 
which it occurs in somatic cells is open to question. It would seem reasonable to 
require mutagenic agents to be carcinogenic and carcinogenic agents to be 
mutagenic should this hypothesis be true. Using the lethal mutation rate on the 
X chromosome as an indicator, both mutation rate and tumor incidence were 
determined simultaneously in the tu^'a strain. When 20-methylcholanthrene 
was administered as an aerosol, the incidence of tumors was increased in the F^ 
generation, but the mutation rate was unchanged significantly (13). On the 
other hand, nitrogen mustard both augmented mutation rate and tumor inci- 
dence (15). The mutation rate in males was increased when formaldehyde was 
fed to larvae, but the tumor incidence remained unchanged in both sexes (11) 
m contrast to the sex difference in lethal mutation rate after treatment with 
formaldehyde. Although negative results are always questionable, the fact that 
mutation rate and tumor incidence were determined simultaneously and one was 
increased with an absence of any effect on the other in the case of methyl- 
cholanthrene and formaldehyde, casts considerable doubt on the idea that the 
neoplastic change is very often the result of a somatic mutation. 

Should somatic mutation be the sole reason for the appearance of tumors, 
the germinal mutants which raise tumor incidence would of necessity be 
mutators. The effect of a mutator was tested in several tumor strains, and no 
relationship between the presence of mutators and an increase in tumor inci- 
dence was found (18). Florida stocks were used for these studies, and fluctuations 
m the mutation rate during the course of experiments made interpretation 
additionally difficult. 

One of the ways in which a neoplastic change could result in an inherited 
change m somatic cells would be through the loss of a gene or enzyme system. 
In order to test whether the germinal tumor mutants consist of a simple loss of 
substance, the tu^''^ vg bw strain was subjected to treatment with both a chemical 
mutagen, formaldehyde, and X irradiation. Progeny were then tested for re- 
verse mutation toward normal. The experiments with formaldehyde were nega- 



Burdette: Tumors in Drosophila 63 

tive with no reversions in 34,331 chromosomes tested. However, when the 
mutation rate was increased approximately threefold by using X irradiation m 
dosage of 2,000 r, two reversals were found in 76,483 tests. Subsequent crosses 
suggest these to be true reversions and not due to the appearance of new 
suppressor mutants. These findings indicate that the structural change which 
produces a tumor mutant may be different than simple deletion and is 
reversible. 

TUMORS AND GENOIDS 

Experiments within recent years on the etiology of leukemia have indicated 
that a filterable agent is of prime importance in the appearance of the disease 
(48) despite previous genetic and radiologic studies to the contrary. This has 
stimulated considerable interest in this facet of the etiology of cancer. The only 
cytoplasmic particle easily available for study in Drosophila melanogaster is 
the genoid for CO.3 sensitivity (66-68, 82). Chromosomes containing tumor 
genes were introduced into cytoplasm containing the genoid by genetic methods, 
using inversions to prevent crossing over, as well as the introduction of the 
genoid through injections into cytoplasm containing chromosomes carrying 
tumor genes. The forte, faible, and Tr strains of the genoid were used in these 
studies. It was found that the presence of genoid in the tu''^ vg bw strain resuUed 
in a diminished incidence of tumors (21, 22). When loss of sensitivity occurred 
in some of the individuals used in the investigation, it was found that the 
lowered tumor incidence remained in the offspring despite the disappearance of 
sensitivity. Whether the presence of the particle in the cytoplasm alters the ex- 
pression of tumor mutants in a permanent manner is an interesting question for 
speculation. Although the plasmagene theory of tumorigenesis has been distin- 
guished chiefly by the absence of experimental data to support the assumption, 
the utilization of viruses with known effects within the cell in conjunction with 
known tumor mutants would seem a profitable future avenue of approach, and 
these resuhs in Drosophila indicate positive results will be forthcoming from the 
combination of genes and genoids. 

SUMMARY 

Hereditary tumors in Drosophila have proved to be a valuable means for 
investigating some of the factors involved in the origin of tumors, but, even in 
this relatively simple experimental animal, with brief life cycle and small 
chromosomal complement, the mechanism is somewhat compHcated (Figure 1). 
The appearance of an atypical growth is naturally modified by the characteristics 
of the organism in which it is encountered, and Drosophila is no exception. The 
tumors studied are composed of aggregates of hemic cells with related stroma, 
and their persistence is modified by the hormones controlling metamorphosis. 
Ecdysone is apparently responsible, at least in part, for their unique regression. 
Susceptibility mutants, single and multiple, are responsible for the appearance 
of the hereditary tumors, and penetrance depends on the relative effectiveness 
of a number of different conditions prevailing during the period of genie action. 
The same environment change may result in different effects on different genes. 



64 



The University of Texas Publication 



Alkyloting cH,-N<^e"H'cHfc. 
Agents * 




f I. Heterochromotin oneuploidy ) 
2. Viral infection 
3 Ambient milieu 
Temperature 



Fig. 1 



In general, certain chemicals such as alkylating agents and carcinogenic hydro- 
carbons may increase the incidence of tumors, whereas caloric restriction and 
suppressor mutants may reduce the incidence of tumors. No general correlation 
between agents increasing incidence of tumors and mutation rate has been 
found, and the germinal mutation to tumor susceptibility is not necessarily a 
deletion or visible chromosomal aberration. The minute localization and spe- 
cificity of this chromosomal change in Drosophila demonstrated by genetic 
studies have sufficient prognostic significance for urging refinement of techniques 
in other organisms in order to discern which intracellular events lead to neo- 
plastic transformation. 

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Burdette: Tumors in Drosophila 67 

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71. Mittler, S. 1952. Influence of amino acids upon the appearance of tumors in tu^^^j stock of 
Drosophila melanogaster. Genetics, 37:606. 

72. - . 1952. Influence of nutrition upon appearance of tumors in tu^'^' stock of 

Drosophila melanogaster. Science, 115:271-272. 

73. . 1952. Nucleic acids and tumor production in tu^oj stock of D. melanogaster. 

Anat. Rec, 113:606. 

74. . 1954. Influence of vitamins upon incidence of tumors in tu^c^ stock of D. 

melanogaster. Science, 120:314. 

75. Newby, W. W. 1948. Abnormal growths on the head of Drosophila melanogaster. J. 
Morphol., 85:177-195. 

76. and R. P. Thelander. 1950. Early development of the head in normal and 

tumorous head D. melanogaster. D.I.S., 24:89-90. 

77. Oftedal, P. 1952. Histology and histogenesis of Drosophila tumors. Science, 116:392-393. 

78. . 1953. The histogenesis of a new tumor in Drosophila melanogaster and a com- 
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79. Oster, I. I. 1954. Factors bearing on the non-malignacy of tumors in Drosophila melano- 
gaster. Cancer Research, 14:478-481. 

80. Plaine, H. L. 1955. The counteraction by cysteine of the effects of X-rays and of tryptophane 
on the action of specific suppressor system in Drosophila melanogaster. Cancer Research, 
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81. . 1955. Influence of tryptophane and related compounds upon the action of a 

specific gene and the induction of melanotic tumors in Drosophila melanogaster. J. Genetics, 
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82. Plus, N. 1954. Etude de la multiplication du virus de la sensibilite au gaz carbonique chez 
la drosophile. Bull. Biol. France et Belg., 88:248-293. 

83. Rapoport, J. A. 1938. Hereditary lethal tumors in Drosophila. Bull. Biol. Med. exp. 
(U.R.S.S.). 6:725-728. 

84. Rizki, M. T. M. 1957. Tumor formation in relation to metamorphosis in Drosophila melano- 
gaster. J. Morph., 100:459-472. 



mmm 



68 The University of Texas Publication 

85. Russell, E. S. 1940. A comparison of benign and "malignant" tumors in Drosophila melano- 
gaster. J. Exp. ZooL, 84:363-379. 

86. . 1942. The inheritance of tumors in Drosophila melanogaster , with special 

reference to an isogenic strain of st sr tumor 36a. Genetics, 27:612-618. 

87. Scharrer, B. and M. S. Lochhead. 1950. Tumors in the invertebrates: A review. Cancer 
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88. Schlumberger, H. 1952. A comparative study of the reaction to injury. I. The cellular 
response to methylcholanthrene and to talc in the body cavity of the cockroach (Periplaneta 
americana). Arch. Path. 54:98-113. 

89. Stark, M. B. 1918. An hereditary tumor in the fruit fly, Drosophila. J. Cancer Research, 
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90. . 1919. A benign tumor that is hereditary in Drosophila. Proc. Nat. Acad. Sci., 

Washington, 5:573-580. 

91. . 1919. A benign tumor in Drosophila. Proc. Soc. Exp. Biol. Med., 17:51-52. 

92. . 1919. An hereditary tumor. J. Exp. ZooL, 27:509-521. 

93. and C. B. Bridges. 1926. The linkage relations of a benign tumor in Drosophila. 

Genetics, 11:249-266. 

94. . 1935. An hereditary lymphosarcoma in Drosophila. Collected Papers N. Y. 

Med. Coll., and Flower Hosp., 1:397-400. 

95. . 1937. The origin of certain hereditary tumors in Drosophila. Am. J. Cancer, 

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96. Stott, G. H. and E. J. Gardner. 1952. The location of the two major genes responsible for 
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97. Strunge, T., D. Gigante, and W. Bernhard. 1951. Hemopathie de type leucemique chez la 
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98. Tannenbaum, A. 1942. The genesis and growth of tumors. II. Effects of caloric restriction 
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99. . 1944. The dependence of the genesis of induced skin tumors on the caloric 

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100. . 1944. The dependence of the genesis of induced skin tumors on the fat content 

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101. Wilson, I. T. 1924. Two new hereditary tumors in Drosophila. Genetics, 9:343-362. 

102. Wilson, L. P. 1947. Effect of dinitrophenol and excess amino acids upon melanotic growths 
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103. . 1949. Increased incidence of a tumor in Drosophila in the presence of high 

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melanogaster. Growth, 19:215-244. 



Sex Balance in Drosophila melanogaster : Aneuploidy of Short 
Regions of Chromosome 3, Using the Triploid Method 



SARAH BEDICHEK PIPKIN 
Howard University 



The present study is the outcome of previous investigations on sex determina- 
tion in Drosophila melanogaster initiated and carried out by Professor J. T. 
Patterson and his colleagues and students. Beginning with his work on X-ray 
induced gynandromorphs, mosaic females, and "aberrant" (i.e., hyperdiploid) 
males and females, Patterson was interested in learning if a primary female sex 
factor exists in the X chromosome (Patterson, 1931, Patterson and Stone, 1938). 
By further studies of hyperdiploid males derived from overlapping X,4 trans- 
locations according to a method described by Muller and Stone (1930), the pos- 
sibility of a major female sex factor was reduced to a short section extending from 
salivary bands 13A2 to 13A6, the g-pl region (Patterson, Stone, and Bedichek, 
1937; Patterson, 1938). Crow (1946) finally ruled out a possible X chromosome 
primary female sex determiner by synthesizing a diploid male hyperploid for 
this region. None of the X chromosome hyperdiploid males studied by these in- 
vestigators was intersexual. An occasional rotation of external genitalia was not 
taken as a sign of intersexuality in X-hyperdiploid males since this rotation is 
sometimes found in diploid males hyperploid for a short autosomal region. Con- 
versely, no diploid female hypoploid for a section of the X chromosome dis- 
played any intersexual features. Mickey (1950) briefly described some diploid 
intersexes arising in an attached-X stock in which detachment had occurred. The 
intersexes were thought probably to be diploid aneuploids with one X chromo- 
some and a fragment of an X, but it was not possible to make cytological studies. 

Meanwhile, Dobzhansky and Schultz (1934) and Pipkin (1940) were able 
to transform triploid intersexes into weakly functional hypotriploid females by 
the addition of long fragments of the X chromosome to the 2X3A intersex com- 
plement. These investigators agreed that short sections of the X in some cases 
shifted the sex type of "duplication" intersexes in the female direction. By the 
overlapping translocation method, Pipkin found that individuals bearing short 
X chromosome sections, covering in succession the entire chromosome, when 
added to the 2X3 A intersex chromosome set are only intersexual flies. Longer 
and longer fragments from either the left or right hand end of the X chromosome 
cause a qualitatively progressive shift in the direction of femaleness until either 
the right or left hand fragment of two different translocations, 17 (broken be- 
tween 7F5 and 8B1 ) and wl3 (broken between 9B1 and 9B3), when added to the 
2X3 A intersex chromosome set, produce weakly functional females (Pipkin, 
1940). A shorter left than right hand fragment is required to produce the 
complete sex shift from intersexuality to femaleness. 

The weakly fertile hypotriploid female with the right hand X fragment of 
X,4 translocation 17 or wl3 retained rudiments (one or two prongs) of male sex 



70 The University of Texas Publication 

combs in some individuals when reared at 23° C. When reared at 18° C, these 
17R or wl3R plus 2X3 A individuals were all sterile, more intersexual in bodily 
appearance, and developed larger sex combs (up to six prongs). Furthermore 
hy pointer sexes lacking a short portion of one of their two X chromosomes, pos- 
sessing three of each autosome, were shifted strongly in the male direction. Thus 
triploid aneuploid studies of the X chromosome show that several parts of the X 
chromosome are concerned with female differentiation and the effects are 
additive. 

Pat+erson, Brown, and Stone (1940) investigated diploid aneuploidy of the 2nd 
and 3rd chromosomes. They succeeded in producing hyperdiploid males and 
females carrying in triplicate each of eight short regions covering successively 
the entire chromosome 2 and eleven regions, covering all but the leftmost short 
section, of chromosome 3. Hypodiploid males and females were obtained for five 
short sections of chromosome 3 but for none of the 2nd chromosome sections. No 
sex shift was found in either autosomal hyperdiploid females or in the hypodip- 
loid males for five regions of chromosome 3 studied. Burdette (1940) studied a 
homozygous duplication female bearing a short heterochromatic region just to 
the left of the centromere of chromosome 3 in quadruplicate, but no change in 
the sexual differentiation of these females was observed. 

With the same 2,4 translocations used by Patterson, Brown, and Stone (1940), 
Pipkin (1947) studied triploid aneuploidy of chromosome 2 in an effort to de- 
termine the role played by this chromosome in sex determination. No shift 
toward maleness w^as observed in hyperinter sexes (2X3A + fragment of 2) for 
each of ten short regions covering successively the whole of chromosome 2. Sim- 
ilarly, hypertriploid females (3X3 A + fragment of 2) showed no signs of inter- 
sexuality. Hypointersexes (2X3 A — fragment of 2) were obtained in numbers 
sufficient for a statistical analysis of the sex types for one region only, and no 
sex type shift in hypointersexes as compared with control sibling intersexes 
(2X3A) was found for this region. Hypointersexes for six other short regions 
and three of the longer regions showed no qualitative shift in a female direction; 
i.e., they appeared intersexual. 

Using the overlapping translocation method, Dubovsky (1939) studied the 
effect of increasingly longer sections of the left arm of chromosome 3 upon dip- 
loid aneuploids. Hypoploids for only one very short region survived. Hyperploids 
for 5 regions included between 61 and 67B lived, but hyperploids for three longer 
regions failed to survive. Patterson and Stone (1952) summarized their views on 
sex determination based on evidence from diploid and triploid aneuploidy. 

Two different investigators have described changes in sexual differentiation 
owing to hypoploidy of the 3rd chromosome. In both cases the deficient regions 
were minimal in length, and the second case combines hypoploidy with the 
action of a mutant. 

Kelstein (1938, 1939), also using the overlapping translocation method, 
studied minute duplications and deficiencies (diploid aneuploids) of a section 
of the right arm of chromosome 3 extending from salivary bands 89A to 94A by 
means of 18 different 3,4 translocations broken in this short section. The indi- 
vidual duplications and deficiencies studied extend only a few salivary bands in 
length and are thus much shorter than those studied by Patterson, Brown, and 



Pipkin: Sex Balance in Drosophila 71 

Stone (1940) in diploid aneuploids and by Pipkin (1947 and this paper) in 
triploid aneuploids. Kelstein described hypoploids for the section from 89D to 
90 A which were female in appearance but possessed male sex combs (the size of 
the sex combs was not given). Hypodiploid males for this region were described 
as invariably having rotated genitalia. No intermediate intersexual forms were 
described (i.e., there was no mention of fragmentation of the male genitalia or 
change in anal plates or of forms with abnormal female genitalia). The author 
does not make it clear if the female-like intersexes were IX or 2X. No cytological 
examinations were described, and no dissections were made to study the internal 
genital tract. Kelstein hsts 130 hypoploid males and 163 hypoploid females for 
this region (XV), but he does not enumerate the number of female-like inter- 
sexes. No doubt is here cast upon the resuhs as reported especially since 
other aneuploid data are in agreement with what has been described by other 
investigators. 

Intersexuality has also been described as resulting from the combination of 
diploid hypoploidy with a mutant. Goldschmidt (1948, 1949a, 1949b, 1949c) 
found low-grade intersexuality in males carrying the 3 chromosome dominant 
mutant Beaded together with any one of several different minutes (tiny de- 
ficiencies) which could be located on either the 2nd or 3rd chromosomes. Neither 
Beaded nor the minute alone produced intersexual effects. The Beaded-Minute 
intersexes were interpreted by Sturtevant (1949) as representing incomplete de- 
velopment of the anal and genital imaginal discs. On the other hand, Gold- 
schmidt (1949c) presented drawings and descriptions of the internal genitaha of 
the Bd-Mn intersexes showing what looks very like seminal receptacles and in 
one specimen, a ventral receptacle and rudimentary vagina. He also pointed out 
that a reduction of the genitalia and anal plates is likewise characteristic of type 
III triploid intersexes. Partly from his work on the Beaded-Minute intersexes, 
Goldschmidt (1955) concludes that "sex determination is actually a function of 
intercalary heterochromatin, a generalized function, not capable of being dis- 
solved into sets of multiple genes with special actions," since Beaded is a hetero- 
chromatic mutant and the minutes are deficiencies in heterochromatic regions. 

A number of autosomal mutants causing intersexuality in diploid (2X2A) 
females but usually not affecting 1X2A males occur in several species of Dro- 
sophila. A recessive mutant in both the 2nd chromosome (the ix mutant, Mor- 
gan, 1943, and its allele, ix\ Meyer, 1959) and in the 3rd chromosome (the tra 
nmtant, Sturtevant, 1945, as well as the dominant allele of tra called //r, Fung 
and Gowen, 1956, 1957) have been described in D. melanogaster . Sturtevant 
(1920) found a recessive intersex mutant in the 2nd chromosome of D. simulans. 
Lebedeff (1939) studied a recessive mutant, ix, in D. virilis, and Newby (1942), 
a dominant mutant, Ix^, not allelic with Lebedeff' s mutant, in the same species. 
Dobzhansky and Spassky (1941) reported such a recessive mutant in D. pseudo- 
ohscura, and, similarly, Hollingsworth (1953) and Spurway and Haldane 
(1954) in D. suhohscura. 

Species hybrids in Drosophila which yield intersexes have been described in 
the repleta group. Cases summarized by Patterson and Stone, 1952, include those 
of Wharton, 1940; Sturtevant, 1946; and Ward and Stone, 1952. In Sturtevant's 
case, an autosomal gene in D. neorepleta was found, capable when present in one 






72 The University of Texas Publication 

dose in a female hybrid between this species and D. repleta of conditioning her 
eggs in a male direction so that the resulting zygotes with two repleta X chromo- 
somes developed into intersexes. 

Bridges' (1921, 1922) discovery of triploid intersexes in D. melanogaster pos- 
sessing two X chromosomes and three of each autosome led him to assign the 
male determining genes to the autosomes. Dobzhansky and Bridges (1928) ruled 
out a conection of the tiny fourth chromosome with sex determination since it 
could be present in single, double, or triple dose in diploids, and its dosage did 
hot affect the sex type of triploid intersexes. The absence of a Y chromosome like- 
wise was long ago found not to affect the sexual differentiation in XO non- 
dis junctional males though these are sterile (Bridges, 1922). Dobzhansky and 
Schultz (1934) concluded that the Y chromosome does not influence the sex 
type of triploid intersexes since the mean sex type of Y 2X3A intersexes does not 
differ significantly from that of sibling 2X3A intersexes in the material studied 
by them (Dobzhansky and Schultz, 1934) . 

As in D. melanogaster, autosomes must play a male-determining role in other 
species of Drosophila. Triploid intersexes similar to those found in D. melano- 
gaster have been reported in D. americana (where motile sperm are found in the 
extreme male type) (Stalker, 1942); D. simulans (Neuhaus, 1939); as hybrids 
between D. melanogaster and D. simulans (Schultz and Dobzhansky, 1933) ; D. 
pseudoobscura and as hybrids between D. pseudoobscura and D. miranda (Dob- 
zhansky and Spassky, 1941). 

The purpose of the present investigation is to complete the triploid aneuploid 
studies of the large autosomes by studying sex balance in the 3rd chromosome. 

MATERIALS AND METHODS 

The 3,4 translocation stocks used in the present experiments with the salivary 
map points of breakage in chromosome 3 include the following: T(3,4) e (Dob- 
zhansky, 1929; Lewis, 1956), broken at 79E; T(3,4) c (Dobzhansky, 1929; 
Lewis, 1951), broken at 86C; Ta(3,4) 13 (Patterson, et al, 1934; Patterson, 
Brown, and Stone, 1940), broken at 67E; Ta(3,4) 12 (Patterson, et al, 1934; 
Patterson, Brown, and Stone, 1940), broken 73C; Ta(3,4) 2 (Patterson, et al, 
1934; Patterson, Brown, and Stone, 1940), broken at 94A3-4; Ta(3,4) 28 (Patter- 
son, et al., 1934; Patterson, Brown, and Stone, 1940; Lewis, personal communica- 
tion), broken at 94D3-4; Ta(3,4) 30 (Patterson, et al, 1934; Patterson, Brown, 
and Stone, 1940), broken at 96E; T(3,4) 85C (Lewis, personal communication), 
broken at 85C; T(3,4) 89E (Lewis, personal communication), broken at 89E; 
T(3,4) H7, broken at 66C; T(3,4) HI, broken in the chromocenter; T(3,4) H3 
broken in the chromocenter; T(3,4) H5, broken at 96E; and T(3,4) H6, broken at 
98A. The T(3,4) H stocks were recently produced by X-rays at Howard Univer- 
sity and their points of breakage here determined for the first time. The older 
Ta(3,4) stocks were checked anew in salivaries by the author to be sure that they 
were the original stocks as labeled. 

The triploid stock developed for these studies had free X chromosomes and con- 
tained no known chromosomal abnormalities. The recessive mutant y^ (yellow 
body with black bristles) marked the X chromosome; ru (roughoid), the left end 



Pipkin: Sex Balance in Drosophila 73 

of chromosome 3; and ca (claret), the right end of chromosome 3. The character 
ru proved to give no difficuUies in classification since it is not only far more pro- 
nounced in its disturbance of facets in 3A flies than in 2A flies, but also the entire 
eye is markedly narowed in triploids and intersexes whereas it is only slightly 
narrowed in most diploid flies. The homozygous y'; ru ca triploid stock was 
rather weak, though not highly inbred. It had to be made up twice during the 
course of the experiments, and it was necessary to use 40 triploid females to one 
bottle in experimental crosses in order to keep down mold infections. 

All of the experimental crosses developed in a B.O.D. incubator; most of the 
work being done at 22° ± 0.5° C. Some special crosses were carried out at 26° ± 
0.5° C. Pans of water were kept in the B.O.D. incubator to keep the relative 
humidity about 75-80%. 

To study aneuploids for either the right or left end of the 3rd chromosome, 
males carrying a 3,4 translocation broken near an end of 3 and bearing the 
normal alleles of ru and ca in the broken 3rd chromosome, also heterozygous for 
an intact 3rd chromosome bearing the marker mutants ru and ca were crossed 
with homozygous y'; ru ca triploid females. A diagram illustrating this type of 
experimental cross appears in Figure 1. For example, in the case of a transloca- 
tion with a break at the right end of chromosome 3, hyperintersexes appear 
either y'; ru or simply ru. Members of the first group, appearing y'; ru, receive 
their 2 y- X chromosomes from the triploid mother, a Y chromosome from the 
translocation male parent, and possess three intact ru ca chromosomes plus the 
right hand fragment carrying the normal allele of ca. The second group of inter- 
sexes, appearing ru, is similar to first with respect to 3rd chromosome content but 
such flies possess one wild-type X chromosome derived from the translocation 
male parent and one y^ X chromosome from the triploid mother. Similarly, hyper- 
intersexes bearing the left hand fragment of a translocation broken near the left 
end of the 3rd chromosome appear y'; ca or simply ca. Hypertriploid females 
carrying a right hand end fragment in excess of 3X3 A appear ru; hypotriploid 
females lacking one dose of this region from the 3X3A complement appear ca. 
Conversely, hypertriploid females carrying a left hand end fragment in excess of 
3X3A appear ca; hypotriploid females lacking one dose of this region from the 
3X3 A complement appear ru. The genetic composition of one group of hyper- 
intersexes and hypertriploid females involving a right hand end region is dia- 
gramed in Figure 2. 




ca 



TRIPLOID 
FEMALE 



Fig, 1 Experimental cross designed to produce aneuploids involving end region of chromo- 
some 3. 



74 The University of Texas Publication 

Aneuploids for interior regions of chromosome 3 were obtained by the over- 
lapping translocation method. Homozygous y^;ru ca triploid females were crossed 
with males heterozygous for two different 3,4 translocations with neighboring 
points of breakage. One translocation carried the recessive marker mutant ru and 
the normal allele of ca; the second translocation, with point of breakage to the 
left of the former, carried the recessive marker mutant ca and the normal allele 
of ru. Such a cross is diagramed in Figure 3. Hyperinter sexes appearing in the 
progeny are either ru ca or f; ru ca, depending on the source of the X chromo- 
somes. Similarly, the hypointersexes are either wild-type or /. Hypertriploid 
females appear ru ca; hypotriploid females, wild-type. The genetic composition 
of one group of hyperintersexes and hypointersexes, and also that of hypertrip- 
loid females appear in Figure 4. 

In some cases the male parents carried ru in the 3,4 translocation broken to 
the left and ca in the 3,4 translocation broken to the right. Here the hyperinter- 
sexes appeared y^ or wild-type; the hypointersexes, y^; ru ca or ru ca; hypertrip- 
loid females, wild- type; hypotriploid females, ru ca. 

3x3A+Section 
HYPER-TRIPLOID FEMALE 

HI 




2x3A+Section 
HYPER- INTERSEX 

X 




lER 
Fig. 2. Genetic composition of ( 1 ) a hypertriploid female carrying an end region of chrome 
some 3 in excess of 3X3 A and (2) of one group of hyperintersexes for an end region. 



^ MALE 




Y _ ca 

HELb niERb 




TRIPLOID 
FEMALE 



Fig. 3. Experimental cross designed to produce aneuploids involving an interior region of 
chromosome 3. 



Pipkin: Sex Balance in Drosophila 



75 



2x3A + Section 
HYPER-INTERSEX 

ru^ CO 

X 




niLLa 



niERb 



2x3A-Section 

HYPO-INTERSEX 

ru 

X ru 




mLb 



HERa 



3x3A + Section 
HYPER-TRIPLOID FEMALE 

coin: 




HIRb 
Fig. 4. Genetic composition of (1) one group of hyperintersexes, (2) one group of hypointer- 
sexes, and (3) a hypertriploid female involving an interior region of chromosome 3. 

Hyper- or hypoploidy of the diploid progeny of f; ru ca triploids and males 
heterozygous for one or two different translocations was determined similarly, 
according to the recessive marker mutant(s) displayed. Diploidy was distin- 
guished from triploidy (or the 2X3A intersex condition) by the wmg texture 
which is fine in diploids and the eye facets which are smaller in diploids than 
the eye facets of the 3 A forms . 

Separate counts were made from each bottle of an experimental cross so that 
homogeneity tests could be made to determine if genetic modifiers were suffi- 
ciently variable in the triploid or translocation stocks to shift the sex types of 
control intersexes (2X3A) hatching in different bottles of the same experimental 
cross, the temperature being kept constant. 

EXPERIMENTAL RESULTS 

1 . Survival of 3rd chromosome aneuploids. 

Survival of 3rd chromosome aneuploids for short regions is summarized in 
the diagram presented in Figure 5. The top line in Figure 5 gives the salivary 
map location of the points of breakage of the 3,4 translocations used in the study. 
The stock numbers of these 3,4 translocations are placed near the bottom line m 
Figure 5. Survival of hyperintersexes, hypertriploid females, and hyperdiploid 



■ 



7^ 



The University of Texas Publication 



SALIVARY LOCATION 

61 66C 67E 



73C 



CENTf^OMERE 

79E 80C 81 F 85C86C 89E 



94A94D96E98AI00 



GENOTYPE 




2x3A-Sectlon 
1x2 A -Section 



H7 13 
PATTERSON 



89E 



2 e HI,H3 85C' 

I _i , I 

GOWEN a FUNG KELSTEIN 

STURTEVANT PATTERSON 



2 28 30, H 5 
I — J 



GOLDSCHMIDT 



Fig. 5. Diagram summarizing survival of various 3rd chromosome aneuploids. 



females, respectively, for each short region, is indicated in the upper section of 
Figure 5. Survival of hypointersexes and hypodiploid males is indicated in the 
lower section of Figure 5. If a single region possessed strong male-determining 
potency, the phenotype of the former group would be expected to be shifted in 
a male direction. Similarly, the phenotype of the latter group would be expected 
to be shifted in a female direction. According to Figure 5, both hyperdiploid 
females and hypertriploid females survived for each region studied. None showed 
any intersexual features. Hyperintersexes survived for each region with the 
possible exception of the region between Ta(3,4) 12 and the chromocenter, a 
point to be discussed later. In all cases hyperintersexes and surviving hypointer- 
sexes appeared typically intersexual. According to the lower section of Figure 5, 
hypointersexes and hypodiploid males for the left hand end region and hypo- 
diploid males for the right hand end region did not live. Further, hypointersexes 
for the interior region 12-e and hypodiploid males for interior regions 13-12, 
possibly 12-chromocenter, 89E-2, and c-89E failed to survive. It should be noted 
that the region 30-H6 was not directly tested owing to the difficulty of introduc- 
ing the marker ca into Ta(3,4) 30. Instead this section is represented in Figure 5 
as yielding hypodiploid males and hypointersexes because hypodiploid males and 
hypointersexes for the longer region 2-H6 survive, as may be determined from 
Table 1. Only one specimen of a hypointersex for region H6R was found. A 
statistical analysis of the sex types of hyperintersexes and hypointersexes, in 
cases where these survived in large enough number will be considered in a latter 
section. 

Table 1 presents the data upon which Figure 5 is based. The first column con- 
tains the genetic composition of the male parents carrying one or two 3,4 trans- 
locations, with the translocation broken to the left appearing first. The 3rd chro- 
mosome marker mutants displayed by the aneuploids can be determined by 
examining the disposition of ru and ca in the male parent. For example, the 40 
hyperdiploid male progeny of y' ; ru ca triploids and ca,\?>lru,\2 males appeared 
r%- ru ca; 55 hyperdiploid females for this region were ru ca; and 208 hyper- 
diploid females were y^; ru ca. A phenotypic appearance of the X chromosome 
marker mutant f is indicated within Table 1 ; a phenotypic appearance of the 
3rd chromosome marker mutants ru and ca must be deduced from the genetic 



Td 
^ fl 

« ^ 

i :§ 

E^ 1 



O O I— I o o 



CSI Tf< CO C— CO 



CNl TJH Tj< 

CO ^ CO 



CM CM vo 

CM r-H 



o o 

f— I o 

r— I O 

LO O 

o o 

o o 

<-^ o 

o o 

ir- o 

o 



P<^0 in 



CO 

o o 

o o 

O '^ 

o r- 

o r- 



o o o 

O CO o 

O CM O 

o ^ o 

CM LO O 

o o o 

O CO o 

o c^ o 

r-H 00 O 

Tf^ LO O 



O r-H 

CO CO 
CO t— 



o o 

o o 

o o 

o o 

o o 



CMOOCOOCM'O'-HOO 



O C^J LO O O C^l o 



r-l CM 

^ On 



O O 

O O 

O -— I 

o o 

O r-l 



CO CO QN CN| 

r— I CO CnJ CO 



r- UO rH CO 

CO t^ CO 'O 



r^ CM LO O 
CO CM 'vO CO 



O CM O O 
'o ^ t— r- < ,— I 



1 CM CM ^ CO 
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78 The University of Texas Publication 

composition of the male parents. In the second column of Table 5 appears the 
region tested and the temperature (22° C or 26° C) at which the experiments 
were conducted. The salivary map limits of each region may be ascertained from 
Figure 5. One group of control intersexes (2X3 A) is included in the last column 
of Table 1. The control intersexes chosen to appear in Table 1 appeared wild- 
type in the case of experiments designed to study end regions of chromosome 
3;c«, in the case of nearly all of the experiments designed to study interior regions 
of chromosome 3. The y^ and y^; ca control intersexes in these two respective 
types of experimental crosses do not occur with as great a frequency as the wild- 
type or ca control intersexes owing to the greater frequency of 1X2A eggs over 
2X2A eggs from the triploid mother. Table 1 is not intended to give information 
for quantitative comparisons of sex types of aneuploid intersexes vs. the control 
intersexes, but to show the total number of the various kinds of aneuploid indi- 
viduals observed. 

The evidence for survival of hypodiploid males for the region 12-H3 rests on 
12 y^; ru ca males listed in Table 1. Only one such /%• ru ca "hypoploid" male 
for the similar region 12-Hl was found and none for the region 12-e. H3 is known 
to be triplo-4, and the presence of an extra chromosome 4 may have favored 
survival of 12-H3 hypodiploid males. On the other hand, these y; ru ca males 
listed as hypodiploids for region 12-H3 might have been derived from non-dis- 
junction of all parts of both 3,4 translocations in the male parent, resulting in 
a sperm with no part of chromosome 3. Upon union of such a sperm carrying 
a Y chromosome, with an ^^^g containing one y X chromosome and two ru ca 
3rd chromosomes, viable y'; ru ca males could result. Such non-disjunctional 
forms have occurred rarely in the progeny of y'; ru ca triploids and males hetero- 
zygous for two different 3,4 translocations with the breakage points rather far 
apart, to be discussed in a later paper. The 12 /,• ru ca males listed in Table 1 
as hypodiploid males for region 12-H3 did not show extreme signs of aneuploidy 
although some of the corresponding y^; ru ca hypodiploid female sibs had reduced 
eyes, facets fused, outstretched wings, certain bristles missing and vaginal plates 
protruding. On the other hand, one /,• ru ca male listed as hypoploid for 12-H3 
showed a slight reduction in number of sex comb prongs on one leg. The author 
is therefore uncertain whether these y'; ru ca males and their y; ru ca and ru ca 
female sibs are hypoploids as Figure 5 indicates, or whether they represent forms 
owing to non-disjunction of both 3,4 translocated chromosomes in the male 
parent. If future work shows that these flies listed as hypodiploids are due to 
non-disjunction, some doubt is cast on the 4 wild-type and 1 y flies listed in 
Table 1 as hyperintersexes for region 12-H3. 

Both control and aneuploid intersexes listed in Table 1 were classified accord- 
ing to five sex types originally described by Dobzhansky (1930) and since used 
in several investigations of triploid intersexes (Dobzhansky and Schultz, 1934; 
Pipkin, 1940, 1947). In the present material only 5 groups were found, ranging 
from extreme male type I to female type V possessing normal vaginal plates, 
female type anal plates, female abdominal coloration, and male sex combs. The 
extreme female type VI, lacking sex combs was absent. Examination of the 
control intersexes in Table 1 shows that the largest female group was type IV, 
composed of intersexes with external genitalia and anal plates of the female type 



Pipkin: Sex Balance in Drosophila 



79 






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80 The University of Texas Publication 

but often protruding, misplaced, and sometimes imperfect, male or intermediate 
abdominal coloration, and male sex combs. Arbitrarily included in this type 
IV class were occasional individuals which showed both vaginal plates and a 
fragment of male external genital apparatus. For example, among the 61 type 
IV wild- type control intersexes occurring in the cross H5/ ru ca males with 
f; ru ca triploid females, 6 displayed mixtures of both male and female gen- 
italia. 

All of the aneuploid intersexes in Table 1 appeared typically intersexual. Since 
there were no obvious shifts in the male or female direction among the hyper- 
and hypointersexes, chi square homogeneity tests (using the method of Brandt 
and Snedecor, described by Fisher, 1932) were made to see if the intersex sex 
types were distributed with the same frequency in aneuploid intersexes as in 
one group of their respective control (2X3A) intersexes. To be sure that any 
shift in sex type found in aneuploid intersexes as compared with their 2X3A 
sibling control intersexes is owing to a dosage change of the short 3rd chromo- 
some section involved in the aneuploidy rather than to genetic modifiers present 
in the triploid or translocation stocks which varied from bottle to bottle of the 
same experimental cross, the following precautions were taken: Chi square 
homogeneity tests were made to determine if the sex type frequency of one 
group of control intersexes varied from bottle to bottle of the same experimental 
cross. Counts from bottles yielding fewer than five intersexes of the chosen con- 
trol group were discarded. In most cases the sex type frequencies of control inter- 
sexes coming from different bottles of the same experimental cross were homo- 
geneous. Where sex type frequencies of control intersexes coming from different 
bottles of the same experimental cross were homogeneous, these were pooled. 
Counts from the occasional bottles giving sex types of the chosen control intersex 
group heterogeneous with the frequencies of sex types of control intersexes of 
other bottles of the same experimental cross were discarded from the calculations. 
The number of such discarded intersexes may be deduced by comparing frequen- 
cies appearing in Table 1 with those for the same experimental cross in Tables 
2 and 4. In the case of wild-type control intersexes occurring in the progeny of 
H5/ ru ca males with /%• ru ca triploid females, counts from a sizable number 
of bottles had to be discarded owing to heterogeneity (presumably due to genetic 
modifiers) but this was an unusual situation. 

Finally, sex type frequencies of hyperintersexes (or hypointersexes) were 
compared with the sex type frequencies of their respective sibling control inter- 
sexes. Pooled counts of both aneuploid and control intersexes were used only 
from bottles in which the control intersex sex type frequencies had proven homo- 
geneous. Table 2 gives chi square homogeneity tests comparing pooled sex type 
frequencies of one group of control intersexes and those of hyperintersexes de- 
rived from the same experimental cross for each of 12 short regions in cases 
where hyperintersexes were numerous enough for statistical studies. P indicates 
the probability that the two sets of sex type frequencies differ owing to chance 
alone; P<0.05 being taken as indicating heterogeneity. The number of degrees 
of freedom, i.e., one less than the number of sex type groups is represented by 
"f" in Table 2. 

According to Table 2, sex type frequencies of hyperintersexes from the left 



Pipkin: Sex Balance in Drosophila 81 

hand end region H7L, the interior region C-89E, and three right hand end re- 
gions; 2 R, 30 R (at 22° C), and H5 R; proved heterogeneous with the sex type 
frequencies of their respective control intersexes. In each case, a sHght shift 
toward more male-hke sex types occurred among the hyperintersexes as com- 
pared with their respective control intersexes. A very slight shift in the female 
direction occurred among the hyperintersexes for the interior region 89E-28 as 
compared with the control intersexes. Homogeneity existed between sex type 
frequencies of hyperintersexes and control intersexes for one right end region, 
H6R, and five interior regions; 13-12, H1-85C; 85C-c; 2-28, and 28-30 (22° C) . 
The latter region, 28-30, forms a part of 2 R since Ta(3,4) 2 and Ta(3,4) 28 are 
broken very close to one another at 94A and 94D respectively. The heterogeneity 
between sex type frequencies of hyperintersexes for region 28-30 and those of 
their control intersexes at 26° C is due to a change in the distribution of the sex 
types of the control intersexes in the female direction in the 26° C series compared 
with the 22° C series. The distribution of sex types of hyperintersexes for region 
28-30 in the two temperature series is seen to be closely similar. 

To indicate the magnitude of sex type shift, means of sex types of those hyper- 
intersexes and their respective control intersexes for regions in which the two 
sets of frequencies were heterogeneous are compared in Table 3. According to the 
last column, the standard deviation of the difference between means of control 
and corresponding hyperintersexes is significant except for the last region, 
89E-28. 

For comparison with the small sex type shifts in the male direction found in 
the 3rd chromosome hyperintersexes, it was of interest to determine if a shift in 
the female direction would be produced in hyperintersexes for a short X frag- 
ment against the particular genetic background of the y%- ru ca triploid stock of 
present experiment. Males hyperploid for a minute X chromosome fragment in- 
cluding the normal alleles of yellow and achaete plus a tiny right hand portion 
including the normal allele of bobbed were crossed to y^; ru ca triploids. (The 
males were of the genetic constitution Y^.X In EN y.Y^ sc«y+.) The gray intersex 
progeny carried 2 complete X chromosomes plus the very short X fragment, a Y 
chromosome, and 3A. The yellow^ intersexes possessed only 2X3A. From this 
cross the gray hyperintersexes were found with the following sex type fre- 
quencies: 52 I, 47 II, 43 III, 50 IV, and 1 V. Yellow control intersexes occurred 
as follows: 119 I, 41 II, 11 III, and 22 IV. The two sets of sex type frequencies 
were heterogeneous with a chi square of 56.681. The gray hyperintersexes had a 

Table 3 
Comparison of mean sex types of hyperintersexes and control intersexes 





Mean, Control 


Mean, 


Difference 


S.D. of 


Region 


Intersexes 


Hyperintersexes 


Between Means 


Difference 


H7 L (22°C) 


1.750 


1.253 


0.487 


0.127 


2R(22°C) 


1.993 


1.245 


0.748 


0.091 


30 R (22°C) 


2.273 


1.424 


0.849 


0.093 


H5 R (22°C) 


1.953 


1.191 


0.762 


0.123 


C-89E (22°C) 


1.699 


1.341 


0.358 


0.094 


89F-28 (22°C) 


2.468 


2.167 


0.301 


0.160 



82 The University of Texas Publication 

mean sex type of 2.484 compared with a mean of 1.672 for yellow^ control inter- 
sexes. The difference between the means was 0.812 ± 0.122, representing a sig- 
nificant shift in the female direction in the hyperintersexes carrying the minute 
X chromosome fragment in excess of 2X3A. These results were to be expected in 
view of the work of Dobzhansky and Schultz (1934) with a minute X fragment 
covering a similar X chromosome region. 

Horaogeneity tests of the sex type frequencies of control intersexes and hypo- 
intersexes for seven short regions of chromosome 3 are presented in Table 4. Four 
of these regions, 2-30, 28-30, 28-H5, and 2-H6, cover much of the same salivary 
map region. Homogeneity between sex type frequencies of hypointersexes and 
their control intersexes exists in all of the cases except region 2-H6 and region 
28-30 (at 26° C). The mean sex type of region 2-H6 hypointersexes was 1.225; 
that of the control intersexes, 1.673; the difference between the means of 0.488 
it 0.137 being barely significant. Table 4 shows that the distribution of sex type 
frequencies of region 28-30 hypointersexes of the 22° C series does not differ 
significantly from that of the control intersexes, but at 26° C hypointersexes for 
this region have a sex type mean of 1.877 compared with 2.267 for control inter- 
sexes. The difference between the latter means is 0.390 ± 0.141, barely signifi- 
cant. In the cases of region 2-H6 and 28-30 (at 26° C), the slight sex type shift 
of hypointersexes as compared with control intersexes is in the male direction. 

Since aneuploid intersexes were characterized by late hatching, an investiga- 
tion was made of the hatching time of 2X3A intersexes of the y^; ru ca stock clas- 
sified according to their sex type, to determine if there is a difference in time of 
hatching (or in survival) of male-like intersexes vs. female-like intersexes. The 
experiment was patterned after that of Dobzhansky (1930), but intersexes were 
classified according to sex types. All eggs were deposited during a two hour 
interval. Development took place at 22° C ± 0.5° C. The sex type frequencies of 
early hatching intersexes on the 12th day after egg laying was 59 I, 33 II, 15 III, 
16 IV, and 1 V. Intersexes hatching late on the 16, 17, 18, 19, and 20th days in- 
clude the following: 17 1, 10 II, 5 III, and 4 IV. The sex type frequencies of early 
hatching intersexes were homogeneous with those of late hatching intersexes 
with a chi square of 0.145 and 0.8>P>0.7. 

None of the hypertriploid females carrying in excess of 3X3A short sections of 
chromosome 3 showed any characteristics of intersexuality. No obvious pheno- 
typic changes were found in hypertriploids for the following sections: H7 L, Hl- 
85C, 85C-C, 89E-28, 89-E, 28-30, 2-28, e-Hl, e-12, or H1-H3. (H1-H3 hyper- 
and hypotriploids could not be distinguished as such since both breaks are in the 
chromocenter, but both groups of aneuploids looked hke normal 3X3 A triploids.) 
Body size was slightly reduced in hypertriploids for 12-13, 12-Hl, and 2-30. 
Wings were held slightly apart and up in hypertriploids for all of the right hand 
end regions 2 R, 30 R, H5 R, and 6 R. In H7-13 and also in C-89E hypertriploids 
the body size was smaller than that of a diploid female; eyes were reduced; 
bristles, thin; legs misshapen; and wings were held apart. Hypertriploids carry- 
ing 13 L in excess of 3X3 A had very large, round, flat eyes with facets pro- 
foundly disturbed, and vaginal hairs were often missing. Hypertriploids for the 
following regions were found to be weakly fertile: 13-12; C-89E; 2 R; 30 R; H5 
R; H1-85C; e-Hl. 



Pipkin: Sex Balance in Drosophila 



83 



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Phenotypic changes are in general more pronounced in hypotriploid females 
lacking a short section of 3 from the 3X3 A complement than in hypertriploid 
females. Nevertheless, hypotriploid females for short regions H7-13; 13-12, 
C-89E, 28-30, 28-H5; e-Hl appeared like normal triploids. Eyes were reduced in 
size; ocelh sometimes missing; wings outstretched; bristles thin; legs misshapen 
in hypotriploids for regions H7 L, 13 L, 89E-28, and Hl-12. Hypotriploids for 
right hand end regions 2 R, 30 R, H5 R, and 28 R had wings sometimes clipped 
medially and eye facets and wing texture intermediate between diploid and 
triploid size. Wing texture was also intermediate between diploid and triploid 
texture in hypotriploids for 6 R. A similar syndrome of phenotypic change was 
sometimes seen in various diploid and triploid aneuploids for the same section, as, 
for example, holding the wings slightly apart in hypertriploid females, hyper- 
diploid females, and hyperintersexes for right hand end regions or the very 
large round eye and dark trident pattern in hypertriploid females, hyperdiploid 
females, hyperdiploid males, and hyperintersexes for H7 L. 

A usually small counterclockwise rotation of male genitalia was found oc- 
casionally in both hyperdiploid and hypodiploid males aneuploid for short sec- 
tions of chromosome 3. Among hyperdiploid males the numbers with rotated and 
normal genitaha were as follows: HZ L, 5 rotated, 76 normal; 13 L, 6 rotated, 35 
normal; 13-12, 1 rotated, 39 normal; 85C-c, 1 rotated, 86 normal; 89E-28, 1 
rotated, 95 normal; 30 R (22° C) 2 rotated, 101 normal; 6 R (22° C), 1 rotated, 
72 normal. Among hypodiploid males the numbers, with rotated and normal 
genitaha were as follows: 85C-C, 2 rotated, 7 normal; H7-13, 8 rotated, 46 
normal. Genitaha were occasionally protruding in H7-13 hypodiploid males. 
The region H7-13 in Figure 5 is marked by "Patterson" to indicate that Patter- 
son, Brown, and Stone (1940) found hypodiploid males with occasional rotated 
and sometimes imperfect genitaha in hypodiploid males for almost this same 
section. Rotation of genitalia was also found in one out of 6 super males that were 
hypoploid for region 28-30. One male hypoploid for region 2-30 and one male 
hyperploid for 6 R showed lack of anal plates and genitaha. 

DISCUSSION 

Although Bridges (1939) found that individuals with 3X4A as well as 2X3 A 
were intersexual, the present work on chromosome 3 and that of Pipkin (1947) 
on chromosome 2 show that no single short autosomal region is responsible for 
the shift toward intersexuality in triploid intersexes. No hypertriploid (3X3 A 
plus a short section) for any region of chromosome 3 (present work) or chromo- 
some 2 (Pipkin, 1947) showed any signs of intersexuality. The normal allele of 
the Hr and tra mutants (Fung and Gowen, 1956, 1957; Sturtevant 1945) is 
located either in the region 12-e or e-Hl, as Figure 5 indicates. There is there- 
fore no evidence from triploid aneuploid experiments that this normal allele 
functions as a male determiner. It may be, of course, a female determiner as 
Stone (1942) suggested possible. 

Small shifts in a male direction were found in the present experiment in hyper- 
intersexes for several short regions of chromosome 3 but for none of chromosome 
2 (Pipkin, 1947). Three slightly different right hand end regions of chromosome 



Pipkin: Sex Balance in Drosophila 85 

3 produced the largest shifts in a male direction in hyperintersexes. These shifts 
are comparable in size to those produced in a female direction by the addition of a 
very short section of the X chromosome to the 2X3A intersex complement. On 
the other hand, none of 7 different hypointersexes lacking a short section of 3 
from the 2X3 A complement showed a shift in the female direction. The sex types 
of one region, 2-H6 even showed a barely significant shift in the male direction 
when compared with the sex types of 2X3A sibling control intersexes. This last 
region forms a part of the right hand end region 2 R which in hyperintersexes 
causes a definite shift in the male direction. Hypointersexes for two short sections 
of the X chromosome, in contrast, were shown by Pipkin (1940) to be strongly 
shifted in the male direction. 

The small shifts in a male direction in hyperintersexes for certain sections of 
the 3rd chromosome may then be interpreted in one of three ways: (1) as sex 
balance shifts owing to dosage changes of the particular aneuploid region in- 
volved; (2) as owing to retardation of development of these late hatching hyper- 
intersexes, assuming that triploid intersexes begin development as males, 
followed by a turning point to female development (Dobzhansky, 1930); (3) 
as owing to possible differential survival on the part of male-like hyperintersexes 
as compared with female-like hyperintersexes for the particular regions involved. 
In connection with this last possibility the experiment of time of development of 
the members of different intersex sex type groups carried out in the present 
investigation shows that among ordinary 2X3A stock intersexes the sex distribu- 
tion of early hatching 2X3A intersexes differs by chance alone from the sex type 
distribution of late hatching intersexes of the same age but subject to the un- 
favorable environment of an older culture. 

Hypodiploid males for several regions showed occasional rotation of genitalia, 
an observation also made by Patterson, Brown, and Stone (1940). This rotation 
is not ,contrary to Kelstein (1938, 1939), taken as a sign of intersexuality because 
occasional rotation also was found in hyperdiploid males for several regions. The 
rotation is thought to be the consequence of genie imbalance and does not repre- 
sent a sex shift. Patterson, Brown, and Stone (1940) report no intersexuality in 
hypodiploid males for a longer region which includes the region XV studied by 
Kelstein ( see Figure 5 ) . 

Third chromosome aneuploid studies of the present investigation as well as 
those of the 2nd chromosome (Patterson, Brown, and Stone, 1940; Pipkin, 1947) 
and X chromosome (Dobzhansky and Schultz, 1934; Pipkin, 1940) support the 
conclusion that dosage changes of portions of the X chromosome are far more 
effective than dosage changes of portions of either of the large autosomes in 
affecting sex balance. 

ACKNOWLEDGMENTS 

This research was supported by Grant C-3453 awarded by The National 
Cancer Institute, U. S. Public Health Service, Bethesda 14, Maryland. Thanks 
are due Dr. David T. Ray of Howard University for assistance in X-raying to 
produce 3,4 translocations; to Dr. Clay G. Huff, of the Naval Medical Research 
Institute for the use of his calculator; to Dr. E. B. Lewis, of the California Insti- 
tute of Technology for certain 3,4 translocations and information concerning 



86 The University of Texas Publication 

their breakage points; to Mr. David W. Ray, Mr. Presley Autry, and Mr. Hollis 
Seunarine of Howard University for invaluable technical assistance; and to Dr. 
Harold E. Finley of Howard University for the many courtesies enjoyed while 
carrying on this work in his department. 

SUMMARY 

A study of triploid and diploid aneuploids occurring in the progeny of males 
heterozygous for one or two different 3,4 translocations and triploids failed to 
reveal any marked shifts in sexual differentiation owing to dosage change of the 
portion of chromosome 3 involved in the aneuploidy. Hypertriploid females with 
no signs of intersexuality survived for each of 14 short regions which covered 
the entire chromosome 3. The mean sex type of hyperintersexes for 5 short 
regions, 3 of which include the right hand end of chromosome 3, were signifi- 
cantly more male-like than the mean sex type of corresponding sibling control 
intersexes with 2X3A only. No sex type shift in the female direction was found 
among the hypointersexes for 7 different short regions. Further there was no 
qualitative shift in sex differentiation among hyper- and hypointersexes occur- 
ring too seldom for statistical analysis of their sex types. An occasional rotation 
of genitalia was found in both hyperdiploid and hypodiploid males for several 
regions. It was not taken as an indication of intersexuality. 

REFERENCES 

Bridges, C. B. 1921. Triploid intersexes in Drosophila melanogaster. Science 54:252-254. 

. 1922. The origin of variations in sexual and sex-limited characters. Amer. Nat. 56: 

51-63. 

. 1939. Cytological and genetic basis of sex. Sex and Internal Secretion, ed. by Allen, 

Danforth, Doisy, pp. 15-63. Williams and Wilkins, Baltimore. 

Burdette, Walter J. 1940. The effect of artificially produced tetraploid regions of the chromosomes 
of Drosophila melanogaster. U.T.P. 4032: 157-163. 

Crow, James F. 1946. Absence of a primary sex factor on the X chromosome of Drosophila. Amer. 
Nat. 80:663-665. 

Dobzhansky, Th. 1929. Genetical and cytological proof of translocations involving the third and 
fourth chromosomes of Drosophila melanogaster. Biol. Zbl. 49:408-419. 

. 1930. Studies on the intersexes and supersexes in D. melanogaster. Izv. Bur. Genet. 

(Leningr.) No. 8:91-158. 

and C. B. Bridges. 1928. The reproductive system of triploid intersexes in Drosophila 



gaster. Biol. Bull. 59:128-133. 

Dobzhansky, Th. and C. B. Bridges. 1928. The reproductive system of triploid intersexes in Dro- 
melanogaster . Amer. Nat. 62:425-434. 

and Jack Schultz. 1934. The distribution of sex factors in the X chromosome of Dro- 
sophila melanogaster. Journal of Genetics 28:349-386. 

and B. Spassky. 1941. Intersexes in Drosophila pseudoobscura. Proc. Nat. Acad. Sci. 

(Wash.) 27:556-562. 

Dubovsky, N. V. 1939. The effect of a gradual increase of aneuploidy on the viability and death 
stage of Drosophila melanogaster. Bull. Biol. Med. Exp. 7(1) :31-33. 

Fisher, R. A. 1932. Statistical Methods for Research Workers. Oliver and Boyd, Edinburgh and 
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Pipkin: Sex Balance in Drosophila 87 

Fung, Sui-Tong Chan and John W. Gowen. 1956. A major sex locus in Drosophila melanogaster. 

Rec. Gen. Soc. of America 25:644. 
. 1957. The developmental effect of a sex-limited gene in Drosophila melanogaster. 

Jour, of Exp. Zool. 134:515-532. 
Goldschmidt, Richard B. 1948. New facts on sex determination in Drosophila melanogaster. Proc. 

Nat. Acad. Sci. (Wash.) 34:245-252. 
. 1949a. The intersexual male of the Beaded-Minute combination in D. melanogaster. 

Proc. Nat. Acad. Sci. (Wash.) 35:314-316. 
. 1949b. Neue Tatsachen zur Analyse der Geschlechtsbestimmung bei Drosophila 



melanogaster. Arch. d. Julius Klaus Sliftg. Ziirich 23:539-549. 
. 1949c. The Beaded-Minute intersexes in Drosophila melanogaster. Jour. Exp. Zool. 



112:233-302. 

. 1955. Theoretical Genetics, p. 93. University of California Press. Berkeley and Los 



Angeles. 
Hollingsworth, M. J. 1953. Intersexes in D. subobscura. D.I.S. 27:94. 
Kelstein, L. V. 1938. The influence of minute deficiencies and duplications on the individual 

development of Drosophila melanogaster. Bio. Zhur. 7: 1145-1169. 
. 1939. Effect of minute deficiencies and duplications of the right arm of Ill-chromo- 

some upon the individual development of Drosophila melanogaster. Bull. Biol. Med. Exp. 

7(l):28-30. 
Lebedeff, G. A. 1939. A study of intersexuality in Drosophila virilis. Genetics 24:553-586. 
Lewis, E. B. 1951. Additions and corrections to the cytology of rearrangements. D.LS. 25: 108-109. 

. 1956. Additions and corrections to the cytology of rearrangements. D.LS. 30:130. 

Meyer, Helen. 1959. Report of new mutants. D.LS. 32:83. 

Mickey, George H. 1950. Studies on intersexuality in Drosophila. Records of the Genetics Society 

of America 19:115. 
Morgan, T. H., H. Redfield, and L. V. Morgan. 1943. The constitution of the germinal material 

in relation to heredity. Yearbook Carnegie Institution 42: 171-174. 
Muller, H. J. and W. S. Stone. 1930. Analysis of several induced gene-rearrangements involving 

the X-chromosome of Drosophila. The Anatomical Record 47: 393-394. 
Neuhaus, M. E. 1939. Triploid stock in D. simulans. D.I.S. 11:47. 
Newby, W. W. 1942. A study of intersexes produced by a dominant mutation in Drosophila 

virilis, Blanco stock. U.T.P. 4228:113-145. 
Patterson, J. T. 1931. The production of gynandromorphs in Drosophila melanogaster by X-rays. 

Jour. Exp. Zool. 60: 173-211. 

. 1938. Sex differentiation. Amer. Nat. 72: 193-206. 

and Wilson Stone. 1938. Gynandromorphs in Drosophila melanogaster. U.T.P. 

3825:1-67. 

. 1952. Evolution in the Genus Drosophila. The Macmillan Company, New York. 

Meta Suche Brown, and W. S. Stone. 1940. Experimentally produced aneuploidy 



involving the autosomes of Drosophila melanogaster. U.T.P. 4032: 167-189. 

, Wilson Stone, and Sarah Bedichek. 1937. Further studies on X-chromosome balance 



in Drosophila. Genetics 22:407-426. 
, Wilson Stone, Sarah Bedichek, and Meta Suche. 1934. The production of trans- 



locations in Drosophila. Amer. Nat. 68:359-369. 
Pipkin, Sarah Bedichek. 1940. Multiple sex genes in the X-chromosome of Drosophila melano- 
gaster. U.T.P. 4032: 126-156. 



88 The University of Texas Publication 

• 1947. A search for sex genes in the second chromosome of Drosophila melanogaster 

using the triploid method. Genetics 32:592-607. 

Schuhz, Jack and Th. Dobzhansky. 1933. Triploid hybrids between Drosophila melanogaster and 
Drosophila simulans. Jour. Exp. Zool. 65:73-82. 

Spurway, H. and J. B. S. Haldane. 1954. Genetics and cytology of Drosophila subobscura IX. An 
autosomal recessive mutant transforming homogametic zygotes into intersexes. Jour, of 
Genetics 52:208-225. 

Stalker, Harrison D. 1942. Triploid intersexuality in D. america Spencer. Genetics 27:504^518. 

Stone, W. S. 1942. The I^^ factor and sex determination. U.T.P. 4228: 146-152. 

Sturtevant, A. H. 1920. Intersexes in D. simulans. Science 51:325-327. 

. 1945. A gene in D. melanogaster that transforms females into males. Genetics 30: 

297-299. 



. 1946. Intersexes dependent on a maternal effect in hybrids between Drosophila 

repleta and Drosophila neorepleta. Proc. Nat. Acad. Sci. (Wash.) 32:84-87. 

1949. The Beaded- Minute combination and sex determination in Drosophila. Proc. 



Nat. Acad. Sci. (Wash.) 35:311-313. 

Ward, C. L. and W. S. Stone. 1952. Studies in the repleta group: the melanopalpa subgroup. 
U.T.P. 5204: 119-128. 

Wharton, Linda T. 1942. Analysis of the repleta group of Drosophila. U.T.P. 4228:23-52. 



Gene-Environment Interaction in Relation to the Nutrition 
and Growth of Drosophila 

FORBES W. ROBERTSON 
A.R.C. Unit of Animal Genetics, Edinburgh 



INTRODUCTION 

In former studies of the genetic variation of body size in Drosophila melano- 
gaster, it has been the practice to supply cultures with excess food since this 
minimises environmental variance when the ordinary live yeast cultures are 
used. However, under natural conditions, or the usual competitive regime in lab- 
oratory bottle or population cage, such conditions would be rather exceptional. 
Indeed there is little doubt that variation in the quantity and quality of the food 
supply comprises the major part of the environmental variation encountered by 
growing larvae. The characteristic genetic attributes of populations will be estab- 
hshed under sub-optimal conditions and hence analyses which are confined to 
the most favourable environment cannot detect gene-environment interactions 
which may be highly relevant to the properties of intrapopulation variance. 
Therefore, gene-environment interactions which are associated with different 
kinds of sub-optimal diet are being studied in some detail and I shall describe 
briefly some of the first results of this work. 

The approach depends on the use of chemically defined, aseptic media upon 
which the larvae are grown. Various workers have contributed to the develop- 
ment of such media including Tatum (1939), Schuhz, St. Lawrence and New- 
meyer (1946), Begg and Robertson (1950), but the later refinements are due to 
Sang (1956) who has determined the quantitative requirements which permit 
growth to the normal size and at the same rate as on the usual live yeast medium. 
If the concentration of one or more essential nutrient is inadequate the length of 
the larval period is increased and body size may be reduced as well, according to 
the nature and severity of the deficiency. The agar gel medium includes 7 B 
vitamins, together with casein as amino-acid source, ribonucleic acid, cholesterol, 
lecithin, fructose and salts, so there are many ways in which a diet can be made 
inadequate. Most relevant, however, will be the suboptimal diets which repro- 
duce the sort of nutritional conditions commonly encountered under competitive 
conditions. Drosophila lives on yeast and, from the published analyses of yeast 
it has been inferred (Sang, in press), that the various B vitamins, with the pos- 
sible exception of folic acid, are unhkely to be limiting factors since they are 
abundant in relation to minimal requirements. Hence variation in the avail- 
ability of other essential constituents is probably more important and, m particu- 
lar, variation in the protein level suggests an obvious starting point. The data 
described here consist of comparisons of the performance of different strains 
when grown on the usual hve yeast medium, on media deficient in casein but 
otherwise supphed with excess of other constituents and also on a medium in 
which the optimum ratio of constituents is maintained but the concentration 



90 The University of Texas Publication 

in the agar gel is reduced. The adverse effects of the latter medium are probably 
chiefly due to protein deficiency. The synthetic medium which has been modified 
in this way is that referred to as Medium C (Sang 1956) . 

GENERAL OBSERVATIONS 

Before giving the results a few general observations are necessary. Body size 
is expressed as three times the natural logarithm of thorax length, measured in 
1 /100mm., since this provides an adequate index of weight. Rough estimates of 
percentage differences in weight can be made by multiplying observed differ- 
ences between means by 100; this enables the relative magnitude of effects to be 
kept in perspective. 

The length of the larval period, where recorded, is measured in days con- 
verted to natural logs, and is based on the egg to adult period minus the duration 
of the pupal period which is comparatively constant and remarkably independent 
of both the duration of the larval period and differences in adult body size due to 
selection. 

A lengthening of the larval period constitutes the first sign that a larval diet 
is inadequate. Especially with protein deficiency there may be a considerable 
lengthening of larval hfe without reduction of final size, but of course, with more 
drastic shortage, body size is reduced as well. There is, therefore, a definite ability 
to regulate body size when protein is a limiting factor and this can be regarded as 
a form of homeostasis. Figure 1 shows the sort of relation which exists between 
body size and length of the larval period with variable protein levels. The two 
wild populations compared evidently differ in their ability to regulate in this 
way. If the protein level falls too low, body size dechnes and progressive reduc- 
tion of body size is associated with a corresponding proportional increase in de- 
velopment time. For a precise comparison of strains with respect to the protein 
concentration at which the regulation breaks down, a range of comparisons is 
needed. But for a quick survey it is sufficient to provide a diet which reduces body 
size appreciably and then strains can be compared in terms of how far body size 
is reduced below the level attained under favourable conditions. 

Genetic differences in this homeostatic regulation of body size are relevant to 
fitness generally. With respect to the general competitive advantage of quick 
versus slow development time there is Httle doubt. With regard to adult body 
size it is known that a sparse larval diet which reduces body size also causes a 
lower rate of egg production. In particular, it has been shown (Robertson 1957) 
that a given reduction in body size is associated with a proportional reduction in 
the number of ovarioles, without affecting their individual rate of production, 
and hence a proportional decline in egg production. Therefore differences in 
percentage reduction of body size below the maximum probably account for the 
major share of what is commonly called fitness with respect to the specific un- 
favourable conditions. 

Finally, in these tests which measure the effects of sub-optimal diets, the 
nutrient gel is homogeneous and always in excess and so the results are due to 
differences in the chemical composition of the diet and competition between 
larvae may be disregarded. 



Robertson: Gene-Environment Interaction 



91 




1-8 



2 3 



24 



19 20 21 2-2 

LOG. DAYS LARVAL LIFE 

Fig. 1. Body size and length of larval period on synthetic media which differ only in casein 
concentration. 

EXPERIMENTAL RESULTS 

These comprise the following: In two different wild populations of Drosophila 
melanogaster, Kaduna and Pacific, mass selection for large and small size has 
been carried out for 5 generations, after which the performance of the selected 
and unselected strains were compared on different media. The Kaduna popula- 
tion was selected for wing length, which is highly correlated with body size, by 
Mr. Barry Latter for other purposes and I have to thank him for kindly providing 
me with subcultures for testing. In the Pacific population selection was for 
thorax length; two large and two small strains were selected simultaneously and 
the comparisons included crosses between large and small strains as well. In ad- 
dition the performance of a number of inbred lines has been compared with 
the population from which they were derived. 

(a) The Kaduna population. To provide a suitable index of deviation of the 
selected strains from the mean size of the unselected population, the differences 
between means of flies reared on the usual live yeast medium is expressed in 
terms of the within-culture standard deviation of the unselected flies. Thus the 
selected strains deviate some 1.5-2a— equivalent to 8-10% difference in body 
weight. The relation between deviation from mean and response to various sub- 
optimal diets is shown in Fig. 2, in which the ordinate measures the decline 
in body size below maximum size. Four treatments were used, two levels of 
protein shortage and two levels of nutrient concentration. The points represent 
the average for 20 — 30 individuals drawn from several rephcate cultures. 

The results are very striking since individuals from both the selected strains 



92 



The University of Texas Publication 



-o I 



z 

=> -0-2 



■0-3- 



KADUNA POPULATION 

VARIABLE NUTRIENT CO NCFMTPATIOM 



-2(r 



-ir 



+ 10- 



< 
> 

uj -O- 



-0-2 



VARIABLE PROTEIN 



•2(r 



+20- 




• -SOX CONC 
30X CONC 




-2/^ CASEIN 
-15% CASEIN 



-03 



Fig. 2. The relation between deviation from the mean body size of the unselected population 
under favourable conditions and decline in body size on media deficient in protein or with reduced 
concentration of nutrients. The deviation is measured in terms of the within-culture standard 
deviation of the unselected flies reared under the favourable conditions of the usual live yeast 
medium. 



suffer a greater decline in body size than those from the unselected population. 
Thus a protein deficiency which results in about 5-6% decline in body size in 
the unselected populations, causes 10% and 20% decline in the small and large 
strains respectively, while a reduction of nutrient concentration which reduces 
the size of the unselected flies by about 20% results in a 30% decline in the 
selected strains. Thus deviation from the mean in either direction lowered the 
ability to regulate body size on these sub-optimal diets. 

An additional point of interest relates to the within-culture variance. It is a 
familiar experience to find that this increases when flies are grown on sub- 
optimal media. In view of the homogeneous nature of the medium and the fact 



Robertson: Gene-Environment Interaction 93 

that about 70% of the variance of body size, under optimum conditions, is 
genetic (Robertson 1957) there is Httle doubt that the greater variance repre- 
sents genetic differences in abihty to cope with the adverse environm.ent. This 
phenomenon is shown in Table 1, which deals with the pooled within-culture 

Table 1 
Within-culture variance of body size (Kaduna) 





Unselec 


ed 


Large 

0.0027 


d.f. 
28 


Small 


Treatment 


a- 

0.0035 


d.f. 

28 


a"- d.f. 


OPTIMUM 


0.0043 28 


2% Casein 
1.5% Casein 
50% Concentration 
30% Concentration 


0.0061 
0.0055 
0.0043 
0.0074 


25 
33 
24 

25 


0.0150 
0.0045 
0.0059 
0.0051 


24 
6 

22 
30 


0.0109 28 
0.0147 33 
0.0073 19 
0.0046 15 



variance of the different series and treatments. Variance on the sub-optimal 
diets is consistently higher than the corresponding variances on Hve yeast media 
and there is also some indication, especially in the small strain, that the relative 
increase in variance is greater in the selected than the unselected populations. 

(b) The Pacific population. The results of a similar comparison of the Pacific 
strains on the usual medium and on 50% nutrient concentration are shown in 
Figure 3; the four possible crosses between a large and small strain are also 
shown. Here the two small strains are rather similar in size, while the large 
strains differ, one (AL) being only about 0.5o- away from the mean. The results 
of these comparisons follow essentially the same pattern as the Kaduna com- 
parisons; the regression of decline in size on squared deviation from the mean 
of the unselected is clearly significant (b = - 0.25 ± 0.008). However, some 
additional points must be noted. Firstly, although the general trend is clear 
enough there is evidently variation with respect to the deviation from the mean 
and relative decline with this treatment. Thus the small A strain is apparently 
no worse than the unselected population in these terms and the same is true of 
one of the large strains (AL). Secondly, the crosses deviate appreciably from 
the intermediate value, as usual in such tests, and all exceed the unselected 
population in body size. Although they differ slightly in percentage decline, 
their average is about the same as that of the unselected flies. Thus, although 
the large and small parent strains may have lower ability to maintain body size 
on this sort of deficient diet, the more normal sized F^ reacquires the properties 
of the unselected population. 

Taken together then, the results of the Kaduna and Pacific tests indicate that 
selection for large and small body size under favourable conditions of excess 
food, results in genetic changes which lower the animal's ability to cope with 
sub-optimal, especially protein deficient, diets. This means that the selected 
individuals grown in such media take relatively longer to reach maturity and 
will lay fewer eggs than the unselected flies, even though their egg production 
may not differ from that of the unselected population when all are grown under 



94 



The University of Texas Publication 



> 
a 
O 

1 -O-l 
< 

2 -0-2 
O 
a. 



•0-3 - 







PACIFIC POPULATION 








50°/ NUTRIENT CONCENTRATION 




AS 


BS 


U 


AL 


BL 




+ 


i 


* 


i 


^ 




-2(r 




-ir o 




+ i(r 


+2r 


o 







G^ 


X 

X 
X 




- 





^ 


-J 


o 

















Fig. 3. The relation between deviation from the mean and response to a medium with low 
nutrient concentration; the crosses refer to the F^^ of matings between a large and a small strain. 

favourable conditions, as is usually the case when such selected strains are 
compared (Robertson 1957). Thus with selection under favourable conditions, 
the effective loss of fitness will be greatly influenced by environmental, espe- 
cially nutritional conditions. 

The characteristic relationship revealed in these tests might, a priori, be re- 
garded as peculiarly characteristic of selection for large and small size as opposed 
to selection for some other character or it might be regarded as a less specific 
by-product of a general loss of balance or co-adaptation whenever the original 
genetic equilibrium is disturbed. It might be argued, for instance, that selection 
has led to an increase in the level of inbreeding and that this accounts for the 
lower performance under adverse conditions. To check this point, a similar 
comparison has been made between the wild Pacific population and 4 highly 
inbred lines derived from this population. If non-specific effects are involved 
such lines should be subject to a greater decline than the unselected flies and 
might even be expected to be a good deal worse than the selected strains. In 
practice it turns out however that they resemble the unselected flies in response 
to diets deficient in protein or with a reduced concentration of nutrients. Table 
2 shows that with a lower concentration of nutrients the averages for the inbred 
lines and the wild stock are almost identical; for the low casein diet the results 
are more variable, and line 10 is missing due to an accident during sterilisation 
of the eggs, but the average decline is close to that of the wild flies and one line. 
No. 4, suffers less decline than the former. It will be noted that the media lead to 



Robertson: Gene-Environment Interaction 95 

Table 2 
Performance of inbred lines and the wild stock on favourable and adverse diets 









Decline 


n body 


size 




Body Size on 












Live Medium 




50% 




Protein 


Genotype 


(Log Units) 




Concentration 




Deficient 


Inbred line 3 


14.044±.008 




—0.393 




—0.300 


4 


14.053±.008 




—0.402 




—0.162 


6 


13.981 ±.008 




—0.450 




—0.271 


10 


13.963+.008 




—0.372 










Average 


—0.404 




—0.244 


Wild population 


14.113±.011 




—0.408 




—0.219 



The means are based on the measurement of about 30 flies shown equally for 3-4 replicated cultures. 

a relatively greater decline in body size than in the other tests. This may be 
partly due to differences in the live medium or the temperature between experi- 
ments, but is probably mainly due to a relatively higher temperature of auto- 
claving since certain amino-acids tend to be inactivated when autoclaved in 
the presence of sugar (Evans and Butts 1949). This circumstance, however, does 
not affect the validity of the results and it is difficult to avoid the conclusion that 
the characteristic behaviour of the selected strains is directly due to change in 
the frequency of genes which affect body size under optimal conditions. It will 
be noted that the inbred lines are smaller than flies of the wild population, some 
7 to 15% smaller, which is quite typical of the effects of inbreeding in this 
species. It has been concluded, on other grounds (Robertson and Reeve 1955), 
that such a decline due to inbreeding is qualitatively different in physiological 
origin and genetic behaviour from a comparable decline due to selection and the 
contrast in reaction to deficient diets of inbred lines and selected strains — 
deviating about equally from the base population — supports that view. 

DISCUSSION 

The leads given by these experiments are being followed up and discussion of 
their wider implications will be deferred until various current tests are com- 
pleted. However, the present data are relevant to two familiar problems which 
have been encountered in earlier studies relating to body size, namely, the 
asymmetry of selection response and the stability of average body size. With 
respect to the former, mass selection for large and small body size leads to 
different rates of response in the two directions. Selection for small body size 
generally proceeds further and faster away from the base population than 
selection for large size which eventually reaches a point at which the response 
to selection stops, although plenty of genetic variation remains and it is easy to 
select down again (Robertson and Reeve, 1952; Robertson, 1955). Now the 
relative deviation from the original population of strains selected for large and 
small size depends on the larval diet. Under favourable conditions a large and 
small strain may deviate about equally from the parent population but when 
the comparisons are made on say, a protein deficient medium, the difference 
between the large and unselected strains diminishes and may even be oblit- 



96 The University of Texas Publication 

erated, whereas the deviation of the small strain increases, so that a more or 
less symmetrical situation is converted into a highly asymmetrical one in terms 
of deviation from the unselected population. This appears very clearly in 
Table 3; the corresponding deviations have been averaged for different treat- 

Table 3 
Deviation of body size from the mean of the unselected population — log units 

Kaduna Pacific 

Treatment Large Small Large Small 

Normal 0.12 —0.09 0.09 —0.07 

Sub-optimal 0.02 —0.21 0.02 —0.09 

Based on average of values for different sub-optimal treatments (Kaduna) or strains (Pacific). 

ments (Kaduna) or for both large or small strains (Pacific). Hence it is likely 
that progressive change in the gene arrays, due to selection for large body size, 
may result in more exacting demands on the available diet, which, looked at the 
other way, becomes effectively sub-optimal and so this sort of gene-environment 
interaction could contribute to the eventual barrier to selection progress. If this 
is true, the actual level at which the barrier or plateau is established will be 
greatly influenced by the composition of the diet provided during selection. Also, 
if the comparisons between the inbred lines and the wild population are typical, 
loss of heterozygosity could be quite unimportant in the development of such a 
situation. Naturally with selection for small size, comparable changes will 
assist rather than hinder the progress of selection. 

It is well known that the average body size of individual wild populations 
maintained in the laboratory remains comparatively stable over long periods 
(Reeve 1954). In unpublished experiments it was found that when samples 
from an initially heterogeneous population were run for many generations 
under competitive conditions, the mean body size of the replicated populations 
was remarkably similar when compared under the usual favourable conditions. 
It is reasonable to assume that the stability of the mean reflects an underlying 
genetic equilibrium such that deviation either way tends to lower fitness, but the 
manner in which this might be effected has remained obscure. Under favourable 
conditions, the genetic variance of body size is largely additive in behaviour, it 
is easy to select either way and the changes produced by a few generations of 
selection do not afford much evidence of loss of fitness as judged by records of 
egg production, viability and so forth. But the present experiments reveal an 
entirely different picture when the larval diet is sufficiently inadequate to 
reduce body size below the maximum level. It is reasonable to infer that progres- 
sive deviation from the mean — measured in terms of body size under optimal 
conditions — will be correlated, on protein deficient diets, with relatively greater 
decline in body size, a longer larval period and a lower rate of egg production, 
which adds up to loss of fitness. 

Naturally it has to be shown that the sort of adverse conditions provided in 
these experiments are relevant to natural conditions and this is being checked. 



Robertson: Gene-Environment Interaction 97 

But in the light of existing information about the nutrition of Drosophila and 
the inter-relations which exist between metabolic processes, it would be rather 
surprising if they were not. 

SUMMARY 

There is a well marked capacity for regulating body size in Drosophila, to be 
regarded as a form of homeostasis. Sub-optimal diets frequently cause a lengthen- 
ing of the larval period but no decline in body size, unless the nutritional de- 
ficiency becomes too severe. Genetic differences in the ability to maintain body 
size have been demonstrated in the performance of strains, selected for a few 
generations for large and small size, with that of the unselected base population 
on chemically defined, aseptic media, deficient especially in protein. Perform- 
ance is measured in terms of the relative decline in body size on such media 
compared with the size attained under optimal conditions provided by the 
usual live yeast medium. There is good evidence that individuals of the selected 
strains are inferior to the members of the unselected population in this respect. 
Comparisons between highly inbred lines and the parent population show 
that the former are, on average, no worse than the outbred population, and 
hence the characteristic relation between deviation from the mean of the wild 
population and decline in body size, under sub-optimal conditions, is peculiarly 
characteristic of selection for large and small body size and cannot be attributed 
to less specific effects such as inbreeding. The results are discussed in relation to 
the asymmetry of the response to selection for large and small body size, the 
level at which the response to selection for large size ceases, the stability of body 
size in populations and the relevance of such gene-environment interactions to 
natural conditions. 

REFERENCES 

Begg, M. and F. W. Robertson. 1950. The nutritional requirements of Drosophila melanogaster. 

J. Exp. Biol. 26:380-387. 
Evans, R. J. and H. N. Butts. 1949. Inactivation of amino-acids by autoclaving. Science 109: 

569-571. 
Reeve, E. C. R. 1954. Natural selection for body size in Drosophila. IXth Congr. Genetics, Abstr. 

Caryologia, Vol. suppl. 1954. 
Robertson, F. W. 1955. Selection response and the properties of genetic variation. Cold Spr. Harb. 

Symp. Quant. Biol. 20:166-177. 
-. 1957a. Studies in quantitative inheritance. X. Genetic variation of ovary size in 

Drosophila. J. Genet. 55:410-427. 

-. 1957b. Studies in quantitative inheritance. XL Genetic and environmental correla- 



tion between body size and egg production in Drosophila melanogaster. J. Genet. 55:428-443. 
and E. C. R. Reeve. 1952. Studies in quantitative inheritance. I. The effects of selection 



of wing and thorax length in Drosophila melanogaster. J. Genet. 50:416-448. 

1955. Studies in quantitative inheritance. VIII. Further analysis of heterosis in 



crosses between inbred lines of Drosophila melanogaster. Z. indukt. Abstamm.-u. Vererb. 
86:439-458. 
Sang, J. H. 1956. The quantitative nutritional requirements of Drosophila melanogaster. J. Exp. 
Biol. 33:45-72. 



98 The University of Texas Publication 

. 1959. Circumstances affecting the nutritional requirements of Drosophila melano- 

gaster. Ann. N. Y. Acad. Sci., in press. 
Schultz, J., P. St. Lawrence and D. Newmeyer. 1946. A chemically defined medium for the 

growth of Drosophila melanogaster. Anat. Rec. 96:540 (Abstr.). 
Tatum, E. L. 1939. Nutritional requirements of Drosophila. Proc. Nat. Acad. Sci., Wash. 25: 

490-497. 



Observations on Variegated Position Effects in Drosophila 

melanogaster 

GEORGE H. MICKEY 

Department of Zoology, Physiology and Entomology 
Louisiana State University 



Variegated eye color traits in Drosophila have been known since the early days 
of radiation genetics (Glass, 1933 and 1934) and numerous investigators have 
demonstrated that both the brown- variegated eye colors and the white-mottled 
mutants result from chromosome breakage and rearrangement of segments such 
that euchromatin is brought into contact with heterochromatin or that hetero- 
chromatic segments are transposed or translocated to a position adjacent to 
euchromatin (Baker, 1952 and 1953; Hannah, 1951; Hinton, 1949; Lewis, 1950; 
Mickey, 1954; Schuhz, 1936; and Slatis, 1955a and 1955b). In other words, the 
irregular distribution of pigment in the eyes which resulted in the mottled or 
spotted distribution was actually a position effect. One case of particular interest 
is described in this paper and an analysis is given of its behavior under varying 
conditions. 

This chromosomal aberration, No. lOC-2, arose in a male germ cell which 
had been irradiated with gamma rays from a Cobalt-60 source.- Aduk wild type 
males of the Oregon-R strain were irradiated and only those germ cells were 
used which were spermatids or mature sperms at the time of treatment The aber- 
ration produced a dominant phenotypic variegation of eye pigment which 
varied under different genetic combinations. The mottling was more conspicuous 
in males than in females. 

Genetic tests showed that stock No. lOC-2 contained a reciprocal translocation 
between the long arm of the Y chromosome and the right arm of the second 
chromosome, the break in the latter being very close to the brown locus. A pre- 
liminary report of this translocation has been published (Mickey, 1955). A dia- 
gram of this rearrangement is shown in figure 1 and a camera lucida drawing of 
a metaphase figure from a ganglion cell appears in figure 2. Cytological analysis 
of salivary gland chromosomes reveals the break in 2R to be at 59C6 on Bridges 
map (Bridges, 1935). The error in identification is not more than one or two 
bands. Figures 3 and 4 show photomicrographs of the salivary gland chromo- 
somes in a structural heterozygote; figure 3 shows the Y.2 chromosome and 
figure 4, the end of the 2.Y chromosome. 

1 Work supported in part by a research grant from the United States Atomic Energy Com- 
mission, Contract No. AT (11-1) -89, Project No. 7, and in part by a grant from Research Funds 
of the Graduate School of Northwestern University. 

2 Grateful acknowledgment is given to Dr. Howard Vogel for aid in treating the experimental 
flies with gamma rays from a Cobalt-60 source at Argonne National Laboratory. 



100 



The University of Texas Publication 
2R 



YL 



•Y 0(S:ii£^2:Ml 



59 C 6 



YL 2R 



Y-2 



IOC-2 STOCK 

Fig. 1. Diagram showing the segmental interchange between the right arm of the second 
chromosome and the long arm of the Y chromosome in the lOC-2 stock. 




ioc-2 d* 



Fig. 2. Camera lucida drawing of a metaphase figure from a ganglion cell showing the 2.Y 
and Y.2 chromosomes. 





Figs. 3 and 4. Photomicrographs of salivary gland chromosomes showing (left) the Y.2 
chromosome protruding from the chromocenter, and (right) the 2.Y chromosome and the tip 
of the normal second chromosome bent around to make terminal adhesion with the heterochro- 
matic Y segment. 



Mickey: Variegated Position Effects 101 

VARIEGATED MALES 

Males of the lOC-2 stock have a chromosome complement of X/Y.2; 2/2. Y; 
3/3; 4/4. The eye color is distinctly variegated or mottled with brown spots on 
wild type red background or, in an individual homozygous for vermilion, brown 
flecks on a vermilion background. Such males are fertile since both parts of the 
Y chromosome are present when the individual is heterozygous for normal 2R. 
Both parts of 2R are present also, one on the Y.2 chromosome and the other on 
the normal second. Random distribution of the 2.Y and normal 2 would be ex- 
pected to produce half the males heterozygous for 2.Y and carrying the Y.2 
chromosome and the other half homozygous for the normal second chromosome 
but also carrying the Y.2, thus being deficient for part of the Y but having a 
duplication for the tip of 2R. The former is just like the lOC-2 parent male, 
whereas the latter carries a duplication of 2R and is sterile because of a deficiency 
for part of the Y chromosome. Furthermore, the latter males might be expected 
to be non-mottled if both 2.Y and Y.2 chromosomes are required for the produc- 
tion of variegation, or perhaps more likely, to be only slightly mottled because 
of the reduced amount of heterochromatin present. 

In the cross of 
lOC-2 males (X/Y.2;2/2.Y) x wild type females (X/X;2/2) 
normal segregation can be diagrammed as follows: 

= normal wild type female 

= deficiency for tip of 2R; lethal 

= wild type male; deficiency for part of Y; 

duplication for tip of 2R; sterile. 
= lOc-2 mottled male 

Thus all females derived from this cross would be expected to be wild type, 
whereas half of the males would be variegated and the other half, wild type if 
the duplication for the tip of 2R survived; but there would be a skewed sex ratio 
with approximately twice as many males as females. Genetic crosses confirmed 
the prediction made on the above assumptions. 

A test was set up to determine whether the 2.Y chromosome could survive 
heterozygously without Y.2 by crossing lOC-2 males which were heterozygous 
for nubbin^ wings to attached-X females carrying yellow and forked on the two 

^ Nubbin is a second chromosomal recessive gene located at approximately 43 on the crossover 
map. It results in very small wings which are thin, with entire margins and venation essentially 
unchanged, but with a tendency to curve upward or downward. The halteres are somewhat 
reduced. (Mickey, D.I. S.— 23). 

X chromosomes and homozygous for nubbin on the second chromosomes as fol- 
lows: X/Y.2;2(nub)/2.Y males x yf/Y;2{nub)/2(nub) females from which F^ 
males of two classes were expected: 

X/Y;2.Y/2 (nub) and X/Y;2inub) /2(nub), 
thus all non-nubbin males should be 2.Y /2(nub) and therefore should be de- 
ficient for the tip of 2R (that is, barring non-disjunction of X and Y.2). The 
actual crosses produced all nubbin males; no non-nubbin males survived. These 



s 


$X;2 


X;2 


X/X;2/2 


X;2.Y 


X/X;2/2.Y 


Y.2;2 


X/Y.2;2/2 


Y.2;2.Y 


X/Y.2;2/2.Y 



1 02 The University of Texas Publication 

genetic results were confirmed by cytological observations. Salivary gland prepa- 
rations showed the condition of Y.2 without 2.Y to be present, but the reverse was 
never observed; in other words, the presence of 2R in triplicate was viable but 
in the hemizygous condition it was lethal. 

Another test was devised to check the viability of combinations deficient for 
the tip of 2R, which involved crossing lOC-2 males with females homozygous for 
speck on the second chromosome, as follows: 

X/Y.2;2(+)/2.Y males x X/X;2(5p)/2(^yo) females 
which should give rise to four types of zygotes: 

1. X/Y.2; 2(5yo)/2( + ) = wild type male; sterile because deficient for part of 

Y; duplication of tip of 2R 

2. X/Y.2; 2(5p/2.Y = lOC-2 mottled male 

3. X/X; 2(^y[?)/2( + ) = wild type female 

4. X/X; 2(sp) /2.Y = lethal because deficient for tip of 2R. 

If class 4 flies were viable then females showing speck should have appeared, 
since the gene would have been hemizygous with no wild type allel to cover it. 
No such females were recovered, which indicated that the loss of the end of 2R 
reproduced lethality. Again, twice as many males as females were expected if 
type 1 males, which carried a duplication for the tip of 2R were viable. One half 
of the males should have been variegated and the other half should have been 
wild type. The cross actually yielded a one to one ratio. 

From the above cross the F^ wild type females (type no. 3) were mated in- 
dividually to lOC-2 males which were heterozygous for speck: 
2X/X;2(^p)/2( + ) X ^X/Y.2;2(^yo)/2.Y. 

The zygotes produced by this cross should have been as follows: 

a. X/X;2{sp) /2(sp) = speck iemale 

b. X/X;2(5yo)/2.Y = lethal because deficient for tip of 2R 

c. X/X;2{ sp)/2( + ) = wild type female 

d. X/X;2( + )/2.Y = lethal because deficient for tip of 2R 

e. X/Y.2;2(sp)/2(sp) = wild type male; sterile because deficient for part 

of Y; duplication of tip of 2R. 

f. X/Y.2;2{sp)/2.Y -10C-2male. 

g. X/Y.2;2( + )/2(sp) = wild type male; sterile because deficient for part 

of Y; duplication of tip of 2R. 
h. X/Y.2;2(+)/2.Y =10C-2male. 
Therefore the ratio expected among females was one speck to one wild type, and 
such a ratio actually was recovered. Among males the expected ratio of one wild 
type to one variegated male was obtained. 

When class 2 males were crossed with speck virgins, the following results 
were obtained: 

Cross: $X/Y.2-2(sp)/2.Yx 9X/X;2(sp) /2{sp) 
Progeny: a. X/X;2(sp) /2(sp) = speck female 

b. X/X;2(sp) /2.Y = lethal; deficiency 2R 

c. X/Y.2;2{sp)/2{sp) = wild type male; duplication 2R; sterile 

d. X/Y.2;2(sp)/2.Y = = lOC-2 male. 



Mickey: Variegated Position Effects 103 

The females were all speck and non- variegated, whereas the ratio among males 
was one wild type to one variegated; but the sex ratio was skewed again, being 
two males to one female. 

Still another genetic test indicated that the 2.Y chromosome without the Y.2 
produced an inviable combination. When lOC-2 males were crossed with females 
heterozygous for the dominant markers Curly and Plum the following results 
were obtained: 



Cross: 5X/Y.2;2( + )/2.Yx 


2 X/X;Cy/Pm 


Progeny: a. X/X;Cr/+ 


= Curly females 


b. X/X;Pm/+ 


= Plum females 


. c. X/X;Cr/2.Y 


- lethal 


d. X/X;Pm/2.Y 


= lethal 


e. X/Y.2;Cr/+ 


= duplication tip 2R; sterile, deficient for 




part of Y 


f. X/Y.2;Pm/+ 


= dupHcation tip 2R; sterile, deficient for 




part of Y 


g. X/Y.2;Cr/2.Y 


= Curly mottled males 


h. X/Y.2;Pm/2.Y 


= Plum mottled males 



Since no variegated females appeared and since both Curly mottled and Plum 
mottled males occurred, the prediction was borne out. 

VARIEGATED FEMALES 

Variegated females occasionally appeared in crosses of lOC-2 males with wild 
type females as a consequence of non-disjunction; but when lOC-2 males were 
crossed with virgin females from an attached-X stock, variegated females regu- 
larly appeared. This was possible since the attached-X females carried a Y 
chromosome and those females receiving both Y.2 and 2.Y could survive and 
produce mottling of eye color. The ratio obtained was one variegated female to 
one wild type female, but all males were wild type. 

NON-DISJUNCTION 

Non-disjunction occasionally occurs involving the X chromosome and the Y.2 
chromosome thus producing a female with variegated eyes. In crosses between 
lOC-2 males and wild type females normal segregation produced a ratio of one 
variegated male to one wild type female as diagrammed earlier. The non-disjunc- 
tion zygotes would be: 

1. X; 2/2. Y = lethal; deficient for 2R and most of Y chromosome 

2. X; 2/2 = wild type male; sterile because the Y is missing. 

3. X/X/Y.2; 2/2 = wild type female carrying Y.2 but not 2.Y and there- 

fore not variegated; but also duplication for tip of 2R. 

4. X/X/Y.2; 2/2.Y = variegated female carrying both Y.2 and 2.Y 

This last type is a variegated female who was produced through primary non- 
disjunction and whose germ cells would give rise to secondary non-disjunction. 
From a cross of lOC-2 male with a variegated attached-X female a few solid 
brown-eyed males and females were obtained. Presumably such individuals 



104 The University of Texas Publication 

could arise by non-disjunction giving genotypes of X/Y.2/Y.2; 2.Y/2.Y for the 
males and ZX /Y.2/Y.2; 2.Y/2.Y for the females, which are homozygous for 
both Y.2 and 2.Y chromosomes. These may be termed homozygous translocations 
or structural homozygotes. The rate of non-disjunction in males was 1 in 1,212 or 
0.0823 per cent. On the other hand the frequency of non-disjunction females was 
7 in 857 or 0.82% which is a hundred times greater. The non-disjunction must 
occur coincidentally with the segregation of the 2.Y chromosome in the same 
cell in order to be detected. 

DISCUSSION 

The variegation exhibited by the 1 OC-2 mutual translocation between 2R and 
the Y chromosome is the result of a break very near the brown locus and placing 
the heterochromation of the Y chromosome adjacent to the brown locus. This 
locus (2-104.5) was first shown by Glass (1933) to be sensitive to chromosomal 
rearrangements. Slatis (1955a) stated that the brown locus is most likely in the 
region between 59D8 and El-2 on Bridges map, but Demerec (1941) gave in- 
formation indicating that the locus might be anywhere within region 59. The 
break in lOC-2 occurred at 59C6. 

SUMMARY AND CONCLUSIONS 

1. The lOC-2 stock contains a mutual translocation between the right arm 
of the second chromosome and the long arm of the Y chromosome. 

2. The variegation or mottling is less obvious in females than in males. 

3. Any male containing a Y.2 chromosome without a 2.Y chromsome is 
viable but sterile. 

4. Males containing both Y.2 and 2.Y chromosomes are viable and fertile. 

5. A fly (male or female) is in viable when it contains a 2.Y chromosome with- 
out a Y.2 chromosome. 

6. Only those flies containing both 2.Y and Y.2 chromosomes have variegated 
eyes. 

7. Appropriate crosses can produce flies which are homozygous for both Y.2 
and 2.Y chromosomes. Such individuals have almost solid brown eyes, in which 
the variegation is barely detectable. 

8. Non-disjunction between the X and Y.2 chromosomes occurs occasionally, 
producing a variegated female. 

LITERATURE CITED 

Baker, W. K. 1952. Position effects of a gene normally located in heterochromatin. Genetics 37: 
564. 

. 1953. V-type position effects of a gene in Drosophila virilis normally located in 

heterochromatin. Genetics 38: 328-344. 

Bridges, C. B. 1935. Salivary chromosome maps. With a key to the banding of the chromosomes 
of Drosophila melanogaster. J. Hered. 26:60-64. 

Demerec, M. 1941. The nature of the gene. Proc. Univ. Pa. Bicentennial Conference 1941:1-11. 

Glass, H. B. 1933. A study of dominant mosaic eye colour mutants in Drosophila melanogaster. 
II. Tests involving crossing-over and non-disjunction. J. Genet. 28:69-112. 



Mickey: Variegated Position Effects 105 

. 1934. A study of dominant eye color mutants in Drosophila melanogaster. I. Pheno- 

types and loci involved. Amer. Nat. 68: 107-1 14. 
Hannah, A. 1951. Localization and function of heterochromatin in Drosophila melanogaster. 

Adv. in Genetics 4:87-125. 
Hinton, Taylor. 1949. The modification of the expression of a position effect. Amer. Nat. 83: 

69-94. 
Lewis, E. B. 1950. The phenomenon of position effect. Adv. in genetics 3:73-115. 
Mickey, G. H. 1954. Cytological analysis of mottled, variegated and dominant Minute mutations 

produced in Drosophila melanogaster by X-rays, gamma rays and fast neutrons. Anat. Rec. 

120:727. 
_, 1955. Further analyses of position effects in Drosophila melanogaster. Genetics 40: 

585. 
Schultz, J. 1936. Variegation in Drosophila and the inert chromosome regions. Proc. Nat. Acad. 

Sci' 22:27-33. 
Slatis, Herman M. 1955a. Position effects at the brown locus in Drosophila melanogaster. Genetics 

40:5-23. 
. 1955b. A reconsideration of the brown-dominant position effect. Genetics 40:246-251. 



Telomeres and Terminal Chiasmata — A Reinterpretation 

M. J. D. WHITE 
Department of Zoology, University of Melbourne, Australia^ 



The phenomenon of chiasma terminahzation (movement of the chiasmata, or 
some of them, towards the ends of the chromosomes) is a very general, but 
probably not universal feature of the late prophase of meiosis. First clearly 
recognized as a distinct process by Darhngton (1929), it has recently been dis- 
cussed by Swanson (1957, pp. 214-218). Darlington (1932) defined the process 
as "expansion of the association of the two pairs of chromatids on one side of a 
chiasma at the expense of that on the other side." When a chiasma terminahzes 
right to the end of a bivalent it becomes a "terminal chiasma," i.e., an association 
of two chromosome ends (each consisting of two chromatids) with a double con- 
necting thread between them. In some species a chromomere or minute dark- 
staining enlargement is seen in the middle of each connecting thread (Fig. 1). 





Fig. 1. Terminal chiasmata. Maternal and paternal chromatids indicated in a and b. On the 
interpretation put forward here, all terminal chiasmata are of type a; on Darlington's interpre- 
tation a configuration like b can arise through terminahzation of a pair of compensating chias- 
mata to the same chromosome end. c, a terminal chiasma with chromomeres in the middle of the 
connecting threads (presumably each such chromomere consists of two halves, one belongmg to 
each chromosome). 

Usually, the connecting threads between the chromosome ends are broken at 
first anaphase. But in some Hemiptera that have pre-reductional separation at 
the terminal chiasma (White, 1954, fig. 97) the chromatids are associated in 
pairs by "half terminal chiasmata" all through the succeeding interphase and 
until the second anaphase. 

The theory of terminal chiasmata which was put forward by Darlington was 
that a special "terminal affinity" existed between chromosome ends. This force 
of attraction was supposed to develop after the early diplotene stage, because if 
it were present at the beginning of diplotene, all chromosome ends would remain 
associated and every bivalent would show two terminal associations in addition 

1 Dedicated with esteem and admiration to Professor J. T. Patterson— and in memory of the 
years 1947-1953 spent by the author at the University of Texas. 



108 The University of Texas Publication 

to true interstitial chiasmata. This assumption of a special affinity between chro- 
mosome ends, which manifests itself only at a particular stage of meiosis and 
presumably only following chiasma formation, seems to be a very special and 
unlikely one. It was put forward before the difference between the properties of 
natural chromsome ends or telomeres and freshly broken ends had been recog- 
nized (Muller, 1932, 1938). 

The essential argument on which the idea of terminal affinity is based rests on 
the "observation" that compensating chiasmata (i.e. reciprocal or complement- 
ary pairs — two strand doubles and four strand doubles in the terminology of the 
Drosophila workers) do not "cancel out" when they reach the same chromosome 
end — i.e., that when a compensating pair of chiasmata terminalize to the same 
end the result is a terminal association ("terminal chiasma") and not a mere 
falling apart of the ends. 

But is this really so? No critical evidence ever seems to have been presented 
on this point. Most cytologists seem to have accepted Darlington's interpretation, 
probably because they have assumed that ( 1 ) compensating pairs of chiasmata 
are common, (2) terminalization is common, (3) falling apart of chromosome 
ends at diplotene to first metaphase is very rare or non existent. 

In figure 2 we show an example of a cell which we interpret as containing a 
pair of chromosomes which were held together at diplotene by a pair of compen- 
sating chiasmata but have cancelled out by first metaphase. That such cells are 
not common is understandable; if they were, a significant amount of non- 
disjunction would undoubtedly result in most types of meiosis. If they occur at 




Fig. 2. A first meiotic metaphase (in side view) in an individual of the grasshopper Trimero- 
tropis gracilis. This particular individual was heterozygous for the Rochester and Standard 
sequences of chromosome 7. Chromosome 4 consists of two univalents situated opposite one 
another, as if they had been paired at an earlier stage. It is suggested that they were associated by 
a pair of reciprocal chiasmata in the same arm which "cancelled out" at premetaphase. But on 
account of the hooked shape of the distal ends of the separating chromosomes it cannot be regarded 
as certain that the two chiasmata terminalized right to the end of the bivalent before cancelling 
out. 



White: Telomeres and Terminal Chiasmata 109 

all, however, they are evidence against Darlington's theory of the terminal chi- 
asma, at least in its original form. 

Callan (1949) in a popular article put forward very briefly a suggestion which 
amounts to a radical alternative to Darlington's theory of terminal chiasmata. 
Because of the way in which it was published, this suggestion seems not to have 
received the attention it deserves. Although we do not think it can be accepted 
precisely in its original form, we believe it points the way to a correct interpre- 
tation. 

Briefly, Callan's suggestion is that the chromosome tips are physically un- 
divided at the stage when terminal chiasmata are seen. If this were so, a terminal 
chiasma would indicate "equational" separation of chromosome ends. It would 
result whenever a single chiasma became terminalized — or when a pair of di- 
agonal chiasmata terminalized to the same chromosome end. On the other hand, 
if two compensating chiasmata were to terminalize to the same end there would 
be no terminal association formed and the ends would simply fall asunder 
"reductionally" (which is what we believe has happened in the cell illustrated 
in figure 2) . 

The idea of the chromosome end or telomere being simply undivided at a time 
long after replication of the rest of the chromonema has been completed is pos- 
sibly a somewhat awkward one from the biochemical standpoint. But there 
seems no biochemical objection to some kind of sister strand union between 
chromatid ends that have already replicated. But the chemical bonding involved 
must be more labile than in true sister strand union; since it is broken at the 
end of first metaphase, when the chromosome ends are torn asunder on the 
elongating spindle. There is no absolutely convincing evidence whether the 
breaking apart of the associated chromosome ends is due simply to tension im- 
posed by stretching of the spindle, or whether some kind of enzymatic action is 
involved which breaks the special "telomeric bonding." But in certain organisms 
at any rate (Schrader, 1944; Hughes Schrader, 1947; Staiger, 1954) the telo- 
meric bonding which we postulate seems able to withstand a very violent stretch- 
ing at premetaphase and is then broken at the end of metaphase when stretching 
is apparently much less violent. We may therefore suspect that some specific 
chemical mechanism (a "telomere-ase") is responsible for breaking the telomeric 
bonds at the beginning of first anaphase. 

Another reason for believing that it is not mechanical tension which breaks 
the telomeric bonding is that metacentric chromosomes appear X-shaped at inter- 
kinesis even when there has been a chiasma in one arm only. The fact that the 
telomeric connections at meiosis are able to withstand the premetaphase tension 
suggests that relatively strong chemical bonding is involved; possibly disulfide 
bridges, since the structure would almost certainly have to be a symmetrical 
one. If the chromosome consists of many short segments bound together by 
calcium bridges (Mazia, 1954), then the telomeric connections are probably not 
due to calcium. 

If telomeres consist essentially of a special kind of sister strand union at mei- 
osis (or of chemical groupings capable of taking part in such unions), it is logical 
to suppose that the telomeres of somatic chromosomes are essentially similar. The 
existence of telomeres was postulated in the first place because natural ends of 



110 The University of Texas Publication 

chromosomes show no tendency to fuse with one another, or to undergo true 
sister-strand union — properties exhibited by artificial ends produced by chromo- 
some breakage. That telomeres should be unable to fuse in these ways is readily 
understandable if they consist, essentially, of a different kind of sister strand 
union, of a more labile type. At what stages of the mitotic cycle the telomeric 
connections would be between chromatids and at what stages they would be 
between sub-chromatids is not clear. But presumably, sub-chroma tid connections 
replace chromatid unions at a particular stage, probably during interphase. 

If the above hypothesis is essentially true, the chromosome ends produced by 
radiation breakage and those which result from mechanical breakage (stretching 
on the spindle) lack telomeric properties because they are incapable of forming 
the special labile type of sister-strand union. Instead, they form unions of a more 
permanent kind which are not broken at each mitotic and meiotic cycle. "Heal- 
ing" of broken ends (such as occurs in maize sporophytes, according to McClin- 
tock, 1942, and in the somatic chromosomes of Ascaris during the cleavage divi- 
sions) would represent a change at the broken surface, probably either a loss or 
a gain of some material, which leaves the exposed end able to take part in a 
telomeric type of sister strand union. 

Various cytologists have claimed to have seen telomeres as specially staining 
structures, at particular stages of mitosis or meiosis. But obviously there can be 
no critical evidence that what they saw were the telomeres themselves rather 
than heterochromomeres or other structures immediately proximal to them. The 
same argument applies to the chromomeres sometimes seen in the middle of the 
threads connecting chromosome ends united in a terminal chiasma (fig. \c). 
All that we are justified in concluding in the latter case is that the region immedi- 
ately proximal to the terminal chromomere possesses considerable elasticity, so 
that it can be greatly extended — not that the terminal chromomere is itself the 
telomere. The latter may be a single nucleotide or nucleotide pair, a single amino 
acid or even an exposed sulfhydryl group at the tip of the chromosome. Terminal 
adhesions between chromosome ends in polytene nuclei or at meiosis, such as 
have been reported for various organisms, are more likely to be due to the termi- 
nal chromomere, or to several chromomeres at the end of the chromosome rather 
than to the telomere in the strict sense we have been considering here. 

It is an observed fact that terminalization is usually much more extreme in 
organisms having a maximum of one chiasma per chromosome arm. On the sup- 
position that compensating chiasmata do not "cancel out" in terminalization, this 
fact was unexplained. But if they can do so, it is understandable that terminaliza- 
tion should be absent, poorly developed or confined to the distal chiasma (or, 
alternatively, to the proximal ones) in species with more than one chiasma per 
chromosome arm. If it were otherwise "accidents" such as that shown in figure 2 
would occur and meiotic nondisjunction would be too frequent. Thus we believe 
that regularity of disjunction is best served by extreme terminalization only 
where there is a maximum of one chiasma per arm and that selection has 
operated to produce these regularities of the meiotic mechanism. 

The ideas expressed here are put forward in a tentative manner only, as an 
attempt to re-open discussion on some fundamental problems of chromosome 
behavior. With the development of techniques for isolating the whole mitotic 



White: Telomeres and Terminal Chiasmata 1 1 1 

apparatus it is not too much to hope that some of these problems may soon yield 
to an experimental approach. 

LITERATURE CITED 

Callan, H. G. 1949. Article on Chromosomes in New Biology 7, pp. 70-88. 

Darlington, C. D. 1929. Chromosome behaviour and structural hj^bridity in the Tradescantiae. 

/. Genetics 21:207-286. 

. 1932. Recent Advances in Cytology. Blakiston. 

Hughes Schrader, S. 1947. The "pre-metaphase stretch" and kinetochore orientation in phasmids. 

Chromosoma 3: 1-21. 
Mazia, D. 1954. The particulate organization of the chromosome. Proc. Nat. Acad. Sci. 40:521- 

527. 
Muller, H. J. 1932. Further studies on the nature and causes of gene mutations. Proc. VI Int. 

Congr. Genet. Ithaca 1:213-255. 
. 1938. The remaking of chromosomes. Collecting Net, Woods Hole. 13:181-195 and 

198. 
Schrader, F. 1944. Mitosis: The Movements of Chromosomes in Cell Division. Columbia U. Press. 
Staiger, H. 1954. Der Chromosomendimorphismus beim Prosobranchier Purpura lapillus in 

Beziehung zur Okologie der Art. Chromosoma 6:419-478. 
Swanson, C. P. 1957. Cytology and Cytogenetics. Prentice-Hall. 
White, M. J. D. 1954. Animal Cytology and Evolution. Cambridge U. Press. 



I 



Dominant Lethal Mutation in Irradiated Oocytes' 

D. R. PARKER 
Division of Life Sciences, University of California, Riverside, California 

Patterson, Brewster and Winchester (1932) in experiments dealing with 
dominant lethal effects, chromosome losses, etc., demonstrated some of the pos- 
sibilities for using Drosophila females in the study of radiation effects on genetic 
material. It is advantageous to be able to recover eggs known to have been in 
meiotic prophase when treated, and, since the completion of meiosis depends on 
sperm entrance, it is possible by holding females virgin to delay completion of 
this process by several days. Patterson and coworkers were able to show a con- 
siderable increase in radiation sensitivity with age. It is unfortunate that the 
significance of this frequently has been overlooked. 

Fuller utilization of Drosophila females as experimental material awaited the 
development of a simple procedure for screening for events depending on chromo- 
some breakage and rejoining. The successful interpretation of the nature of radi- 
ation-induced detachment of attached-X's made independently by Parker (1953, 
1954) and by Herskowitz and Muller (1953) and Herskowitz (1954) provided 
the needed technique. The morphological description of oogenesis by King, 
Rubinson and Smith (1956) has been found useful in work on the effects of aging 
on sensitivity. There has been good agreement between their descriptions of the 
distribution of stages and experimental findings in this laboratory. 

It is possible to distinguish at least four different levels of response in oocytes. 
The highest sensitivity is found in the oldest cells in females aged three or more 
days after eclosion. The oldest cells in newly emerged females show lower 
frequencies of rearrangement and of both dominant and recessive lethals. The 
next preceding stage will show about the same level of rearrangement (ie. de- 
tachment of attached X) at lower doses but will be destroyed by higher dosages 
(eg 4000 r). Younger than these are stages in which crossing over may be in- 
duced, but with no appreciable frequency of induced detachment of attached X's 
(Parker, in discussion of Whittinghill, 1955). The most useful stages are the 
ones designated as stages 7 and 14 by King, et al. The oldest oocytes in newly 
emerged females are in stage 7, while virgin females aged two or more days have 
one stage 14 oocyte in each ovariole. Thus, by limiting the number of eggs col- 
lected from any one female, a fairly homogenous sample of cells may be 
obtained. 

The differences in response of stages 7 and 14 to irradiation are both qualita- 
tive and quantitative. Stage 14 shows a higher incidence of all types of genetic 
damage which have been looked for, with break-rejoining delayed until fertiliza- 
tion, while breaks induced in stage 7 rejoin in about 10-15 minutes (Parker 
1955; Parker and Hammond, 1958; King, Darrow and Kaye, 1956). In stage 7 
most interchromosomal exchanges involved homologues or chromosomes which 

1 The experimental work reported here was carried out at the University of Mississippi, and 
was supported by Contract No. AT(40-1)-2163 with the U. S. Atomic Energy Commission. 



1 14 The University of Texas Publication 

presumably are partially homologous, as X and fourth chromosomes (Sandler 
and Novitski, 1956), while stage 14 yields relatively more interchanges between 
nonhomologous chromosomes (eg., X and second chromosomes) and more gross 
deficiencies (Parker and McCrone, 1958). Similarity, because of differences in 
shapes of the survival curves, there is reason to believe that the mechanisms of 
origin of dominant lethals may differ in these two stages (Parker, 1955; King, 
1957). 

A considerable literature has developed on hypotheses attempting to relate 
the origin both of dominant and of recessive lethals to chromosome breakage and 
rejoining. The resolution of these problems has been delayed in part by diffi- 
culties in interpretation of events taking place in mature Drosophila sperm. 
Perhaps the different conditions found in oocytes will permit the development of 
a fruitful approach to the problems of lethal mutation. This report represents a 
first attempt on the part of this writer to develop such an approach. 

TECHNIQUES OF EGG COUNTING 

In determining the frequencies of dominant lethals, eggs were collected from 
individual treated or untreated Oregon-R females over a period of approximately 
24 hours. Where aged females were treated, these were mated immediately with 
two Oregon-R males each. Females treated 0-12 hours after emergence were 
stored at room temperature (74°) for two days, then mated with two Oregon-R 
males each. In earlier experiments (Table 1 ) matings were made in 25 x 95 mm 
vials containing the ordinary culture medium of the laboratory to which 
powdered charcoal had been added to give a light gray color to facilitate egg 
counting. In all later experiments, because of difficulties in making counts in 
these vials, etherized flies were placed in empty vials which were bound together 
in groups of seven and inverted on petri dishes containing the charcoal medium. 
Following a suggestion of Dr. C. W. Edington, mycostatin was used to inhibit 
yeast growth. A solution of 250,000 units of the antibiotic in 200 ml of 50% 
ethyl alcohol was sprayed lightly on the surface after eggs had been laid. About 
26-28 hours were allowed for egg hatching, after which egg counts were made, 
tabulating totals of hatched and of unhatched eggs. Where necessary, plates 
could be stored at about 5° until counts could be made. At this temperature, 
larvae are killed and counts can be delayed without affecting the accuracy of 
the data. Mycostatin was not found to affect egg hatchability, as Oregon-R gave 
95.4 ± 1.2% hatch with mycostatin and 95.2 ± 1.1% without. 

The number of eggs any individual female could lay for the count to be tab- 
ulated was 10-30 inclusive. Any group of eggs in which none hatched was con- 
sidered the product of an uninseminated female. Any error introduced by this 
assumption, at the dosage used, would be smaller than the one introduced by 
considering this the result of complete killing. Where more than 30 eggs were 
laid, some may have been in earlier stages at the time of treatment. As there were 
few cases of exclusion of counts, about equally frequent at each dosage used, 
these exclusions do not change the percentage hatch very appreciably.. 



Parker: Dominant Lethal Mutation 115 

DOSE RESPONSE IN DOMINANT LETHAL PRODUCTION 

The first series of egg counts involved stage 7 material which had been 
irradiated with 250 KVP X-rays, with 3 mm Al-filtration, at a dose rate of about 
300 r/minute. Through an error in calibration of the dosimeter, there is prob- 
ably an error in total doses given, but the error factor would be common to all 
points. The remaining treatments, both stage 7 and 14, were given with a Snook 
machine operated at 60 KVP (1 mm Al-filter) at a dose rate of 100 r/minute. 
All treatments were based on readings taken with a Victoreen r-meter which 
had been calibrated against a known Cobalt-60 source at Oak Ridge National 
Laboratory. The results are given in Table 1. Figure 1 shows the shapes of the 
survival curves, and includes points plotted from the data of Sonnenblick (1940) 
for stage 14. There is a marked difference in sensitivity of the two stages, as well 
as in shapes of the two survival curves. These have been interpreted as "two-hit" 
and "one-hit" curves, respectively for stage 7 and stage 14. Sonnenblick's points 
show agreement with the theoretical curve at lower doses. Above about 1000 r, 
his points lie above the line, giving a curve suggesting a mixture of cells with 

Table 1 
Dominant lethal dose response 



Stage 
Treated 


Dose 
in r-units 


Number 
Eggs 
Laid 


Number 

Eggs 
Hatched 


Percentage 
Hatch ± S.E. 


Corrected 

Percentage 

Hatch 


7 


control 


467 


456 


97.6+0.7 


100.0 


7 


900 


466 


444 


95.3±1.0 


98.7 


7 


1,800 


317 


^ 271 


85.5+2.0 


87.6 


7 


3,600 


318 


184 


57.9+2.8 


59.3 


7 


control 


392 


358 


93.9+1.2 


100.0 


7 


1,500 


.. • 549 


459 


83.6+2.0 


89.0 


7 


3,000 


424 


269 


63.3+2.3 


67.4 


7 


6,000 


197 


64 


32.5+3.3 


34.6 


7 


control 


447 


415 


92.8+1.2 


100.0 


7 


700 


930 


848 ■ 


91.2+0.9 


98.2 


7 


1,400 


517 


428 


82.8 + 1.7 


89.2 


7 


2,800 


765 


487 


63.7+1.7 


68.6 


7 


4,900 


543 


173 


31.9+2.0 


34.3 


14 


control 


395 


374 


94.7+1.1 


100.0 


14 


250 


522 


357 


68.4+2.0 


72.2 


14 


500 


279 


97 


34.8+2.9 


36.7 


14 


control 


1,054 


952 


89.5+0.9 


100.0 


14 


50 


574 


503 


87.6+1.4 


97.9 


14 


100 


568 


430 


75.7+1.8 


84.6 


14 


150 


383 


265 


69.2+2.4 


77.3 


14 


200 


645 


405 


62.8+1.9 


70.2 


14 


250 


252 


146 


57.9+3.1 


64.8 


14 


300 


400 


208 


52.0±2.5 


58.1 


14 


500 


233 


83 


35.6+3.1 


39.8 


14 


600 


321 


80 


24.9+2.4 


27.8 



116 



The University of Texas Publication 

FIGURE I 



0.9 

o.a 

0.7 
0.6 
0.8 



> 




K 




3 




CO 




z 


0.1 


o 


0.O9 


H 


o.oe 


o 




< 


0.07 


tc 




u. 


0.06 




0.0S 




0.03 



DOSE IN kr 

Fig. 1. Survival curves for irradiated oocytes. Triangles represent three replications of the 
experiment irradiating stage 7 material. Circles represent irradiated stage 14 material, the filled 
circles being taken from the data of Sonnenblick (1940) . 

two markedly different sensitivities. His egg collections may have included a few 
which had been pre-stage 14 when treated. King, Darrow and Kaye (1956) 
show the same types of curves, again with a marked concavity in the stage 14 
curve. 



EFFECT OF VARYING THE 
INTENSITY OF IRRADIATION ON SURVIVAL 

Knowledge of the time required for chromosome break rejoining in the two 
treated stages of oogenesis would allow some predictions to be made of effects of 
dose-fractionation on the frequency of dominant lethals which might depend on 
the rejoining of broken ends of chromosomes. Parker and Hammond (1958) con- 
cluded that breaks in stage 7 rejoin in about 10-15 minutes; those in stage 14 
rejoin about the time of fertilization. Thus, the conditions in stage 7 would lead 
one to expect an intensity effect if the "two-hit" dominant lethals depended on 
the interaction of two independently induced breaks. On the other hand, failure 



Parker: Dominant Lethal Mutation 117 

of early rejoining of breaks in addition to the "one-hit" nature of the response 
in stage 14 would lead to the prediction of no intensity effect. 

Irradiations in this series were made with a G. E. Maximar-100 machine 
operated at 100 KVP with a 1 mm Al-filter, giving a HVL of 1.3 mm Al. Dosages 
were monitored as in the previous irradiations. The dose rate was approximately 
300 r/minute. The dosage used in stage 7 was 3000 r, while that in stage 14 was 
300 r. These were chosen as each will give around 60 percent survival. Treat- 
ments were either given in one continuous dose or were divided into two equal 
fractions with a measured time interval between treatments. Table 2 gives the 

Table 2 

Effects of dose fractionation on dominant lethality: Dosages divided in two equal fractions 
separated by variable time interval 

Interval Number Number 

Stage Total Between of Eggs of Eggs Percentc^ge Percentile 

Treated Dose Fractions Laid Hatched Hatch ± S.E. Increase 

7 3,000 1,091 599 54.9±1.1 

7 3,000 45min. 1,219 753 61.8±1.4 6.9±1.8 

7 .. 172 161 93.6+1.9 . .... 

7 3,000 350 202 57.7±2.6 

7 3,000 15min. 305 200 65.4±2.7 7.7±3.8 

7 3,000 30min. 277 156 68.7±3.1 11.0±4.1 

7 3,000 60min. 309 222 71.8±2.6 14.1±3.7 



3,000 





1,091 


599 


3,000 


45 min. 


1,219 


753 







172 


161 


3,000 





350 


202 


3,000 


15 min. 


305 


200 


3,000 


30 min. 


277 


156 


3,000 


60 min. 


309 


222 







721 


693 


3,000 





2,454 


1,460 


3,000 


30 min. 


2,848 


1,905 


300 





961 


524 


300 


15 min. 


828 


456 


300 


30 min. 


1,045 


558 


300 





2,447 


1,320 


300 


4 hours 


619 


300 



7 .. 721 693 96.1+0.7 

7 3,000 2,454 1,460 59.5 ±1.0 

7 3,000 30 min. 2,848 1,905 66.9±0.9 7.4±1.3 

14 300 961 524 54.5±1.7 

14 300 15 min. 828 456 55.1±1.7 0.6±2.4 

14 300 30 min. 1,045 558 53.4±1.5 —1.1 ±2.3 

14 300 2,447 1,320 53.9±1.0 

14 300 4 hours 619 300 48.5±2.0 — 5.4±2.2 



results of these treatments. There seem to be effects of the type expected, with 
significant increases in percentage of hatch in the stage 7 series where the in- 
terval between fractions was long enough (10-15 minutes) to allow restitution 
and rejoining to occur in the first breaks produced before the second group were 
induced. As expected, fractionation did not increase survival in stage 14. The 
control values (continuous treatment) are somewhat lower in the stage 7 ma- 
terial than those given in Table 1. Perhaps this was due to the higher intensity 
used (300 r/minute vs. 100 r/minute) . 

CENTRIFUGATION 

Sax (1943), and Wolff and von Borstel (1954) have shown that aberration 
frequency in Tradescantia and in Vicia may be increased by post irradiation 
centrifugation which might move broken ends of chromosomes apart making 
restitution less likely, while centrifuging before irradiation lowered aberration 



118 The University of Texas Publication 

frequency, presumably by a packing effect which lowered mobility of the 
chromosome fragments produced by subsequent irradiation. 

Drosophila females can withstand centrifugal forces of the order of 1200 g 
without any observable effect on fecundity or hatchability of non-irradiated eggs. 
Small numbers of females were placed in a paper capsule closed Vv^ith a soft 
cotton plug. The capsule was then placed in the centrifuge tube with the plug 
down. The centrifuge used had a radius of six inches, and was operated at ap- 
proximately 2600 rpm for 5 or for 20 minutes giving a RCF of about 1150 g. 
Flies were centrifuged either immediately before or after irradiation, otherwise 
the conditions of irradiation were similar to those in the fractionation series: 
Stage 7 was given 3000 r; stage 14 received 300 r. The results are found in 
Table 3. 

Table 3 

Effects of centrifuging (1 150 g) on the induction of dominant lethals. 
(B, before and A, after irradiation) 

Nmnber Nvunber 

Eggs Eggs Percentage Percentile 

Stage Dose Centrifuging Laid Hatched Hatch ± S.E. Decrease 



7 





20 min. 


1,177 


1,101 


93.5+0.7 




7 


3,000 


none 


3,153 


1,701 


53.9+0.9 




7 


3,000 


A-20 min. 


2,329 


931 


40.0±1.0 


13.9+1.3 


7 


3,000 


none 


1,188 


733 


61.7±1.4 




7 


3,000 


A-20 min. 


818 


381 


46.6±1.7 


15.1+2.2 


7 


3,000 


B-20 min. 


800 


471 


58.9+1.7 


2.8±2.2 


7 


3,000 


none 


827 


423 


51.2+1.7 


.... 


7 


3,000 


A-5 min. 


1,043 


529 


50.7+1.5 


0.5+2.3 


14 


300 


none 


1,648 


770 


53.3+1.3 




14 


300 


A-20 min. 


1,661 


827 


49.8+1.2 


3.5+1.8 



No effect of pre-irradiation centrifuging can be seen, while post-irradiation 
centrifuging resulted in a lowered hatchability of stage 7 only. Since few inter- 
chromosomal exchanges involving non-homologous chromosomes are produced 
in this stage, little "packing effect" would be expected, and the absence of a 
pre-irradiation centrifugation effect is not too surprising. Breaks in stage 14, on 
the other hand, remain open for a long time after being centrifuged, and the 
small amount of movement resulting from centrifugation might have little 
effect on the distance between broken ends at a time when rejoining becomes 
possible. The results are, therefore, compatible with the idea that chromosome 
breakage is a factor in the production of a significant fraction of dominant 
lethals. 

HYPOTHESES OF DOMINANT LETHALS 

If dominant lethals arise as a result of chromosome breakage, what events 
must follow for the lethal change to be effected? The low frequency of inter- 
chromosomal aberrations induced in females (eg. Glass 1955) suggests that 
mechanisms involving aneucentric rearrangements between nonhomologous 



Parker: Dominant Lethal yjutation 119 

chromosomes play at best only a minor role in these cases. Particularly must this 
be the case with the "one-hit" dominant lethals in stage 14, the stage where a 
somewhat higher frequency of translocation between nonhomologous chromo- 
somes might be expected (Parker and Hammond, 1958). Precisely the same 
argument can be applied to large deficiencies. 

The suggestion was made by A. R. Whiting (1945) for Habrobracon, that 
dominant lethals may arise through formation of bridges in anaphase II of 
Meiosis. This interpretation was later adopted as a working hypothesis by 
Parker (1955) and in a modified form by King (1957). 

The strong inference that rearrangement depends in part on the proximity of 
breaks, leading to the low incidence of translocation in females, would permit 
the occurrence of rearrangements involving breakage and rejoining in 
chromatids, either sisters or homologues in early prophase; in late prophase per- 
haps only sisters or chromatids effectively behaving as sisters, i.e. homologues 
lying together distal to a crossing-over point but proximal to the resultant termal- 
izing chiasma. 

An exchange giving rise to a dicentric chromosome and an acentric fragment 
might yield a dominant lethal especially if the resulting bridge were in anaphase 
II and involved the prospective egg nucleus. Anaphase I bridges might be 
eliminated in the same manner as when formed by crossing over in inversion 
heterozygotes (Sturtevant and Beadle, 1936), although anaphase I bridges 
formed in autosomes might not be eliminated since apparently only acrocentric 
chromosomes give this result (Novitski, 1952) . 




H " S 

Fig. 2. Two possible modes of origin of Anaphase II bridges. S, sister union; H, aneucentric 
rejoining of homologues distal to a chiasma. 

THE PRODUCTION OF COMPOUND X'S BY IRRADIATION 

The description by Novitski (1954) and Sandler (1954) of the origin of 
reversed acrocentric compound-X chromosomes suggested a technique of ap- 
proach. Reversed acrocentrics arise spontaneously with a low frequency in fe- 
males heterozygous for a long inversion (scute^) if the normal-sequence X 
chromosome has the long arm of the Y attached. In the absence of an attached 
arm, this type of recombination does not take place. A great many of Sandler's 



120 



The University of Texas Publication 



cases showed homozygosis for markers near the point of union of the two X's, 
whereas the mother had been heterozygous for those loci. This showed that two 
meiotic events had been necessary for the orgin of these chromosomes: sister 
union in the proximal heterochromatin of one, with an appropriate crossover 
distal to that point. 

Since the hypothesis of dominant lethals involves the formation of sister 
unions following breakage of two sister chromatids, the irradiation of inversion 
heterozygotes with no attached Y arms suggested itself as a means of testing for 
these radiation-induced sister unions. 

Females of the composition y car bb/ sc^ w^ cv v f were irradiated and mass 
mated to "Muller 5" {sc^^ B In S w"" sc^) males, and allowed to lay eggs for 24 
hours. These females were irradiated either within 12 hours after emergence, or 
5 days later, giving results on stage 7 and stage 14 oocytes respectively. All non- 
Bar female offspring were tested for the presence of a compound X by remating 
to Muller 5 males — these females were recorded as to phenotype, and for loci 
which became homozygous in subsequent generations. Various dosages were 
used in the treatments in the hope of distinguishing "one-hit" and "two- hit" 
effects. 

Two series of irradiations were made on Stage 7 material, and one on stage 14. 
The results of these irradiations are given in Table 4. 

It would be difficult to conclude from these data whether or not there is a 
two hit effect in stage 7. In both series, at lower doses the increases in frequency 
with dose seems to exceed the linear expectation, however, at higher doses this is 
not the case. From only two points, it would be impossible to interpret the 



Table 4 



Induction of compound X's by irradiation of inversion heterozygotes. 
(In (/) sc^ f V cv iv^/y car bb X Muller-5 {sc^'^ B InS w^ sc^) 



Stage 
Treated 


Dose 


Regular Classes 
Bar?? Non-Bard" d + 


Compound X 

y yf 


y car 


Non-disjunct ional 

Cl-sses 
Non-Bar?? Barcf d" 


Percentage 
Compound X's 


7 





508 


421 














7 


1,000 


720 


579 








1 






7 


1,500 


1,438 


1,281 


1 




1 


5 


3 


0.074 


7 


2,000 


438 


346 


1 




1 






0.25 


7 


3,000 


1,679 


1,061 1 


7 


2 




13 


4 


0.36 


7 


4,000 


256 


151 


2 










0.49 


7 


6,000 


120 


76 




1 


1 


1 


1 


1.0 


7 


750 


6,990 


4,610 


1 






20 


9 


0.008 


7 


1,500 


7,072 


4,655 


2 


3 


3 


6 


15 


0.07 


7 


2,050 


5,702 


3,883 


4 


1 




42 


7 


0.05 


7 


3,000 


4,366 


2,879 


4 


3 


1 


38 


31 


0.11 






Totals for stage 7 1 


26 


10 


12 








14 


250 


7,999 


6,096 


3 


7 


3 


2 


151 


0.09 


14 


500 


2,615 


1,787 




3 


2 


4 


104 


0.11 






Totals for 


stage 14 


3 


10 


5 









Parker: Dominant Lethal Mutation 121 

response of stage 14. However, as pointed out in the preliminary report (Parker, 
1957) the data are not inconsistent with the expectations of two-hit and one-hit 
for stages 7 and 14 respectively. 

It is impossible at present to make any valid estimate of the frequency of 
dominant lethals arising as a result of similar breakages and rejoinings. Meiotic 
drive (Sandler and Novitski, 1957) must limit the recovery of compound X's. 
Variations in the coefficient of non-randomness found in experiments of Novitski 
(1951), Novitski and Sandler (1956), from a low of about .67 to a high of at 
least .92 (with the possibility of much higher values not yet detected) would not 
allow a specific estimate of meiotic drive in the present case. 

It would be necessary, further, to make corrections for crossing over distal to 
the unions to get a measurement of their frequency in the case of compound X's, 
while crossing over proximal to these would affect their fates to dominant 
lethality. 

The events, to give rise to recoverable compound X's, must occur in hetero- 
chromatin and the resulting females must be fertile. These events seem to occur 
with different frequencies in the two X chromosomes used. There is no basis for 
estimating the frequency of their occurrence in euchromatic parts of 
chromosomes. 

However, in spite of these difficulties, some conclusions can be drawn. The fate 
of these unions should not be affected by the time of their production, as in 
neither case is meiosis carried beyond metaphase I before these events have 
occurred. There should, therefore, be similar differences in sensitivity of stage 7 
and of stage 14 oocytes, whether measured by dominant lethals or by production 
of compound X's. At the dosages used, for dominant lethals, doses giving similar 
survival differ by about a factor of 10, while for compound X production, there 
is about a three or four fold difference. 

The compound X's heterozygous for all markers occurred largely in the stage 
7 irradiations, there being 27 out of 49 in this stage as opposed to 3 out of 18 in 
stage 14. This might be expected if stage 14 represents late diplotene or diakinesis, 
with only sister chromatids being in contact in the proximal heterochromatin, 
and if stage 7 represents a much earlier prophase where wide separation of 
homologues has not yet occurred. 

One apparent anomaly is the high frequency of carnation homozygotes as 
compared to forked. The presumed origin of these is given in Figure 3. Recovery 
of the forked class (So) should be favored since the compound X will be separat- 
ing at anaphase II from an anaphase I bridge. On the other hand, the carnation 
class would be separating at anaphase II from scute 8, and its recovery would be 
affected adversely by meiotic drive. 

DISCUSSION 

Lea (1946) summarized the evidence available to him, concluding that much 
of the dominant lethal damage to irradiated sperm resulted from chromosome 
breakage. A. R. Whiting (1949) found that heavily irradiated eggs when fertil- 
ized by untreated sperm would give rise to patroclinous males, showing that 
death at ordinary dosages resulted from nuclear rather than from cytoplasmic 



122 



The University of Texas Publication 
y + 4- + + car ^ ^^ >vbb> 




Fig. 3. Possible modes of origin of compound X's by irradiation of inversion heterozygotes. 
S-^, sister union in the normal sequence; S.„ sister union in the inverted sequence; H, eucentric 
rejoining of homologues. 



damage. Also in Habrobracon, von Borstel and Moser (1956) were able to show 
that for lethal effect with ultra-violet the nucleus was more sensitive than the 
cytoplasm. Schwartz and Bay (1956) show that radiation doses high enough to 
prevent mitosis will allow cells to survive and for a while to carry on otherwise 
relatively normal metabolic activities, implying that death results from chromo- 
some damage which is expressed when the cell divides. 

On the other hand, von Borstel and Pardue (1956) and von Borstel (1958) 
have evidence that most radiation-induced dominant lethal effects occur earlier 
in development than does death from aneuploidy in the progeny of non- 
irradiated translocation heterozygotes and triploids. They suggest that dominant 
lethality may be associated with interference with DNA synthesis. However, 
their aneuploids differ from the kind postulated by classical dominant lethal 
theory: no anaphase cleavage bridges were involved. Until this difficulty can be 
overcome their argument must remain unanswered. It may be possible by 
studying the time of death of progeny of various kinds of compound X's where 
it is known that crossing over will produce anaphase II bridges with various 
frequencies (Novitski, 1954, 1955) to see where in ontogeny death will occur. 

Herskowitz and Schalet (1956, 1957) have presented evidence which they 



Parker: Dominant Lethal Mutation 123 

believe will account for at least 1/9 of the dominant lethal damage mduced at 
2000r as being the result of "half-translocation" and of non-disjunction and 
chromosome losses. Their estimate may be a good one in that it is not inconsistent 
with the figures given by von Borstel and Pardue (1956) , where about 80 percent 
of dominant lethals induced in Habrobracon oocytes die early as do about 
90 percent of those induced in Drosophila sperm, presumably leaving about 
10-20 percent for later death which could be due to simple aneuploidy. However, 
dominant lethals of this sort cannot account for the majority of deaths actually 
induced. It is definite that the response of stage 14 is "one hit," hence we may 
rule out "two hit" events as accounting for any appreciable number of dommant 
lethals in this stage. (Table 1; also King, Darrow and Kaye, 1956.) On the 
other hand, stage 7 gives "two hit" dominant lethals, as shown by the dose 
response curve and by the intensity effect (Tables 1 and 2; King, Darrow and 
Kaye, ibid) . However, the low frequency of interchromosomal rearrangements 
induced in this stage (Parker and McCrone, 1958) would tend to rule out the 
significance of the contribution of "half translocations" to dominant lethality in 
this stage. 

King (1957) has suggested a modified version of A. R. Whiting's (1945) hy- 
pothesis of induction of bridges in anaphase II of meiosis. He accounts for the 
origin of each bridge as a one hit event, and assumes that at least two bridges are 
necessary if induced by irradiation of stage 7, requiring the additional hypothesis 
that the egg nucleus is not oriented: either end nucleus may be able to function 
as the egg nucleus. He explains the "one hit" curve obtained with stage 14 as 
multiple breakage so that enough bridges are formed to insure some passing to 
opposite poles at anaphase I. 

There are several reasons for believing this hypothesis incorrect. Novitski 
(1954) in work dealing with the fate of anaphase II bridges showed that the pro- 
duction of nullo-X eggs by tandem-ring compound-X females might be explained 
as the result of loss of the X due to the formation of double anaphase II bridges. 
He was able to show that symmetrical double bridges lag and are lost on the 
spindle while single and asymmetrical double bridges are lethal presumably due 
to breakage and inclusion of a fragment in the egg nucleus. Since there is normal 
disjunction of compound X's at anaphase I, meiosis would always produce at 
least two bridge-free nuclei, and the phenomena of lethality and chromosome loss 
would not be expected unless one made the unlikely assumption that bridges aris- 
ing by crossing over behave differently from radiation-induced bridges. 

Evidence for the determination of the egg nucleus has been suggested by von 
Borstel (1957). Binucleate eggs occur occasionally in Habrobracon, while the 
corresponding binucleate oocyte has not been found. In stocks producing a high 
frequency of binucleate eggs (and in no others) meiosis frequently is not blocked 
at metaphase I, occurring in the interior of the egg rather than at the surface, 
allowing the possibility of survival of more than one nucleus, von Borstel postu- 
lates an agent, present in the cortical cytoplasm, which is responsible for disinte- 
gration of the polar nuclei, the egg nucleus escaping destruction by being pushed 
farther toward the interior by each of the meiotic divisions. 

Were King's hypothesis of a non-determined nucleus correct, it would be 
difficult to explain the very low frequency of binucleate eggs in non-irradiated 



124 The University of Texas Publication 

Drosophila. Some binucleate eggs would give rise to gynandromorphs and /or 
autosomal mosaics. Mosaics arising in this manner are much rarer than those 
involving chromosome loss. While the recognition of these may depend on the 
two egg nuclei being non-sister (i.e., product of the same first but a different 
second meiotic division), a higher frequency of involvement of sister nuclei 
would be of little consequence to the hypothesis of dominant lethals, as these 
would be the ones involved in single anaphase II bridges. 

Still another basis for at least partial rejection of King's hypothesis is the 
intensity effect reported here and by Abrahamson and Herskowitz (1955, 1957). 
If, as in King's hypothesis, all bridges are produced by "one hit" events, there 
would be no interaction between the two struck targets, hence one would predict 
a "two hit" curve but no intensity effect. 

Sonnenblick (1940) dealt with egg samples that were largely in stage 14 at the 
time of treatment, and there is very good agreement between his data and those 
reported here. Lea (1946) has used his description as a basis for the hypothesis 
that no rejoining of breaks occurred in oocytes, giving rise to cleavage bridges 
following sister union occurring after the completion of meiosis. The evidence 
supporting this hypothesis was the failure to obtain translocations by irradiation 
of females (e.g. Glass, 1955) . However, it was found that breaks induced in stage 
14 could rejoin (Parker and Hammond, 1958; Parker and McCrone, 1958), and 
Parker (1955) was inclined to reject the Lea interpretation because of the 
incorrectness of this assumption. There are reasons for modifying this position. 
Since rejoining is delayed until the removal of the block on meiosis, and meiosis 
is then completed rapidly, perhaps some breaks miss an opportunity to rejoin at 
this time. The failure to find an increase in stage 14 in induction of compound 
X's comparable to that found in dominant lethals casts serious doubt on the 
hypothesis that in stage 14 most dominant lethals arise following sister fusion of 
isochromatid breaks. Perhaps most breaks are chromatid breaks, and many dom- 
inant lethals do arise in the manner suggested by Lea and by Sonnenblick. Parker 
and McCrone concluded that translocations induced in stage 14 involve chroma- 
tids, as these show evidence of meiotic drive as would be expected if the re- 
arranged chromatid were separating from an intact one at anaphase II. If all or 
most of the breakage were isochromatid, one might expect either both sisters 
would be involved in each translocation, giving no basis for meiotic drive, or one 
sister might frequently fail to undergo restitution or rejoining and meiotic drive 
would operate in the opposite way to that actually found. 

We should then expect "dominant lethals" to be a mixture of events, possibly 
not all involving breakage and/or loss of genetic material. Non-disjunction and 
losses of whole chromosomes may be radiation-induced and therefore contribute 
somewhat to dominant lethal effects. Meiotic drive will favor the aneuploid 
segregation of any translocation which is induced. It is impossible at this time to 
make any valid estimate of the frequencies of these events, but the evidence of 
von Borstel and Pardue (1956) must limit these to not more than 10-20 percent 
of all dominant lethals produced at ordinary dosages. Undoubtedly anaphase II 
bridge formation occurs, although again determination of its frequency is pre- 
vented by the phenomenon of meiotic drive as well as by lack of information on 
breakage and rejoining in euchromatin in oocytes. We may expect these single 



Parker: Dominant Lethal Mutation 125 

anaphase II bridges to break (Novitski 1955) and their subsequent fate should 
be similar to that of broken chromatids which failed to restitute or rejoin before 
completion of meiosis. This final category of dominant lethals may constitute the 
great majority of the "one hit" deaths found in stage 14, and possibly make up 
a part of those produced in stage 7. This might account for the smaller than 
expected intensity effect found in this stage (Table 2). If failure to rejoin or 
restitute is a major cause of dominant lethality, then much of the difference in 
rate of production of dominant lethals in the stages worked with would be due to 
differential rejoining rather than breakage. Differences in the two stages may not 
be explainable on so simple a basis when rearrangement frequency is used as the 
measure of radiation sensitivity. 

Because of the probable variety of events leading to dominant lethal mutation, 
it would be difficult at this time to develop a mathematical treatment of the 
phenomenon. An equation based on the dual target model of King (1957) may 
give a fairly good superficial fit to the data, but this may be spurious, due to 
intensity effects operating differently in treatments of different durations, rela- 
tive amounts of "one-hit" and "two-hit" killing, and possibly a variety of other 
causes. 

SUMMARY 

Dominant lethals induced in stage 7 were found to follow a "two hit" curve, 
while those in stage 14 were "one hit." Evidence of participation of breakage and 
rejoining of chromosomes in the origin of dominant lethals is given by the finding 
that fractionation of the irradiation decreases while centrifuging after irradiation 
increases the incidence of dominant lethals in stage 7. Evidence that sister unions 
may occur following irradiation is given by the induction of compound X's by 
irradiation of inversion heterozygotes. This shows that anaphase II bridge forma- 
tion might account for some dominant lethality. Dominant lethals must arise 
through a variety of mechanisms, and there must be differences in the relative 
frequencies of the various kinds in the two stages studied. It is not possible to 
estimate the frequencies of the various events in oocytes which would lead to 
dominant lethality, hence no mathematical treatment is attempted. 

LITERATURE CITED 

Abrahamson, S. and I. H. Herskowitz. 1955. The effect of x-ray intensity and dose on egg mor- 
tality following irradiation of female Drosophila. D.I.S. 29: 101. 

. 1957. Induced changes in female germ cells of Drosophila. II. Oviposition rate and 

egg mortality in relation to intensity and dosage of x-rays applied to oocytes. Genetics 42: 
405-420. 

von Borstel, R. C. 1957. Nucleocytoplasmic relations in early insect development. The beginnings 
of embryonic development: 175-179. A.A.A.S., Washington, D.C. 

. 1958. Dominant lethal mutations in Habrobracon and Drosophila. Proc. X Int. 

Cong. Genetics 2:303. 
and H. Moser. 1956. Differential ultra-violet irradiation of the Habrobracon egg 



nucleus and cytoplasm. Progress in radiobiology: 211-215. Oliver and Boyd, Edinburgh. 
and M. L. Pardue. 1956. On the nature of radiation-induced dominant lethal muta- 



tions in Habrobracon and Drosophila. Genetics 41 : 665 (Abstr.) , 



126 The University of Texas Publication 

Glass, H. B. 1955. A comparative study of induced mutation in the oocytes and spermatozoa of 
Drosophila melanogaster. I. Translocations and inversions. Genetics 40:252-267. 

Herskowitz, I. H. 1954. The relation between x-ray dosage and the frequency of simulated heal- 
ing of chromosome breakages in Drosophila melanogaster females. Proc. Natl. Acad. Sci. 
U. S. 40:576-585. 

and H. J. Muller. 1953. Evidence against healing of x-ray breakages in chromosomes 

of female Drosophila melanogaster. Genetics 38: 669. (Abstr.). 

and A. Schalet. 1956. Half -translocations induced by irradiation of oocytes as a basis 



of dominant lethals in D. melanogaster. Genetics 41:647. (Abstr.). 

1957. Induced changes in female germ cells of Drosophila. V. The contribution of 



half-translocations and nondisjunction to the dominant lethality induced by x-raying oocytes. 
Genetics 42:649-660. 

King, R. C. 1957. The problem of dominant lethals in Drosophila melanogaster females. Proc. 
Natl. Acad. Sci. U. S. 43:282-285. 

, J. B. Darrovs^ and N. W. Kaye. 1956. Studies on different classes of mutation induced 



by radiations of Drosophila melanogaster females. Genetics 41 : 890-900. 

, A. C. Rubison and R. F. Smith. 1956. Oogenesis in adult Drosophila melanogaster. 



Growth 20: 121-157. 

Lea, D. E. 1945. Actions of radiations on living cells. Cambridge University Press. 

Novitski, E. 1951. Non-random disjunction in Drosophila. Genetics 36:267-280. 

. 1952. The genetic consequences of Anaphase bridge formation in Drosophila. 

Genetics 37:270-287. 

. 1954. The compound X chromosomes in Drosophila. Genetics 39: 127-140. 

. 1955. Genetic measures of centromere activity in Drosophila melanogaster. J. Cell. 



Comp. Physiol. 45, Supp. 2:151-169. 
and L. Sandler. 1956. Further notes on the nature of non-random disjunction in 



Drosophila melanogaster. Genetics 41: 194^206. 

Parker, D. R. 1953. Observations on x-ray induced detachments of attached-X chromosomes in 
Drosophila. J. Tenn. Acad. Sci. 28:185. (Abstr.). 

. 1954. Radiation-induced exchanges in Drosophila females. Proc. Natl. Acad. Sci. 

U. S. 40:795-800. 

. 1955. The origin of dominant lethals in irradiated oocytes of Drosophila. Genetics 



40:589. (Abstr.). 
. 1957. Radiation induction of compound X's in Drosophila oocytes. Genetics 42: 



387-388. (Abstr.). 

and A. E. Hammond. 1958. The production of translocations in Drosophila oocytes. 



Genetics 43:92-100. 

and J. McCrone. 1958. A genetic analysis of some rearrangements induced in oocytes 



of Drosophila. Genetics 43: 172-186. 

Patterson, J. T., W. Brewster and A. M. Winchester. 1932. Effects produced by aging and 
x-raying eggs of Drosophila melanogaster. J. Heredity 23:325-333. 

Sandler, L. 1954. A genetic analysis of reversed acrocentric compound X chromosomes in Dro- 
sophila melanogaster. Genetics 39:923-942. 

and E. Novitski. 1956. Evidence for genetic homology between chromosomes I and 

IV in Drosophila melanogaster, with a proposed explanation for the crowding effect in 
triploids. Genetics 41:189-193. 

. 1957. Meiotic drive as an evolutionary force. Amer. Nat. 91: 105-110. 



Sax, K. 1943. The effect of centrifuging upon the production of x-ray induced chromosomal aber- 
rations. Proc. Natl. Acad. Sci. U. S. 29: 18-21. 



Parker: Dominant Lethal Mutation 127 

Schwartz, D., and C. E. Bay. 1956. Further studies on the reversal in the seedling height dose 

curve at very high levels of ionizing radiations. Amer. Nat. 90:323-327. 
Sonnenblick, B. P. 1940. Cytology and development of the embryos of x-rayed adult Drosophila 

melanogaster. Proc. Natl. Acad. Sci. U. S. 26:373-381. 
Sturtevant, A. H., and G. W. Beadle. 1936. The relations of inversions in the X-chromosome of 

Drosophila melanogaster to crossing over and disjunction. Genetics 21 : 554-604. 
Whiting, A. R. 1945. Dominant lethality and correlated chromosome effects in Habrobracon eggs 

x-rayed in diplotene and in late metaphase I. Biol. Bull. 89:61-71. 
. 1949. Androgenesis, a differentiator of cytoplasmic injury induced by x-rays in 

Habrobracon eggs. Biol. Bull. 97: 210-220. 
Whittinghill, M. 1955. Crossover variability and induced crossing over. J. Cell. Comp. Physiol. 

45, Supp. 2: 189-220. 
Wolff, S., and R. C. von Borstel. 1954. The effects of pre- and post-irradiation centrifugation on 

the chromosomes of Tradescantia and Vicia. Proc. Natl. Acad. Sci. U. S. 40: 1 138-1 141. 



I 



Mammalian Chromosomes in Vitro, XI. Variability 
Among Progenies of a Single Cell' 

T. c. HSU 

Section of Cytology, The University of Texas M. D. Anderson Hospital 
and Tumor Institute, Houston 

Hsu and Klatt (1958) observed that in several cell strains of murine origin 
cells exhibited structural changes from normal mouse karyotype as well as hetero- 
ploidy. Prominent among the structural changes were the formation of dicentric, 
metacentric, subtelocentric, and minute chromosomes. Chu and Giles (1957) 
found similar changes in human cell strain HeLa; and Chu, Sanford and Earle 
(1958) recorded ring chromosomes in certain mouse lines in addition to other 
anomalies. Moreover, in all the strains studied, none was found to be cytologically 
homogeneous. 

Strain L (Earle, 1943) is probably one of the oldest existing cell lines. Since 
Sanford and her associates (1948) isolated a single L-cell to produce a clone, 
practically all the laboratories working with tissue culture materials have carried 
Clone 929 one time or another. A number of laboratories still employ Clone 929 
as one of their principal materials. It is conceivable that workers in different 
laboratories have used different environments, such as nutrition, temperature, 
mechanical and chemical handling, toxic substances, etc., to cultivate the cells, 
either consciously or unconsciously. These treatments, or sometimes abuses, 
would serve as selection forces to influence the composition of cell populations, 
and genotypes unfit for one environment may gain popularity when conditions 
alter. Thus even if the original Clone 929 was a relatively homogeneous popula- 
tion, the cytological picture of the sublines carried in various laboratories may 
well be different. In fact Hsu and Klatt (1958) already reported a few such cases. 

The present communication presents observational results from a survey of 
twelve L-lines to substantiate the concept of genetic polymorphism. 

MATERIAL AND METHODS 

All the cell strains employed in this study were sublines of Clone 929. Strain L. 
They were kindly supplied by various investigators to whom we are extremely 
grateful. The following list summarizes the origin of these strains: 

Clone 2071, from Dr. Wilton R. Earle, National Cancer Institute, Bethesda, 
Maryland. 

Clone A929, from Dr. Charity Waymouth, Roscoe Jackson Memorial Labora- 
tory, Bar Harbor, Maine. 

Clone 929, from Dr. Donald J. Merchant, University of Michigan, Ann Arbor. 
Michigan. 

Li, Lo, Ls, Lc, Lb, Lob. and Le, all the preceding strains from Dr. Sergey 

1 Supported in part by Grant DRG-269C from the Damon Runyon Memorial Fund for Cancer 
Research and Grant P-133 from the American Cancer Society. 



130 The University of Texas Publication 

Fedorcff, University of Saskatchewan, Saskatoon, Canada. The origin of these 
Hnes will be described by Fedoroff and Cook (in press). Briefly, Strains Lb and 
Lob originated from Strain L^, and Strains Lc and Le were derived from Lb- 

L-P55, from Dr. Charles M. Pomerat, The University of Texas Medical 
Branch, Galveston, Texas. 

L-P55-N2, a subline of LP-55, established in our laboratory. 

The cell lines, upon arrival, were subcultured in Blake bottles with 10% horse 
serum in Eagle's basic medium. Cytological preparations were made as soon as 
practicable so as to minimize possible changes in genotype induced by our system 
of culture handling. The technique of preparing slides was the colchicine-hypo- 
tonic squash. For each strain 50 cells were carefully studied in regard to their 
chromosomal composition. 

RESULTS AND DISCUSSION 

As has been reported, Strains L-P55 and L-P55-N2 possessed one or two dicen- 
tric chromosomes per cell. It was found out later that one of the constrictions was 
in reality a deep secondary constriction, so that the chromosome was a long sub- 
telocentric. Regardless of the nature of the constrictions, the chromosome was 
still an excellent marker and will be henceforth referred to as the D-chromosome 
(fig. 1). This element was also observed in Strains L., Lg, Lqb, Lb, and the stock 
from Michigan University. In the Michigan strain, however, a number of the 
cells contained a special chromosome with a median centric constriction and two 
subterminal secondary constrictions (fig. 2). Probably this chromosome (T-chro- 
mosome) was a derivative of the D-chromosome. The two types of chromosome 
never appeared in one cell, but cells with two D's or two T's have repeatedly 
been observed in this line. The proportion of cells containing D-chromosome 
and those containing T-chromosome was approximately 4:1. 

As has been mentioned previously, Strain Lb and Lqb, both of which contained 
the D-chromosome, were actually substrains of L^. Yet the D-element was not 
observed in L^. It is very unlikely that rearrangement which had produced the 
D-chromosome could be formed repeatedly, so that Strain L^ either had contained 
D-chromosome in its past or still contained such chromosome at a very low 
froquency when the sample was taken. In either case it suggests that both Lb 
and Lob were not clone derivatives. 

Minute chromosomes in Clone 2071 from National Cancer Institute were also 
found in a number of strains. There were at least two types of minutes, one 
slightly larger than the other, and both may appear in a single cell. In the strain 
from the Roscoe Jackson Laboratory many cells contained a medium sized telo- 
centric chromosome that had a secondary constriction lying close to the centro- 
mere. This chromosome was also present in several lines. It is of course not pos- 
sible to establish identity of two morphologically similar marker chromosomes 
present in two different strains, but it is probably not unsafe to conclude that most 
marker chromosomes were not characteristic of any particular strain, but were 
shared by a number of strains. The only obvious exception was the T-chromo- 
some of the Merchant line which has not been found elsewhere. 

Quantitatively, none of the strains analyzed was identical with another. Table 



Hsu: Mammalian Chromosom.es 



131 




■■'.,:. .Wi^l'V- # ^* 



#.% 






- % v"^ 






.# 



f. 






^' 



1^ 









■I 



Fig. 1. A cell from the Michigan University subline showing the D ehroniosonie (arrow). 
Fig. 2. A cell from the Michigan University subline showing the T-chromosome (arrow). 



132 The University of Texas Publication 

I presents data obtained from twelve strains showing mean values of total chro- 
mosome number, of number of metacentrics, subtelocentrics, D-chromosomes, 
and minutes. The total chromosome number was rather similar among all the 
strains ranging from the lowest mean value of 67 (Waymouth) to the highest 
of 73 (2071), a difference of six chromosomes. The number of metacentric chro- 
mosomes per cell, however, varied more significantly. The lowest mean value 
found was 12.5 (L-P55-N2), and the highest, 21.5 (Lb), a difference of nine 
chromosomes. Variation in the number of subtelocentrics was relatively small, 
but that of the D-chromosome and of the minutes ranged from completely absent 
to at least one per cell. 

It should be added that the strains also differed in the frequency of high poly- 
ploids. In most strains high polyploids occupied less than 1 per cent of the popu- 
lation, but in strain L^ the value reached as high as 24.8 per cent. 

In Table 1 an attempt was also made to calculate the percentage of biarmed 
chromosomes per cell. The values ranged from approximately 24 per cent to 32 
per cent. It is understood that these chromosomes were not the only ones that had 
structural alterations. 

Our results substantiate the conclusion that most cell populations in tissue 
culture are genetically polymorphic. Of special interest is the fact that all the 

Table 1 
Mean values and standard deviations of chromosome types in 12 sublines of Strain L, Clone 929 





Source 


Total 


Number 


Number 


Number 


Number 


Percent 


Strain 


of 


Chromosome 


of 


of 


of 


of 


Biarmed 


Designation 


Material 


Nmnber 


Metacentrics 


Subtelocentrics 


D-Chromosomes 


Minutes 


Chromosomes 


Lc 


Fedoroff 


71.66± 


13.68+ 


3.58H= 


0.00 


1.10+ 


24.21 






1.85 


1.18 


0.61 




0.63 




Lg 


Fedoroff 


72.74± 


15.00+ 


3.24d= 


0.02+ 


0.44+ 


25.13 






1.82 


1.20 


0.82 


0.14 


0.54 




Lqb 


Fedoroff 


68.14± 


14.18± 


3.28+ 


0.14d= 


0.86+ 


26.00 




3.00 


1.66 


0.78 


0.35 


0.70 




Li 


Fedoroff 


67.96± 


14.68+ 


1.42+ 


0.00 


0.38+ 


23.71 


1 




1.93 


1.30 


0.73 




0.49 




l; 


Fedoroff 


67.30± 


13.50+ 


2.84+ 


0.80+ 


0.00 


27.86 






2.38 


1.61 


0.91 


0.45 






Lb 


Fedoroff 


67.48± 


21.52+ 


2.06+ 


0.06+ 


0.60+ 


35.11 






2.91 


2.71 


0.55 


0.24 


0.76 




Le 


Fedoroff 


71.80± 


14.56+ 


2.68+ 


0.00 


0.68+ 


23.01 






1.09 


1.37 


0.75 




0.54 




L(M) 


Merchant 


67.36± 


13.44+ 


2.24+ 


0.44+ 


1.06+ 


23.97 






3.28 


1.86 


0.74 


0.50* 


0.82 




L(W) 


Waymouth 


67.30± 


18.18+ 


3.76+ 


0.00 


0.22+ 


32.68 






2.89 


1.45 


1.06 




0.46 




2071 


Earle 


73.06± 


17.68+ 


3.74+ 


0.00 


0.38+ 


29.34 






1.70 


1.39 


0.75 




0.53 




L-P55 


Pomerat 


67.88± 


12.94+ 


3.78+ 


1.20+ 


0.00 


25.42 






1.81 


1.32 


0.86 


0.61 






L-P55-N2 


Hsu 


67.86± 


12.50+ 


2.06+ 


1.60+ 


0.00 


23.83 






1.87 


1.02 


0.37 


0.49 







* T-chromosomes included in this category. 



Hsu: Mammalian Chrom.osom.es 133 

lines reported here were derivatives of a single cell. Since some marker chromo- 
somes were found in different laboratories which had had no direct contact with 
one another prior to the time when the cultures were shipped to this laboratory, 
indication is therefore strong that these chromosomes were present in the original 
population of Clone 929 in the National Cancer Institute. Perhaps each was pres- 
ent at a relatively low frequency but various new environments facilitated the 
selection of special genomes. The populations of somatic cells in vitro are thus 
analogous to the populations of Drosophila in the well-known population cages. 

It should also be borne in mind that each line was represented by only 50 cells. 
Variation in genotype should indeed be much greater in each population than 
indicated in Table I. Some genotypes may be at a very low frequency and can be 
brought to distinction only when the existing superior genomes are not fit for 
survival. Indeed, experimentally produced changes in genomes have been ob- 
tained in Strain L-P55 when the populations were subjected to media containing 
various sugars to replace glucose (Hsu and Kellogg, in press) . In L-P55, the low- 
est chromosome number per cell registered was 63, and such cells occupied less 
than 5 per cent of the population. In the special "sugar" lines a large proportion 
of cells showed chromosome numbers in the vicinity of 63, with the lowest num- 
ber registered as 59. 

Recently Puck (1958) and Tjio and Puck (1958) claimed that some samples 
of serum used in preparing growth medium for in vitro cells contain toxic sub- 
stances which might induce mitotic anomalies and cause cytological hetero- 
geneity. Application of their procedure may enable investigators to maintain 
relatively homogeneous populations as experimental materials. 

SUMMARY 

Twelve sublines of Strain L, Clone 929 were analyzed in regard to their total 
chromosome number per cell, numbers of metacentric and subtelocentric chro- 
mosomes, D-chromosomes, and minute chromosomes. Statistically none of the 
lines was identical with another, yet marker chromosomes were found in differ- 
ent lines which had never had contact with one another. This indicates that the 
marker chromosomes must have a long history and they might all have been 
present in the original 929 clone in the National Cancer Institute before stocks 
were distributed. Different genotypes developed in different laboratories when 
conditions favored their survival. 

EPILOGUE 

It has been over seven years since I left Professor Patterson's laboratory and 
took a seemingly completely different field of biological research from my pre- 
vious interest in Drosophila genetics. The present article constitutes the eleventh 
paper on the topic of mammalian chromosomes in vitro^ but it is interesting to 
note from this study that a bottle of mammalian cells may also be excellent ma- 
terial for studying population and evolutionary genetics. One can never escape 
from the influence of such a stimulating teacher. 



1 34 The University of Texas Publication 

REFERENCES 

Chu, E. H. Y. and N. H. Giles. 1957. Comparative chromosomal studies on mammalian cells in 
culture. I. The HeLa strain and its mutant clonal derivatives. J. Nat. Cancer Inst., 20:383- 
401. 

Chu, E. H. Y., K. K. Sanford and W. R. Earle. 1958. Comparative chromosomal studies on mam- 
malian cells in culture. II. J. Nat. Cancer Inst., 21:729-751. 

Earle, W. R. 1943. Production of malignancy in vitro. IV. The mouse fibroblast cultures and 
changes seen in the living cells. J. Nat. Cancer Inst., 4: 165-212. 

Fedoroff, S. and B. Cook (in press). Effect of human serum on tissue cultures. II. Development of 
resistance to toxic human serum in strain L cells. 

FIsu, T. C. and D. S. Kellogg, Jr. (in press). Genetics of in vitro cells. Genetics and Cancer Mono- 
graph for 13th Annual Symposium on Fundamental Cancer Research, M. D. Anderson 
Hospital and Tumor Institute. 

FIsu, T. C. and O. Klatt. 1958. Mammalian chromosomes in vitro. IX. On genetic polymorphism 
in cell populations. J. Nat. Cancer Inst., 21:437-473. 

Puck, T. T. Genetic studies on somatic mammalian cells. Proc. X. Intern. Cong. Genet., 2:225. 

Sanford, K. K., W. R. Earle, and G. D. Likely. 1948. The growth in vitro of single isolated tissue 
cells. J. Nat. Cancer Inst., 9:229-246. 

Tjio, J. H. and T. T. Puck. 1958. Genetics of somatic mammalian cells. II. Chromosomal consti- 
tution of cells in tissue culture. J. Exp. Med. 108:259-268. 



( 



Heterochromatic Control of Position-Effect 
Variegation in Drosophila' 

WILLIAM K. BAKER AND JANICE B. SPOFFORD 
Department of Zoology, The University of Chicago, Chicago, lUinois 

It is particularly fitting that a discussion of somatic variegation be included 
in the volume honoring Professor Patterson's eightieth birthday. He was not 
only among the first of the Drosophila geneticists (Patterson 1928) to become in- 
terested in the genetics of somatic tissue but he picked this topic to initiate his 
brilliant series of genetic investigations. His vision is attested to by the fact that 
now, thirty years later, this field is at the forefront of genetic research and the 
subject of a recent symposium (Hollaender 1958) . 

The subject of genetically controlled variegation is intriguing to a student of 
developmental biology because of its relevance to the problem of differentiation 
of cellular function. The variegation caused by position effect is of signal interest 
in this connection because its expression can be altered by a host of inherited 
factors as well as by certain environmental agents. The work being reported is 
only an exploratory effort designed to uncover some of the complicating factors 
involved in V-type position effects. Novel types of genetic control of variegation 
were discovered during this investigation which may play an important role in 
cellular differentiation. They also prove to be a source of embarrassing variability 
if their presence is not recognized; a criticism which is applicable to certain of 
the experiments being reported herein. 

These experiments were initiated with the thought in mind that since extra 
heterochromatin suppresses white variegation in Drosophila melanogaster^ an 
opportunity was provided to determine if there were separable units of suppres- 
sing activity within the heterochromatin of the Y chromosome. In other words, 
could heterochromatin be subdivided into functional units like euchromatic 
chromosome regions? Different fragments of Y chromosomes, inserted into the 
genome of three different types of flies, were used to test this notion. The end 
effect, their influence on the expression of variegation, was measured both vis- 
ually and chromatographically. The latter procedure provided some evidence on 
the chemical differences between pigmentation in variegated, white, and wild- 
type eyes. 

MATERIAL AND METHODS 

The rearrangement used to produce the white-variegated eyes was derived 
from N^*^*"°® (Demerec 1940) which is a transportation of the white region, in 
reversed order, into the heterochromatin of 3L. It was carried as a duplication, 
Dp(w'"), the white region of the X chromosome being structurally normal but 
carrying the w allele. At the beginning of the experiments (1956) it appeared 
that the duplication was lethal when homozygous in either males or females. 

1 Work performed under contract No. AT (11-1) -431 for the Atomic Energy Commission. 



136 The University of Texas Publication 

However, in the spring of 1957 (after co-isogenic stocks had been made) viable 
and fertile females which were homozygous for Dp(w"') were discovered in the 
stock cultures. It has been conclusively established that homozygous males, if 
viable, are sterile; probably they are not viable. 

Most of the Y fragments utilized originated from exchange between an inverted 
X chromosome and a normal Y, or as products of exchange between X Y^ and a 
free Y chromosome, or from spontaneous or induced breakage of the sc*-Y:bw+ 
chromosome arising in experiments previously reported (Baker, 1955, 1957). A 
brief description of each of the Y fragments follows: 

Y<^^:bb+. Arose from X-irradiation (1500r) of sperm bearing the sc^Y:bw+ 
chromosome. It is a moderate size ring-Y similar in size to the three other Y^^'s 
to be described. (See Figure 7.) 

Y^:y+ bb+-7. Induced by X-irradiation of sc^Y:bw+ sperm (1750r). Cyto- 
logically it appears as a small single-armed chromosome about twice the size of 
chromosome 4. 

Y^. A small two-armed chromosome. 

Y'^^-H. A ring chromosome that arose from X-irradiation (451r) of sc^Y:bw+ 
sperm. 

Y^:bb+-8. Spontaneously arose in X''Vsc^Y:bw+ males. It is a small two- 
armed chromosome. 

Y^:y+ bb+-6. A two-armed chromosome produced by X-irradiating sc^Y:bw+ 
sperm with 175 Or. 

Y^^:bw+ bb + . A ring chromosome arising from sc^ Y:bw+ sperm treated with 
45 Ir of X rays. 

YcL_J5 ^ j.jj^g chromosome spontaneously arising in X'^'/sc^ Y:bw+ males. 

Ys.ys #2. A two-armed chromosome of moderate size formed by exchange be- 
tween an XY^ chromsome and the sc^Y. (see Muller 1948) . 

g(.vi.Ys A small two-armed chromosome formed by exchange between a nor- 
mal Y and the sc^^ inverted-X chromosome. (Muller 1948) . 

ycL p^ ring chromosome which arose in the crosses giving rise to the Y^-Y^ #2 
chromosome. (Muller 1948). 

Y^— 13. A large acrocentric spontaneously arising in X^Vsc^Y:bw+ males. 

Y^:y+ bb + — 5. This large acrocentric arose spontaneously in X''Vsc^Y:bw+ 
males. 

Y. The Y chromosome from the M-5; Cy; Ubx^^^VXa stock, (see Lewis, 1952, 
for a description of Ubx^'^'^.) 

sc^^-Y^ #2. This large acrocentric chromosome arose by exchange between a 
free Y and the distal end of the Muller-5 chromosome (Parker and McCrone 
1958). 

The study was designed to determine the effect on variegation of these 15 dif- 
ferent Y chromosomes in three different genotypes resulting from the matings 
listed below: 

Y^wyY^y + /Y; Dp(w'")/+ $ $ X yw/Y^ ? 9 which produces 

(1) YVy.Y^y+ZY^; Dp (w"0/+ males. 
XY"VY^ 5 $ X y w/y w; Dp(w'")/+ 9 2 which produces 

(2) y w/Y^; Dp(w"0/+ males. 



Baker and Spofford: Heterochromatin and Variegation 



137 



X-Y'VY^ $ S X yw/Y;Dp(w"0/+ ? ? which produces 
(3) yw/Y^;"Dp(w"0/+ females. 
(The general notation XY'^ designates an X chromosome to which is attached a 

Y that is lacking the fertility factors present in the free fragment; e.g. X Y^/Y^' 
orX-YVY«.) 

It is well known that there are numerous genetic modifiers of variegation; 
therefore, if a critical study was to be made of the effects of the Y fragments per 
se, co-isogenicity of the stocks used was a necessary prerequisite. This involved 
the synthesis of 20 different stocks that were as nearly co-isogenic as possible 
within the limits of the experimental technique employed. It is not feasible to 
present the detailed breeding scheme which gave rise to these co-isogenic stocks; 
however, a schematic diagram outlining the protocol used is shown in figure 1 . 
Each block in this diagram represents a stock and within the block is given the 
genotype of each chromosome in the order (from the left) ; the X and Y, the 2nd, 
and the 3rd. The subscripts to the chromosomes (a, b, c, etc.) indicate homolo- 
gous chromosomes that need not be identical in their genetic composition. Certain 
limitations of this scheme should be noted. The Y chromosome in all cases comes 
from the Muller-5; Cy; Ubx^^VXa stock but no effort was made to insure that the 

Y in all strains came from a single Y of the original stock. No control over the 
heterogeneity of the 4th chromosome was attempted and, of course, the Cy and 
Ubx^''^ inversions do not completely ehminate the recovery of crossover products 
providing an additional unmeasured source of variability. Finally, there is free 
exchange between the third chromosome bearing the duplication and its homolog. 
This may be an influential factor in causing variability since the remainder of 



^_w^ yw 



•^0 



y Wq -t-o +a 

y Wb' +b' +b 



l^. ±e. ±d 



^ 



y Wq y wq -|^ 
Y ® y Wq' +a 



y^^- y_Wa .+a. Dp(w^) 
Y ** y Wq'+q' +a 



XY-F yjl. ±0. ±0 
yF * yF' +0' +a 



-d ' +c 



M-5 . Cy; Ubx' 
Y ' Xa 



+g. +f 



y£w_j^* XX.+i . ±± 
Y " Y • +j ' -l-i 



15 DIFFERENT Y'"'s USED 



Y^w yYl-yt^ Ui. -4-a. Dp(wm) 



Y +a +a 



Fig. 1 . The breeding scheme for synthesis of the coisogenic stocks. 



138 The University of Texas Publication 

the chromosome 3 bearing the duplication was not made co-isogenic with the 
normal 3 from which the other stocks were derived. 

The matings in the isogenization scheme were so scheduled that stock #3, stock 
*5, and most of the 15 different stock #4's were synthesized at about the same 
time. (These are the stocks necessary to produce the three different genotypes of 
flies in which variegation was examined.) This enabled the study to be made 
shortly after isogenization was completed, thus reducing the effect of newly arisen 
modifiers. After the stocks were synthesized, no controlled breeding pattern was 
utilized (e.g., brother-sister matings), and the heterozygosity of the duplication 
was not insured by outcrossing. At that time we did not suspect that flies homo- 
zygous for the duplication would rather suddenly become viable nor did we sus- 
pect that the homo- vs. heterozygosity of a mother would influence the occurrence 
and extent of variegation in her heterozygous offspring (Spofford, 1958, 1959). 

There is one other potential source of genetic variability that was desirable to 
control. If nondisjunction of sex chromosomes takes place during the breeding 
scheme, then an individual fly might contain a Y chromosome in addition to the 
expected Y-fragment. Or, similarly, two Y-fragments might occur in a fly where 
only one is anticipated. Since the effect on variegation of different Y fragments 
was the main purpose of the study, tests had to be made to determine the presence 
of extra Y's or extra Y^'s. This entailed making a progeny test of each of the 
individuals scored for variegation. The labor involved when the Y'^ did not have 
a visible marker was enormous and the results, as to be expected, were only par- 
tially successful. When the flies to be scored for variegation had eclosed, they 
were mated, in general, according to the following schemes: 

( 1 ) Y^w yYV+/Y^; Dp (w"^) /+ males X X-Y'^/w females. 
If the white male offspring from this cross were sterile, it could be assumed that 
no free complete Y was present in their father. The fertility of the other class of 
F^ male offspring assured the presence of at least one Y^ in the father. The white 
female offspring were crossed to wild-type males and their offspring examined. If 
no wild-type males appeared, then their mothers, and therefore the original male 
parent, did not contain an extra Y^. For if an extra Y^ had been present in the 
male, some of his white daughters would receive this chromosome and by sec- 
ondary nondisjunction would form a few wild-type sons when crossed to wild- 
type males. 

(2) yw/Y^; Dp(w™)/+ males were crossed to any type of virgin female to 
see if they were fertile. Since most of the Y^'s contained either the Y^ or the Y^ 
fertility factors but not both, this cross tested for the presence of an extra complete 
Y. Of course, no test can be made for the possibilities that more than one Y^ or no 
Y'^ existed in these sterile males. 

(3) yw/Y^; Dp (w"^) /+ females X y BY'VY^ males. Fertility of the Fi males 
indicates the presence of at least one Y'^ in the mother. These Fi males were 
crossed to y f females and the resulting sons were checked for fertility, indicating 
the presence or absence of a complete Y in either of the original parents. The 
resulting daughters, which should be y BY'^/y f were mated to wild-type males 
and any non-yellow males in their progeny would indicate the probable presence 
of an extra Y^ in either original parent. 



Baker and Spofford: Heterochromatin and Variegation 1 39 

It can be seen from these mating schemes that the presence of an extra Y chro- 
mosome can be detected with a good deal of assurance; however, detection of an 
extra fragment depends on securing a sufficient number of offspring to insure the 
appearance of secondary exceptions. Since within almost every one of the 48 
genotypes examined in this study there was sufficient homogeneity m the appear- 
ance of the variegated eyes, it was possible to detect aberrant phenotypes pro- 
duced by extra Y's or extra Y^'s. Progeny tests were successfully completed on 
many of the individuals exhibiting the "normal" phenotype for each genotype 
and on at least some of the phenotypically aberrant flies. The latter tests revealed 
the expression of variegation resulting from an extra Y chromosome. Thus it 
seemed justified to exclude from the data other similarly aberrant individuals 
on which progeny tests were not completed or were ambiguous, and likewise to 
include flies with "normal" expression of variegation in which the progeny tests 
were unsuccessful. 

The males undergoing the progeny tests remained with their mates for three 
days before they were removed and scored for variegation; females remained five 
days. Therefore, all measurements of variegation were made on males that were 
from three to four days of age and females five to six days. Two types of measure- 
ments were made: a visual examination of the extent of pigmentation in each 
eye, and a chromatographic analysis of the pteridines in each individual head. 
All the visual estimates were made by one of us (J.B.S.). These estimates took 
into account primarily the size of the area that was pigmented but also, in certam 
cases, the intensity of pigmentation. If the visual estimates were to be a function 
of the amount of pigmentation in the eye, the latter consideration was necessary 
in particular variegated eyes in which only a "tinge" of pigmentation was present. 
Each eye was graded separately; a value of was assigned to a completely color- 
less eye and a value of 1 to a totally pigmented eye, with a variegated eye falling 
in between these values. 

After the visual estimates were made, the fly was decapitated and its head im- 
mediately squashed on Whatman No. 1 fiUer paper for chromatographic analysis. 
On each 15 x 15 cm. sheet of paper, 10 heads with variegated eyes and one head 
from an Oregon-R male (3-4 days old) were squashed at 1 cm. intervals, 1.5 cm. 
from the bottom of the paper, following the procedure of Hadorn and Mitchell 
(1951). The chromatograms were equilibrated by placing in the bottom of the 
chromatographic jar the aqueous phase of the butanol-acetic solvent (4 butanol : 
5 H.O : 1 glacial acetic acid) and allowing two hours for equilibration at which 
time the organic phase was layered on top of the aqueous phase until the bottom 
of the chromatographic paper was immersed in the developing solvent. The front 
was allowed to ascend for 21/2 hours at 23° C, at which time it had reached 12.5 
cm. from the line on which the heads were located. The chromatograms were 
then removed from the jar and allowed to dry at 23° C. 

The ultra-violet fluorescence of the substances chromatographed was ex- 
amined with a "Shannon" lamp and a Corning No. 9863 filter. Two rectangular 
areas for each head were cut out of the paper for elution of the fluorescent sub- 
stances they contained. One area 3 x 0.8 cm., included the head and extended 
vertically to an Rf value of about 0.22. This area contains all of the visible 



140 The University of Texas Publication 

pigments and fluoresces orange and yellow (Fl-1 and Fl-2, see Hadorn and 
Mitchell, 1951 ) . For the purposes of this paper, we shall call this the area contain- 
ing the "yellow" fluorescing substances. The other area was 1.5 x 0.8 cm. with its 
lower boundary at about Rf = 0.30 and its upper boundary at Rf = 0.42. This 
area includes the substances F1-4A, 4B and 5 and is called by us the area of "blue" 
fluorescing substances. 

The rectangles cut out of the chromatograms were rolled in a loose spiral, each 
placed in the bottom of a 10 x 75 mm. test tube, and extracted in 1 cc of a 2% 
glacial acetic acid solution for 2 hours at 23° C. After this period the paper was 
removed and the fluorescence of the solution measured by means of a Farrand 
Photofluorometer. A Corning No. 9863 filter was used between the UV-lamp 
and the cuvette containing the solution being measured. Between the cuvette and 
the photo tube (in that order) filters Nos. 4308 and 3385 were inserted for the 
"blue" substances, and Nos. 3385 and 3389 for the "yellow" substances. The 
standard used was 1 ^gm/ml of anthranilic acid dissolved in 2% acetic acid. Prior 
to each reading the fluorometer was adjusted to give a galvanometer reading of 
5 for 1 ml. of the standard. Thus the units of fluorescence given in this paper are 
in units of anthranilic acid fluorescence. The experimental readings have each 
been corrected for fluorescence of the filter paper by subtracting the fluorescent 
reading of a paper blank whose size and Rf location were the same as for the ex- 
perimental area being measured. This correction is valid since the photoflu- 
orometer has a linear response to concentration of fluorescing substances over the 
range employed. 

RESULTS 

Chromatographic Analysis of Variegated Eyes 

An examination in ultraviolet light of the chromatograms of single heads con- 
taining white, variegated and wild-type eyes reveals, in general, the following 
features. White eyes show no fluorescence. In variegated eyes, the amount of 
fluorescence associated with the visible pigments in the "yellow" area is directly 
related to the amount of pigmentation in the eye; whereas in the "blue" region 
from variegated eyes there is more fluorescence than in wild-type if the eyes are 
about half pigmented. Thus, one of the striking features of variegated eyes is 
their accumulation of certain of the pteridines far in excess of that found in wild- 
type. 

Figure 2 shows the relations between the amount of the "yellow" and the 
"blue" fluorescent substances, measured photofluorometrically, and our visual 
estimate of the amount of pigmentation in 55 individual heads of one genotype. 
This example is atypical only in the fact that this one genotype of males shows a 
wide variation of expression of variegation in individual flies, all the way from 
practically white eyes to those showing almost complete pigmentation. The linear 
relation found between the amount of the "yellow" substances and our visual 
estimates of the amount of pigmentation supports the claim that these estimates 
are reasonably accurate assessments of the extent of pigmentation since the "yel- 
low" fluorescence is associated with the visible pigments on the chromatograms. 
The excessive amount, over wild-type, of the "blue" fluorescing substances is 



Baker and Spofford: Meter ochromatin and Variegation 



141 



2.0 



1.6 



1.2 



< 

0^0.8 
111 



< 
1 0.4 



0.5 



1.0 



Y^w yYS'VY^-13 C? / 

/ oo 
o / o 

yellow O C^'O O 
O / 





««.••»** 



3 
BLUE 



Fig. 2. Relation between the amount of "yellow" and "blue" fluorescence and the extent of 
pigmentation in Y^w yYLy+/YL-13; Dp(w"^)/+ males. 

clearly evident at intermediate values of variegation. Therefore, it is reasonable 
to conclude that the pigmented areas of variegated eyes are chemically different 
(at least quantitatively) from wild-type. 

There are three chemical compounds present in the "blue" fluorescing area: 
F1-4A which is 2-amino-4-hydroxy-6-(r, 2'-dihydroxypropyl)-pteridine (bio- 
pterin) ; F1-4B which is 2 amino-4-hydoxypteridine; and the sepia pteridine (Fl- 
5) which is N«-lactyl-7, 8-dihydro-2-amino-4-hydroxypteridine-6-carboxyl acid 
(see Forrest and Mitchell 1955) . The presence of these three compounds in wild- 
type and variegated eyes was detected by comparing th Rf's of spots from chrom- 
atograms of heads with those made using the pure F1-4A and F1-4B, kindly sup- 
plied by Dr. Hugh S. Forrest. (The Rf's for the sepia pteridine were determined 
from chromatograms of sepia flies.) Four types of chromatograms were used to 
confirm the identification: one-way ascending in butanol : acetic : HoO (4:1:5) ; 
one-way descending in propanol : 7% NH.OH (2:1); and two-way, ascending 
and descending, using these two solvents. For the two-way chromatograms, 20 
heads of each genotype were homogenized in 7% NH.OH, shaken with chloro- 
form, and part of the water-soluble portion spotted on the paper. Since no quanti- 



142 The University of Texas Publication 

tative comparison of the amount of fluorescence between wild and variegated 
eyes was made for each of these pteridines, we are not certain as to the extent 
which each of these substances contributes to the greater fluorescence in the 
"blue" area in variegated flies. However, it appears that the major portion of the 
difference is caused by a greater amount of the sepia pteridine in variegated 
eyes. This would explain the "brownish" appearance of the pigmented areas in 
many eyes. 

Effects of Y -Fragments on Variegation 

The main body of data gathered in this study concerns the effect of different 
fragments of Y chromosomes on the expression of variegation. Four criteria of 
effect were utilized: a visual estimate of the extent of pigmentation, the amount 
of "yellow" and "blue" fluorescence, and the penetrance of variegation. The latter 
criterion deserves further comment. In many of the genotypes, especially with 

Y fragments that enhance the extent of pigmentation only slightly or not at all, 
many of the flies will have completely white eyes even though they carry the 
duplication containing w"\ These cases have been spoken of as those in which the 
variegation has gone to completion. In the crosses which gave rise to the indi- 
viduals studied, the duplication was carried in heterozygous condition and in 
only one of the parents, with the result that half of the offspring would be ex- 
pected to carry it. If half of the offspring were variegated and half white, then 
the penetrance is said to be 100%; however, if more offspring were white than 
variegated the penetrance figure is less than 100%. This, of course, assumes that 
the third chromosome bearing the duplication is just as likely to be included in 
the egg nucleus as the normal chromosome 3 and that there is no net influence on 
viability; reasonable assumptions, but ones for which we have no evidence. In 
the tables that follow, some of the penetrance figures are given as a range between 
two percentages. This is done in those cases in which supposedly replicate samples 
are statistically different from one another in their degree of penetrance. 

Let us examine (Table 1 ) the results of these measurements in Y^w yY^y+/Y^; 
Dp(w"')/+ males. The Y^'s are arranged in increasing order of effectiveness in 
enhancing pigmentation (based on the visual estimate). Practically all of the 
attached-XY males without a fragment have white eyes. Only two out of the 
estimated 275 males of this genotype showed any pigment and these pigmented 
areas covered only a few ommatidia. In contrast, the males containing a free 
fragment all showed a high degree of penetrance even though, with many frag- 
ments, the median extent of pigmentation was less than a fifth of the area in each 
eye. One of the most impressive features of these data is the marked differences in 
the effectiveness of many of the Y fragments in suppressing variegation; some 
have very little effect, and others, e.g. sc^'Y^, are just as effective as a complete 

Y chromosome. It is also of interest to note that the amount of fluorescence in the 
"blue" region is highest in males with fragment Y^-13. These males produce an 
intermediate value of pigmentation as judged by both the visual estimate and 
the amount of "yellow" fluorescence. This again attests to the accumulation of 
certain of the pteridines in variegated eyes. 

In table 2 are presented the data obtained from measurements on yw/Y^; 
Dp(w"')/-H females. It is obvious that, with the exception of the last three frag- 



Baker and Spofford: Heterochromatin and Variegation 



143 



merits listed, the eyes of these females have very little pigmentation. In addition, 
even in those cases where the eyes are heavily pigmented the penetrance is far 
from complete. Especially noteworthy is the observation that many of the frag- 
ments are more effective than the complete Y chromosome in suppressing 

Table 1 
Measurements on Y^w yYLy+/YF; Dp(w>»)/+ males 



Y Fragment 



None 

YcL:bb+ 

YS:y+bb+-7 

ys 

YcL_14 

YS:bb+-8 

YS:y+bb+-6 

YcS:bw+ bb+ 

YcL_i5 

ys-Y^ 

scvi.ys 
ycL 
YL-13 
YS:y+bb+-5 

Y 

scSi.yL 



Flies Showing 
Pigmentation 



Measurements on Individug 



N 


Per Cent 


275 


1 


199 


40-68 


178 


81 


63 


82 


419 


74-92 


123 


40-97 


171 


100 


109 


100 


484 


80-100 


787 


78-100 


302 


90-100 


216 


74-100 


435 


94 


395 


100 


68 


95 


218 


100 



Total 
Number 



Median 
Visual* 



2 


<0.01 


50 


0.05 


19 


0.05 


34 


0.09 


46 


0.10 


40 


0.10 


53 


0.18 


45 


0.20 


73 


0.24 


60 


0.27 


38 


0.30 


95 


0.33 


57 


0.75 


50 


1.10 


52 


1.20 


47 


1.30 



Median 
Blue Fl.f 



0.55 
0.65 
1.18 
1.38 
0.95 
0.95 
1.40 
1.60 
1.70 
1.75 
1.78 
3.45 
1.73 
2.35 
2.50 



Median 
Yellow Fl. 



0.05 
0.10 
0.28 
0.20 
0.30 
0.40 
0.38 
0.40 
0.30 
0.33 
0.43 
0.75 
0.85 
1.00 
0.95 



White eves=:0, wild-type eyes = 2.0. 

l»gm/ml of anthranilic acid in elution solvent gives a readmg ot 5.U. 



Table 2 

Measurements on yw/Y^; Dp(wni)/+ females 







Flies Showing 
Pigmentation 




Measurements on Individuals 






Total 
Number 


Median 
Visual 


Median 
Blue Fl. 


Median 


Y Fragment 


N 


Per Cent 


Yellow Fl. 


None 




724 













ys.bb+-8 




618 


4 


21 


0.01 


0.10 


0.07 


YS:y+bb+- 


-6 


621 


5-20 


23 


0.02 


0.26 


0.10 


ys.ys 




800 


6 


44 


0.03 


0.35 


0.15 


Y 




242 


7-19 


27 


0.03 


0.35 


0.20 


ycs.bw+bb+ 


488 


21 


50 


0.03 


0.35 


0.25 


YcL-15 




738 


0-25 


38 


0.04 


0.40 


0.10 


YS:y+bb+- 


-5 


436 


19 


57 


0.06 


0.60 


0.20 


ycL_i4 




488 


11-33 


28 


0.05 


0.97 


0.20 


YcL:bb+ 




433 


3 


12 


0.07 


0.15 


0.04 


YS:y+bb+- 


-7 


716 


10 


55 


0.08 


0.30 


0.20 


scvi.ys 




338 


4 


30 


0.16 


0.52 


0.07 


ys 




172 


14-62 


33 


0.20 


1.55 


0.35 


ycL 




84 


62-84 


43 


1.30 


2.90 


1.30 


YL-13 




752 


51-100 


37 


1.39 


5.45 


1.60 


scSi.yL 




620 


49-88 


36 


1.93 


3.30 


2.20 



144 



The University of Texas Publication 



variegation in this genotype. Once again it is found that females carrying Y^- 13, 
which have eyes that are a little over half pigmented, accumulate the "blue" 
pteridines to a high degree. 

The measurements on yw/Y^; Dp(w™)/+ males are presented in table 3. The 
most obvious feature of these males is the small amount of pigmentation in their 
eyes irrespective of the Y fragments they carry. However, in spite of the small 
size of the pigmented areas, the penetrance in many cases is moderately high. 
For example, even though the median extent of pigmentation in males carrying 
the Y^:y+ bb+-5 was only a two-hundredth of the area in each eye, some of the 
replicates showed 100% penetrance. It is interesting to observe that in these 
males the complete Y is definitely the most effective enhancer of pigmentation. 

The data in the tables previously discussed illustrate quite dramatically that 
the relative effectiveness of a fragment may be quite different in the three dif- 
ferent genotypes of flies in which studies were made. In order to make not only 
this point clear but also to indicate similarities in patterns of effectiveness of 
certain of the fragments in the three genotypes, all the data have been sum- 
marized in figure 3. The size of the black area in each of the circles in this figure 
indicates the visually estimated median value for the area of pigmentation in 
both eyes combined. A glance at this figure will suffice to show that a fragment 
which is effective in enhancing pigmentation in one genotype may be quite inef- 
fective in one of the other genotypes, or vice-versa. This is indicative of a highly 
complex set of interactions which determine effectiveness of heterochromatin in 
suppressing variegation. However, certain fragments behave according to par- 
ticular patterns. For example, Y^^, sc^'Y^ and YL-13 are most effective in at- 
tached-X females, least effective in normal X males, and have intermediate 
effectiveness in attached-XY males in as far as the visual estimate of pigmenta- 



Table 3 
Measurements on y w/Y^'; Dp(wm)/+ males 





Flies Showing 
Pigmentation 




Measurements 


on Individuals 






Total 
Number 


Median 
Visual 


Median 
Blue Fl. 




Y Fragment 


N 


Per Cent 


Median 
Yellow Fl. 


None 


600 













YS:y+bb+-5 


467 


21-100 


36 


0.01 


0.10 


0.10 


Y<=L:bb+ 


494. 


5 


59 


0.04 


0.25 


0.04 


YcL 


862 


56-95 


75 


0.06 


0.30 


0.05 


YcL_i5 


908 


3 


74 


0.06 


0.48 


0.10 


YcL_i4 


880 


26-76 


45 


0.06 


0.60 


0.15 


YS:y+bb+-6 


659 


15-67 


65 


0.07 


0.50 


0.20 


YS:y+bb+-7 


238 


69-94 


39 


0.07 


0.80 


0.15 


YcS:bw+bb+ 


285 


32 


89 


0.08 


0.81 


0.17 


YL-13 


638 


34^75 


43 


0.10 


1.10 


0.15 


ys 


728 


14-56 


39 


0.10 


1.15 


0.25 


YS:bb+-8 


1000 


54-94 


100 


0.11 


0.89 


0.26 


scSi.yL 


542 


52 


27 


0.14 


0.30 


0.20 


scvi.ys 


294 


40-100 


37 


0.17 


2.15 


0.30 


ysys 


851 


19 


45 


0.45 


2.20 


0.30 


Y 


610 


4-62 


55 


0.65 


2.70 


0.45 



Baker and Spofford: Heterochromatin and Variegation 145 



Pigmented 
Y-Frogment Flies Visual Fluorescence 



None 1% V J 0°/» V J 0°/' 

ycLbb* 40-68»/ 



YSy*bb^-7 8l%(v_yl=) I0 7o( 'o 69-94 7o(^c=: 

ySbb* 82%<^^i=i blue 14-62% C^ I 1 l4-567o(^i=: 

YcL.|4 74-92 '/oC^Cz:^ 11-33 7o(_)c^ 26-76 %V^a 

yS b b"*"- 8 40- 97 %*^^ C=3 4 »/o V^ ' 54-94 % V_y cz 

YS.y+bb"*"-6 IOO%(_Jc=l 5-20 7o<^^D .15-67 % (^a 

Y*^^:bb* bw"*" 100 %V_)c=i 2l7oV^D 32 7o V^i= 

Y*^"— 15 80-100 %V,^ I 1 0-25 7oV^_ya 3 7o V^_ya 

yS yS 78i007oV ^1 1 6 7oV_^)a l9 7o^^l= 

r>)" C^' C^r 

7o'^l==3 4 7oV^ /c=3 40-I007o^_yc: 

Vol^^ ( I 62-84 7ol^Pi I 56- 95 7o V_/ ° 

4 7^^^ , «ii-inn%(^p . 34-75 7o(,^i:z 



sc^'y^ 90-100 



74-100 

yL-IS 9' 

YS:y*bb*"-5 100 7. (^c=l l9 7o*^^C3 21-100 7oV^ I 

Y 95 vS^ r I 7-|9 7o(^^D 4-62 7o'^l I 

scSIyL 100 %^^^ft I 49-88 7.^^1 -i 52 7<,V^^'d 

Y^w yY'-yVY'", Dp(wm)/+ (5* ^_wA'^ , Dplw'")/^ ^ y w/y'^, Dp(w'")/-t- cT 

Fig. 3. A summary of the data on how 15 different Y fragments affect the penetrance, the 
pigmentation, and the amount of certain pteridines in variegated eyes of flies of three genotypes. 

tion and the amount of "yellow" and "blue" fluorescing substances are con- 
cerned; whereas other fragments, like Y^Y^ and Y^:bb+-8 are less effective in 
attached-X females than in the other two genotypes. These factors are brought 
out to indicate that not only can many of the Y fragments be differentiated from 
each other by their effect within a given genotype but also they can be distin- 
guished by their pattern of response among the three genotypes. In particular, 
note the differences in the four Y''^ fragments. 

Before proceeding further with an analysis and discussion of these resuks, it 
is instructive to learn the extent of variation encountered in the three measure- 
ments among individuals of supposedly the same genotype. The data in the 
previously discussed tables and figure were given in terms of medians. This was 
felt necessary in view of the fact that in certain of the genotypes and in particu- 
lar with the "blue" fluorescence measurements there were long tails on the 
frequency distributions. Histograms showing the distributions for the visual 
estimates, the "yellow" fluorescence substances, and the "blue" fluorescence are 
pictured in figures 4, 5 and 6 respectively. Let us consider first the histograms 



146 



The University of Texas Publication 



Dp(w'^/-(- U 




Fig. 4. Frequency histograms of the visual estimates of pigmentation. The figure above each 
histogram indicates the total number of individuals sampled. 



of the visual estimates. After the precautions which had been taken to attain co- 
isogenicity of the stocks used and the constancy of the environmental conditions 
under which the flies were reared, it is certainly disturbing to find such wide 
variation in the extent of variegation as observed, for example, in attached-XY 
males bearing the Y or the Y^-13 chromosomes. In these cases not only were 
flies obtained whose eyes were almost completely pigmented but other individ- 
uals had eyes with very little pigment. Part of this variation may be caused by 
inadequate visual estimates of the intensity of pigmentation since the distribu- 
tions of the measurements of "y^l^o^" fluorescence (figure 5) were less broad. 
However, most of the variation is the result of pigmentation differences in the 
flies per se and not caused by errors of estimation or experimental errors in 
measurements since in many of the genotypes these variations were not observed. 
A discussion of the possible factors which may be at the basis of this variation will 
be deferred until later. It is obvious from figure 6 that the variability encountered 
in the measurements of the "blue" fluorescence is much greater than that observed 
in the "yellow" fluorescence or with the visual estimates. In general, there is little 
variability in the amount of "blue" fluorescence when there is very little eye 
pigmentation as estimated either visually or by "yellow" fluorescence. However, 
in eyes with moderate amounts of pigmentation the value of "blue" fluorescence 
varies greatly (see figure 2). This would not appear to be caused by the insta- 
bility of these pteridines during chromatography since in many instances most 



Baker and Spofford: Heterochromatin and Variegation 



147 



of the heads of one genotype were chromatographed simultaneously under the 
same conditions. It seems more Hkely that either these pteridines accumulate 
under some sort of threshold conditions or that after accumulation they are 
rapidly used in synthesis and converted to other compounds. 

Let us return now to the main finding of this study: many of the Y fragments 
have radically different potencies in enhancing pigmentation in variegated eyes. 
The question arises as to whether the effectiveness of a fragment is positively 
correlated with the amount of heterochromatin it contains, or whether there is 
evidence of regions of heterochromatin which are more functionally active than 
other regions. One way of measuring the amount of heterochromatin is by cyto- 
logical study of the fragments. Figure 7 contains a series of photomicrographs of 
the Y fragments as seen in aceto-orcein smears of the ventral ganglion of larvae. 
It can be seen from this figure that the four ring fragments which contain only 
the Y^ fertility factors are about the same size. Yet we noted that they had quite 
different effects on the extent of variegation. For example, consider the amount 
of fluorescence in the "yellow" area of the chromatograms of attached-XY 
males. Y'^^-IS produces over six times as much "yellow" fluorescence as does 
Y^L~14, which in turn has twice the amount present in Y''^:bb+ males. In at- 
tached-X females, Y*"^^ produces over six times as much "yellow" fluorescence as 
does any of the other ring-Y^ fragments. Consider also the fragment Y^:y + bb+-6 
which, as can be seen in figure 7f, has two arms and contains the nucleolus 



Y^*f yY'-y*/Y'^iDp(w'")/-|- d* 

12 



Dp(wm)/+ Cf 




YELLOW 
FLUOR 



Fig. 5. Frequency histograms of the amount of the "yellow" fluorescing substances. 



148 



The University of Texas Publication 




*bb*-7 



BLUE 
FLUORESCENCE 



Fig. 6. Frequency histograms of the amount of the "blue" fluorescing substances. 

organizer. This moderate-sized fragment is very ineffective in enhancing pig- 
mentation in all three of the genotypes studied. One is led to the conclusion that 
there are regions of heterochromatin which are functionally more active than 
other regions, thus suppression of variegation is not entirely a matter of the 
quantity of heterochromatin. In fact, the observation that in attached-X females 
and in attached-XY males some of the fragments are more effective than even a 
complete Y leads one to believe that there may be some regions of heterochroma- 
tin that act as suppressors of the regions that are actively involved in promoting 
pigmentation. 

DISCUSSION 

The variegated-type of position effect has been studied in Drosophila for three 
decades. In spite of the impressive array of facts that have been gleaned, none of 
the testable hypotheses that have been proposed has withstood empirical exami- 
nation. Our experiments only add complexity, not clarity, to the issues involved, 
but one hopes that the marshaling of new facts will lead eventually to a general 
explanation. 

Since practically all of the V-type position effects involve a euchromatic- 
heterochromatic chromosomal rearrangement and since heterochromatic addi- 
tion to, or subtraction from, the genome changes the expression of the gene 
concerned, the functions of heterochromatin within a cell would appear to be 
the key to an explanation. Schultz (1956) has suggested, and he and his co- 
workers have obtained evidence, that heterochromatin is concerned with nucleic 



Baker and Spofford: Heterochromatin and Variegation 



149 




^% 



i%- 



\' S. 



\ -t. 



^r 



^ 



■Ir 



\ ■ 



V 

% 




-4 \^^ 



f 8 h 






'^i*.^" 








ri!i-,i / ■ 






% .!• 



- \ 










ti 



Fig. 7. The cjrtological structure of the different Y fragments. 

(a) YcL:bb+/X-X female, (b) YS:y+bb+-7/X male, (c) YVX-X female, (d) Ycl_i4/x 
male, (e) YS:bb+-8/X-X female, (f) YS:y+ bb+-6/X-X female, (g) YcS.bw+ bb+/X-X 
female, (h) Y^L-IS/X-X female, (i) YS-YS#2/X-X female, (i) scVi-Y^/X male, (k) Y^l/X 
male. (1) YL-13/X-X female, (m) YS:y+ bb+-5/X male, (n) Y/X male, (o) scSi.YL#2/X 
male. 



150 The University of Texas Publication 

acid metabolism. The importance of this finding should not be underestimated 
but its relevance to an explanation of position effect is not clear at this stage of 
our knowledge. The well-known fact should be recalled that the deletion or addi- 
tion of large heterochromatic regions has no effect on gene expression of the w+ 
locus unless a euchromatic-heterochromatic rearrangement has occurred. One 
would conclude that functioning of large heterochromatic regions is not essential 
for normal gene expression. On the other hand, once a break is produced in the 
heterochromatin and a locus normally situated in euchromatin is juxtaposed to 
the heterochromatic break, then the gene expression of this locus is altered. 
There is no evidence to indicate that any deletion of heterochromatin is neces- 
sary to produce this effect; an interruption of its continuity is sufficient. Neither 
the heterochromatic break alone nor the close proximity of a euchromatic gene 
to heterochromatin per se is a sufficient cause for the effect; both of these primary 
factors must be present for variegation to occur. 

The gene concerned must not only be close to the region of interrupted hetero- 
chromatin, it must be on the same member of the homologous pair of chromo- 
somes if its action is to be affected. It may not be generally realized that the 
variegated-type of position effect exhibits, like position pseudoalleles, the cis- 
trans phenomenon. If one considers the heterochromatic break in the rearrange- 
ment causing position effect as a region of mutant heterochromatin (h), then it 
has been shown that w+ h/w h+ produces variegated eyes whereas w + h+/w h 
flies, derived by exchange between the locus and the break point, have normal 
eyes (Judd 1955). Therefore, the primary action of the disturbed heterochroma- 
tin extends linearly along the chromosome with no interaction between horao- 
logs. This is also exemplified by the spreading effect (see Lewis 1950 for a 
review). This barrier between the gene action of each homolog does not neces- 
sarily imply a physical barrier between homologous chromosomes. Perhaps the 
chromosome products of a particular region of one homolog which are released 
into the cytoplasm retain a linear continuity encompassing part of the disturbed 
heterochromatin as well as the affected loci. If the effect on gene expression 
is mediated through synthetic activities occurring in the cytoplasm, and if 
the chromosome products of each homolog are released independently, then a 
competent spatial barrier in the cytoplasm may result. 

Now if both the primary factors for variegation are present, then its ex- 
pression and penetrance are secondarily influenced by the addition or subtraction 
of heterochromatin from the genome. If one supposes that the postulated linear 
pieces of chromosomal product are RNA, then the demonstrated effect of hetero- 
chromatin on nucleic acid metabolism might be at the root of these secondary 
effects. 

In this connection, it is interesting to note that our data indicate that the 
heterochromatin of the Y chromosome contains regions which are much more 
effective than others in influencing the extent of pigmentation. In fact, some of 
the Y fragments are more effective than a complete Y. A study of the nucleic acid 
metabolism in cells with these fragments might prove illuminating. It should 
not be forgotten that even these secondary effects on variegation take different 
directions depending on the type of gene involved in the rearrangement. Extra 
heterochromatin increases pigmentation and subtraction of heterochromatin 



Baker and Spofford: Heterochromatin and Variegation 151 

decreases pigmentation in variegated eyes caused by rearrangement involving a 
gene normally located in euchromatin, like the white locus. However, just the 
opposite relation is found with position effects of genes normally located m 
heterochromatin; e.g., the light locus (Schultz 1936). Since we have not as yet 
conducted studies on the effect of these same Y fragments on variegation at light, 
we do not know whether the fragments which are most effective in suppressing 
variegation at white are the same fragments most potent in enhancing light 
variegation, or vice-versa. We may be deluding ourselves in conceiving that the 
heterochromatin concerned secondarily with variegation has a single function; 
more than likely, it is multifunctional with spatially separated units. 

The secondary factors which affect the expression of variegation are not lim- 
ited to heterochromatic components of the genome of the individual in question, 
but extend to the genotypes of the parents (Spofford 1958). The details of these 
parental effects will be presented in another paper (Spofford, 1959), suffice it to 
say that their cause is not restricted to the amount or to the suppressing quality of 
any extra heterochromatin in the parents. For example, the extent of pigmenta- 
tion in the variegated offspring is dramatically influenced by whether or not the 
mother was homo- or heterozygous for Dp(w'"). In addition, it is influenced by 
which parent contributes the duplication and by whether or not the mother is 
homozygous for a recessive modifier which segregates independently from the 
duplication. Thus there is an array of hereditary factors, some intrinsic, others 
not, which mimic one another in their effects on variegation. The environmental 
temperature has an action similar to the hereditary factors (Gowen and Gay 
1933). 

It seems clear that unless these secondary factors are under experimental con- 
trol, the interpretation of data on position-effect variegation may be ambiguous. 
The secondary factors which were not under control when our studies were initi- 
ated were the parental effects just mentioned; in fact, their presence was only 
discovered during the course of the experiments. One of these factors which could 
be responsible for a portion of the variation observed between individuals of a 
given genotype is whether or not the mother was homozygous for the duplication. 
The duplication was of maternal origin in the X/Y^ males and the XX/Y^ fe- 
males. An examination of the data on the extent of pigmentation and "yellow" 
fluorescence (figures 4 and 5) shows no more intra-genotypic variation among 
individuals of these two genotypes than in XY/Y^ males although the pene- 
trance figures (tables 1, 2 and 3) suggest less variabihty in the latter males. One 
can only conclude that an unmeasured portion of the variability in X/Y^ males 
and XX/Y^ females should be attributed to this cause. 

The parental effect which might make inter-genotypic comparisons ambiguous 
is the observation that more pigmentation in the variegated eyes is produced if 
the duplication and the sex chromosome bearing the mutant allele of white are 
both contributed by the father rather than the mother. In our experiments, the 
XY/Y^ males received these two elements from their father, whereas the X/Y^ 
males and the XX/Y^ females received them from their mothers. Therefore, the 
general increase in pigmentation in XY/Y'^ males over the other two genotypes 
may be at least partly the result of this parental effect. However, note that with 



152 The University of Texas Publication 

four of the fragments Y^, Y*'^^ Y^-13, and sc^^-Y^, more pigmentation was present 
in the attached-X females than in attached-XY males. 

Finally, the high degree of pigmentation in XY/Y^:y+ bb+-5 males is un- 
doubtedly the result of the "residual effect" of this frgament. Spofford (1959) 
has shown that this fragment in attached-X mothers produces more pigmentation 
in variegated offspring — irrespective of the Y fragment present in the offspring — 
than several other Y fragments in attached-X mothers. 

Two interesting inter-genotypic comparisons should be mentioned. It was 
noted that the penetrance of variegation was higher in both types of males than 
in the attached-X females. This observation can be extended to free-X, y w/y w; 
Dp(w"') females since they also show low penetrance (unpublished data). It 
appears that penetrance is inversely correlated with the dosage of the mutant 
allele of white. If these white alleles are amorphs, there is a balance, relating 
the number of sets of X-chromosome genes to gene expression, which requires 
more activity of Dp(w"') in females than in males for pigment formation. 

Another inter-genotypic comparison of interest is between the variegation seen 
in Y^w yY^y+/0 males and y w/Y males (see tables 1 and 3). Although the 
former males contain as much, and probably more, heterochromatin than the 
latter males, only very rarely do their eyes show any pigment at all and then 
only just a trace. The X/Y males are quite heavily pigmented. This observation 
raises again the possibility that a certain amount of structural integrity of the 
heterochromatin must be retained if its functions in suppressing variegation are 
to be maintained. Three alternatives are suggested: (1) The Y heterochromatin 
is subdivided into separate functional units, in so far as their effect on variegation 
is concerned, and during the synthesis of the attached-XY chromosome (Lindsley 
and Novitski 1950) some of the potent regions were lost. (2) Irrespective of 
whether or not the Y heterochromatin is delineated into functional units, the 
effectiveness of the whole depends on the structural integrity of its parts. (3) The 
activity of the hypothetical functional units in the Y may be suppressed by a re- 
arrangement which associated them with euchromatin. It would be interesting to 
see if the XY^-Y^ chromosome of Parker (1954), derived from detachment of 
an attached-X with the sc^Y, shows this same reduced activity towards suppres- 
sion of variegation. In any event these observations should serve to warn 
one against the possible pitfalls of using attached-XY chromosomes to increase 
the amount of active heterochromatin in a genome. 

One additional finding of ours deserves further comment. The increased 
amount of the sepia pteridine in variegated eyes over that found in wild type 
points to the possibility that steps in the synthesis of pigment are being affected 
which occur later than the step ostensibly controlled by the white locus. This 
conclusion is reached because the sepia pteridine is not found in adult flies with 
white eyes no matter whether they are homozygous for w or en bw (Hadom and 
Mitchell 1951). Since even bw flies do not contain any of this pteridine, one 
would guess that one of the steps in the synthesis of the red pigments affected by 
Dp(w'") follows after the step controlled by the bw locus. One might still retain 
the one gene-one action hypothesis and explain this dichotomy of action of the 
white locus by assuming that its primary function is the production of a co- 
enzyme, or perhaps a polypeptide, which is a component of at least two different 



Baker and Spofford: Heterochromaiin and Variegation 153 

enzymes controlling the synthesis of pigment. One of the enzymes may control 
a reaction leading to intermediates that are common to both the red and the 
brown pigments: whereas, the other enzyme might be concerned with the me- 
tabolism of the sepia pteridine. Further speculation at this time seems fruitless, 
but at least the possibility should be kept in mind that the white locus may 
not be directly involved in the synthetic pathways leading toward pigment 
formation. 

The altered gene expression caused by Dp(w'") is distinguishable from that 
seen with either the w'^ or the w^ mutant alleles. Hadorn and Mitchell, loc. cit., 
found a reduced value of Fl-4 plus Fl-5 (the sepia pteridine) in these mutants as 
compared with wild type. This fact in itself suggests that variegation is not the 
result of somatic mutation but rather altered gene action, often in a clonal pat- 
tern. The results of further experiments addressed to this topic will be reported 
in a subsequent paper. 

ACKNOWLEDGMENTS 

The authors wish to express their appreciation to Miss Carmen Kanapi, who 
greatly assisted in the tedious chromatographic analyses; to Dr. Eileen S. Gersh 
who not only provided the duplication stock but many hours of stimulating 
discussion; and to Dr. Hugh S. Forrest for the pteridine samples. 

SUMMARY 

The effect of 15 different fragments of the Y chromosome (Y^'s), in suppres- 
sing position-effect variegation was studied in Drosophila melanogaster. The 
chromosomal rearrangement responsible for the variegation is an insertion of the 
w+ locus into the centromeric heterochromatin of 3L and is designated Dp(w'"). 
The suppressing action of these fragments on the size of the white areas in the 
variegated eyes was recorded visually and chromatographically in flies of the 
following genotypes who were otherwise coisogenic: y w/Y^; Dp(w"0/+ males, 
yw/Y^; Dp(w"0/+ attached-X females, and Y^wyY^y+ZY^ Dp(w-)/+ 
attached-XY males. 

Many of the Y^'s were found to differ markedly in their potency; some even 
more effective than a complete Y. Cytological examination indicates that sup- 
pressing activity is not necessarily correlated with the amount of heterochroma- 
tin in the fragment. Thus the conclusion is drawn that Y heterochromatin may be 
subdivided into functional units. 

Variegated eyes with an intermediate extent of pigmentation accumulate an 
amount of the sepia pteridine greatly in excess of that found in wild-type eyes; 
whereas, white eyes contain none of this compound. This suggests that the action 
of w+ which is being altered by position effect occurs at a rather late stage in the 
synthesis of the red pigments. 

LITERATURE CITED 

Baker, W. K. 1955. On the structure of the sc^:Y:bw+ chromosome of D. melanogaster. Dro- 
sophila Info. Service 29:101-103. 

Baker, W. K. 1957. Induced loss of a ring and a telomeric chromosome in Drosophila melanogaster. 
Genetics 42:735-748. 



154 The University of Texas Publication 

Demerec, M. 1940. Genetic behavior of euchromatic segments inserted into heterochromatin. 
Genetics 25:618-627. 

Forrest, H. S., and H. K. Mitchell. 1955. Pteridines from Drosophila. III. Isolation and identifica- 
tion of three more pteridines. Jour. Amer. Chem. Soc. 77:4865-4869. 

Gowen, J. W., and E. H. Gay. 1933. Effect of temperature on ever-sporting eye color in Drosophila 
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Hadorn, E., and H. K. Mitchell. 1951. Properties of mutants of Drosophila melanogaster and 
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Judd, B. H. 1955. Direct proof of a variegated-type position effect at the white locus in Drosophila 
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Lewis, E. B. 1950. The phenomenon of position effect. Adv. in Genetics 3: 73-1 15. 

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Muller, H. J. 1948. The construction of several new types of Y chromosomes. Drosophila Info. 
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Parker, D. R. 1954. Radiation-induced exchanges in Drosophila females. Proc. Nat'l. Acad. Sci. 

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, and J. McCrone. 1958. A genetic analysis of some rearrangements induced in oocytes 



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Patterson, J. T. 1928. The effects of X-rays in producing mutations in the somatic cells of Dro- 
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Schultz, J. 1936. Variegation in Drosophila and the inert chromosome regions. Proc. Naf 1. Acad. 
Sci. 22:27-33. 

■ . 1956. The relation of the heterochromatic chromosome regions to the nucleic acids of 

the cell. Cold Spring Harbor Symp. Quant. Biol. 21 : 307-328. 

Spofford, J. B. 1958. Parental control of position-effect variegation. Proc. Xth Intern. Congr. 
Genetics 2:270. 

. 1959. Parental control of position effect variegation: I. Parental heterochromatin 

and expression of the white locus in compound-X Drosophila melanogaster. Proc. Natl. 
Acad. Sci. 45 (in press) . 



Genetic Studies on the Cardini Group of Drosophila 
in the West Indies 

WILLIAM B. HEED AND NIKAM B. KRISHNAMURTHY 
University of Arizona, and Bangalore, India 

INTRODUCTION 

Geographic isolation is the main prerequisite to the initiation of genetic di- 
vergence of sexually reproducing populations. Island populations afford an ex- 
cellent opportunity to study the effects of isolation, especially when the forms 
under study can be crossed in the laboratory. The cardini group of Drosophila, 
restricted chiefly to the Neotropical Region, exhibits an interesting distributional 
pattern in the West Indies, part of which has recently been discussed (Heed, 
1957a), and contains an array of forms which can be subjected to laboratory 
analysis. 

The present report deals mainly with the genetic and cytological affinities of 
the forms inhabiting the Lesser Antilles and Puerto Rico, described below as the 
dunni subgroup of the cardini group. The series is named after D. dunni, de- 
scribed by Townsend and Wheeler (1955) from Puerto Rico. The taxonomy of 
the series has not been completely worked out and will be reported separately. 
The geographic origin, stock number and symbol of each of the strains studied 
is given below; additional collection data are shown in Table 1 7. 

D. acutilabella: St. Petersburg, Fla. (F), 2099.1; Floral City, Fla. (Fl), 
2302.12; Cuba (C), 2380.2; Jamaica (J), H137.5; Haiti (H), H135.7. 

D. belladunni: Jamaica ( Jb) , H356.3. 

D. dunni subgroup: Puerto Rico (PR), 2327.1; St. Thomas (ST), H253.5; St. 
Kitts (SK), H126.1; Guadeloupe (GU), H252.7; Martinique (MA), H248.1; St. 
Lucia (SL), H122.1; Barbados (BA), H247.1; St. Vincent (SV), H240.1; Gre- 
nada (GR),H239.6. 

Figure 1 illustrates that, in general, each of the islands has its own pheno- 
typically distinguishable population and that the change in abdominal pattern 
from north to south forms a more or less regular sequence or cline. Such an 
obvious cline in abdominal pattern is a rare phenomenon in Drosophila species. 
The chain of islands of the Lesser Antilles and Puerto Rico may be exactly super- 
imposed on the state of Florida, for they extend in an arc only 734 miles. The 
majority of the main islands are separated from one another by 20-30 miles of 
open water. This affords the isolation necessary to provoke divergence of an 
otherwise more or less continuous genetic system. The adaptive significance of 
the darker flies to the south and lighter flies to the north is not known. A study of 
the microhabitats on the islands in conjunction with an analysis of the inheri- 
tance of the abdominal pattern may yield some information on this, for most 
assuredly the chne is the result of present day selective values superimposed upon 
the past history of the group. The tests made to date indicate that the inheritance 
of the abdominal pattern depends on a number of genetic factors, apparently of 



156 



The University of Texas Publication 



unequal effects, and is difficult to analyze. Further tests must be made before a 
report on this phase of the study is possible. 

Another interesting feature of the dunni subgroup which can be briefly men- 
tioned is that the larvae of every island stock exhibit a low rate of cannibalism. 
Larvae have been observed to feed on other apparently normal larvae (as well as 




Fig. 1. Abdominal patterns of the dunni subgroup females in the Lesser Antilles and Puerto 
Rico; the pattern of belladunni from Jamaica is also shown. 



Heed and Krishnamurthy : Cardini Group of Drosophila 157 

tumorous larvae, which are fairly frequent) and also to scrape their way into 
puparia and feed on their contents. This phenomenon has not been previously 
reported in Drosophila and deserves further investigation. It is interesting that 
the cardini group is one of the groups of Drosophila that have skipping larvae. In 
the case of the dunni subgroup the skipping habit would have a distinct selective 
advantage, on the assumption that cannibalism per se is a detriment to the popu- 
lation. The assumption however may not be correct. Also there is the possibility 
that the larvae are predatory on larvae of other species. This has not yet been 

tested. 

The following report is divided into four sections. The first two sections briefly 
describe the distribution of the group on the islands and the collection data as it 
concerns the dunni subgroup. The purpose is to show that this subgroup is char- 
acteristic of the Lesser Antillean and Puerto Rican faunas both in natural and 
domestic environments. The last two sections concern the hybridization tests, 
carried out mostly by the senior author, and the cytological analyses, made by 
the junior author. 

TAXONOMY AND DISTRIBUTION 

The cardini species group of the subgenus Drosophila was estabhshed by 
Sturtevant in 1942 for Drosophila cardini Sturtevant, with similis Williston, 
albirostris Sturtevant and metzii Sturtevant indicated as other possible members. 
Since that time the following species have been added to the group: cardinoides 
Dobzhansky and Pavan, neocardini Streisinger, polymorpha Dobzhansky and 
Pavan, acutilabella Stalker, parthenogenetica Stalker, dunni Townsend and 
Wheeler, neomorpha Heed and Wheeler, nigrodunni Heed and Wheeler, and 
procardinoides Frydenberg, while albirostris and metzii have been placed in the 
tripunctata group. 

Our studies of the forms on the Lesser Antilles and Puerto Rico showing a cline 
in abdominal pattern show that they constitute a dunni subgroup, clearly differ- 
ing from the remainder of the cardini group. This is a natural subgroup, the 
members being more closely related to one another than to other members of the 
group. A distinguishing feature is that nearly every island population has its own 
distinct phenotype, especially in the abdominal pattern. The arrangement of 
bristles on the palpi (5-6 long bristles on the antero-lateral margin) and the 
cannibahstic habits of the larvae are also common to all members of the 
subgroup. 

The members thus far described are dunni Townsend and Wheeler (1955) 
from Puerto Rico, and nigrodunni Heed and Wheeler (1957) from Barbados. 
Drosophila similis, described by Williston from St. Vincent, may be conspecific 
with the form that we have studied from that island. 

The new species described below, Drosophila belladunni from Jamaica, is not 
included in the dunni subgroup since it will not cross as well with those forms as 
they do with one another. 

Drosophila belladunni, new species 
This new species from Jamaica is sympatric with and very similar in abdom- 
inal pattern to D. acutilabella Stalker (1953). The bristle arrangement of the 



158 The University of Texas Publication 

palpi and the male genitalia are more similar, however, to those of the members 
of the dunni subgroup. There are no good obvious phenotypic characters that will 
always distinguish the females of these two species in Jamaica; characters that 
sometimes work are the larger size and more extensive abdominal pattern of 
acutilabella females. The abdominal pattern of belladunni of both sexes will dis- 
tinguish them from all members of the dunni subgroup (Fig. 1 ) . 

Males of belladunni and acutilabella can be distinguished by the following 
characters as based upon belladunni. 

Labellum. No acute process at tip as in acutilabella. 

Palpi. With 5-6 long bristles on antero-lateral margin, the subapical bristle 
the longest, the others of about equal length. There are 5-6 bristles on the antero- 
lateral margin in acutilabella, but the first 3 are longest and stoutest, with the 
apical one the longest of the three, the remaining 2-3 bristles gradually becoming 
shorter posteriorly. 

Abdomen. Tergites 2, 3, and 4 with narrow posterior dark bands interrupted 
dorsally; 5th and 6th solid black except on lateral margin (Fig. 1); not known 
to be polymorphic. The pattern in acutilabella is sometimes very similar to that 
of belladunni in Jamaica, but also sometimes with dark lateral extensions con- 
necting 3rd and 4th segments at angle of tergite and sometimes with light para- 
median extensions on 2nd and 3rd tergites. 

Genitalia. Forceps with Q-7 primary teeth and 7-8 secondary teeth; lower tip 
of anal plate with 3-4 medium length bristles. D. acutilabella with about 6 pri- 
mary and 9-10 secondary teeth on forceps, the lower tip of anal plate with sub- 
apical extension bearing one exceptionally stout long curved bristle. 

Chromosomes. Metaphase plate preparations show 2 pairs of V's, one smaller 
than the other, and 2 pairs of rods of which one pair stains darker than the other. 
The lighter staining rod is the X-chromosome; the darker staining rod is the dot 
(4th) chromosome with added heterochromatin. Salivary chromosomes show 5 
arms and a dot. This description is based upon stock No. H356.3. D. acutilabella 
has 2 pairs of V's, one pair of rods, and one pair of dots. 

Distribution and Types. Drosophila belladunni belongs to the cardini species 
group of the subgenus Drosophila. It is known only from Jamaica, B. W. I., where 
it is usually collected in association with acutilabella. In the summer of 1958 
belladunni made up 11% of 196 males of the two species collected at three local- 
ities in the lowlands along the north coast. At Hardware Gap (4,380 feet), how- 
ever, belladunni made up 98% of the 56 males collected. From this data bella- 
dunni is considered an older member of the Jamaican fauna than acutilabella. 

Holotype male and paratype males and females, from stock No. H356.3d from 
Hardware Gap, Jamaica, British West Indies, deposited in the University of 
Texas collection. 

DISTRIBUTION 

The distribution of the species of the cardini group in the West Indies is not 
fully known, but enough collecting has been done to emphasize several points 
(Table 1). Drosophila acutilabella is definitely absent on Puerto Rico; it is lim- 
ited to southern Florida and the three islands indicated. Members of the dunni 
subgroup inhabit the Lesser Antilles and Puerto Rico, including the Virgin 



Heed and Kris hnamur thy: Cardini Group of Drosophila 159 

Table 1 
Distribution of the cardini group species in the Caribbean region 





Southern 
Florida 


Cuba 


Jamaica 


Haiti 


I'lieito Rico 


Lesser 
Antilles 


-r.irndad and 
South America 


cardini 
acutilabella 


X 

X 


X 
X 


X 
X 


X 
X 


X 


Xf 


X 


belladunni 






X 










dunni subgroup 
polymorpha 
Other spp.* 










X 


XJ 

x§ 


X 
X 



* cardinoides, neomorpha and undescribed sp. 

f Grenada, St. Vincent, St. Lucia and Martinique only. 

X All 8 islands visited. 

§ Grenada only. 

Islands; they definitely do not extend to Trinidad or to the mainland of South 
America. 

The closest relatives of the dunni subgroup are acutilabella and belladunni, 
described above. They are sibling species, living sympatrically in Jamaica, and 
with the females being indistinguishable. However, belladunni is more frequent 
in the highlands while acutilabella is more frequent in the lowlands. 

Hispaniola has been sampled only once and it is not known whether a member 
of the subgroup inhabits this large island. Similarly, collections in Cuba have 
been too fragmentary to be significant. 

Drosophila polymorpha is the only cardini group species known to bridge the 
87-mile gap between Trinidad and Grenada, aside from the widely distributed 
cardini, but it has not yet dispersed any farther north. It is interesting that this 
species is not polymorphic in Trinidad or Grenada as it is in Brazil (da Cunha 
1949), the light form being the only one ever collected in the islands. Here is 
more evidence that marginal populations cannot afford the luxury of poly- 
morphism. 

POPULATION SAMPLES 

The dunni forms are well established on all the islands collected, judging from 
the fact they are usually among the first three most common forms collected over 
natural or artificial bait. Tables 2 and 3 Hst a few examples of the frequencies of 
the most numerous species from two general habitats on the islands. The wood- 

Table 2 
The three dominant species from woodland habitat samples 





PR 

Jan. 31 


SK 
Jan. 21 


SL 
Jan. 17 


BA 

Jan. 2 


sv 

July 24 


No. Species 
No. Individuals 


7 
351 


9 

515 


9 

579 


5 
322 


7 
1,206 


Percent willistoni group 
Percent dunni subgroup 
Percent others 


56.7 

22.2 
17.9* 


59.6 

25.2 
7.0t 


54.2 
23.8 
7.9$ 


93.5 
5.3 
0.6$ 


14.6 
66.7 
17.6t 



simulans (very few melanogaster) ; f melanogaster., % sturtevanti. 



160 The University of Texas Publication 

Table 3 
The three dominant species from domestic and semi-domestic habitat samples 





PR 
Aug. 18 


PR 
Feb. 4 


GU 
Aug. 3 


MA 

July 31 


SL 
Jan. 18 


BA 
Jan. 5 


GR 

July 22 


No. species 


9 


14 


15 


13 


10 


9 


13 


No. individuals 


890 


394 


690 


665 


1,448 


793 


831 



42.9 


13.6 


11.0 


12.6 


15.6 


15.2 


(4.9) 


(7.8) 


13.4 


75.0 


49.7 


(6.3) 




42.0 


64.4 






30.4 



Percent willistoni group 68.5 
Percent dunni subgroup 6.0 

Percent melanogaster 18.5 
Percent ananassae ... 10.2 ... ... ... 22.1 33.0 

Percent others . . . 22.1* 9.3f . . . 3.8$ 

* mediodiffusa: 7 ornatipennis: ± metzii. 

land habitat includes such areas as the Caribbean National Forest in Puerto Rico, 
a government reserve forest in St. Vincent, and Turner's Hall Wood in Barbados. 
The domestic and semidomestic habitats include local secondary woods, cacao 
and banana fincas, orchards and garbage dumps. The dunni forms are more 
consistently common in the relatively undisturbed woodlands and they may be 
considered a characteristic part of the native fauna of the islands. 

The sample from Barbados in Table 2 from an orchard is a selected sample in 
favor of the dunni forms. They were very numerous on a pile of freshly cut 
citrus branches and twigs. The sample from St. Lucia however, from a baited 
local secondary woods, is not selected. It is possible that the dunni forms are 
presently being transported from island to island by man and by other means, 
although not at high enough rate in most cases to obviate either the phenotypic 
identity of most island types or the genetic sterility found in some of the crosses. 

HYBRIDIZATION TESTS 

Two sets of mass matings were made independently by both investigators 
(Tables 4 and 5). Table 4 gives higher total numbers because the larvae were 
treated with yeasted kleenex. Table 5 includes a Puerto Rico strain and no control 
(homogamic) matings were made. A strain from St. Lucia was later tested by 
one set of mass matings (approx. 20 males and 20 females) with four other island 
strains. The tests were run for only one month and were not run to extinction as 
in the above crosses. The results are listed separately in Table 6. Twenty-five sets 
of 10 pair matings each were made and the results are listed in Table 7. 

The first generation hybrids from all the original crosses except those involv- 
ing the St. Lucia strain were inbred. The tests that gave an Fo generation or more 
are listed in Table 8. Backcross tests were made with most of the hybrid females 
to usually both types of original strain males when the hybrid stock would not 
inbreed. The backcross offspring were allowed to inbreed for several weeks. After 
this time the number of vials that showed larvae out of the total number of vials 
for each set was recorded (Table 7). The original hybrid males were not tested 
for fertility in backcrosses. 

From the tables it can be seen that all degrees of genetic relationship between 



Heed and Krishnamurthy: Cardini Group of Drosophila 



161 



Table 4 

Total number of hybrids and controls from two mass matings of approximately 20 $ and 20 i}, 
each with yeasting method. Results of the second mass mating are shown in parentheses 





^ST 


GU 


BA 


SV 


GR 


9 


9 ^ 


5 $ 


9 $ 


9 6 




498:362 


64:46 


273:205 


7:2 


0:2 


ST 


(892:809) 


(23:31) 


(168:117) 


(0:2) 


(1:1) 




39:18 


567:391 


587:559 


79:64 


20:26 


GU 


(29:32)f 


(917:740) 


(856:782) 


(26:25) 


(62:58) 




47:19 


937:215* 


746: 741 


151:183 


9:41 


BA 


(39:15)t 


(1,548:329)* 


(1,501:1,278) 


(13:61) 


(10:101) 




87:52t 


405:48* 


642:602 


730:675 


711:659 


SV 


(no count) 


(232:36)* 


(835:609) 


(696:601) 


(578:570) 




42:21f 


828:5* 


798:666 


541:444 


628:511 


GR 


(10:13)t 


(768:10)* 


(852:592) 


(944:825) 


(706:638) 



* One-half of males with abnormal abdomens. 

-j- Approximately one-sixth of males with abnormal abdomens 



Table 5 - 

Total number of hybrids from two mass matings of 20 9 and 20 $ each without yeasting method. 
Results of the second mass mating are shown in parentheses. 





^PR 


ST 


GU 


BA 


SV 


GR 


9 


9 $ 


9 $ 


9 $ 


9 $ 


9 $ 


9 $ 






31:12 


8:14 


15:18 


1:1 


0:0 


PR 




(81:67) 


(18:17) 


(50:46) 


(2:0) 


(0:0) 




230:152 




46:64 


142:153 


9:19 


0:0 


ST 


(109:54) 




(74:45) 


(111:104) 


(1:3) 


(4:1) 




6:1 


53:10 




141:129 


56:44 


55:44 


GU 


(17:10) 


(14:8) 




(264:145) 


(11:5) 


(38:37) 




7:5 


5:1 


362:113 




45:83 


77:141 


BA 


(4:10) 


(15:8) 


(137:18) 




(30:67) 


(35:51) 




1:0 


8:6 


34:3 


13:30 




20:29 


SV 


(0:0) 


{7:7) 


(119:2) 


(150:124) 




(38:45) 




1:0 


13:12 


45:0 


20:29 


40:22 




GR 


(5:3) 


(17:13) 


(90:2) 


(107:101) 


(16:6) 





Table 6 
SL crosses for one month only 



SLxSL 


94 


74 


SLX ST 


6 





SLX GU 


50 


12 


SLxSV 


48 


10 


SLxBA 


259 


261 



STX SL 
GUx SL 
SVx SL 
BAX SL 





75 



105 




48 


99 



162 The University of Texas Publication 

the islands are exhibited from complete fertility in the F. as in the case of PR X 
ST (and reciprocal) and SV X GR (and reciprocal), to strong sexual isolation 
especially in the cases of PR and ST with SV and GR and with some oi the tests 
with SL. A few dissections were made to check for sexual isolation; however, 

Table 7 
Pair matings in the dunni subgroup 





No. vials fertile 


Tote 


dNo. 


x\ ^0^' P 


Average per 


fertile ? 


? cT 


from 10 pairs 


?? 


cfcf 


a 1 : 1 value 


9? 


dd 


STxST 


6 


334 


288 


3.40 >.05 


55.7 


48.0 


GUxGU 


7 


378 


314 


5.92 >.01 


54.0 


44.9 


SVx SV 


8 


473 


374 


11.58 <.01 


59.1 


46.8 


GRX GR 


9 


920 


856 


2.30 >.05 


102.1 


95.1 


BA X BA 


9 


142 


151 


.276 >.50 


15.8 


16.8 


STXGU 


1 


4 


3 




4.0 


3.0 


STx SV 















STx GR 















ST X BA 


6 


39 


42 


not significant 


6.5 


7.0 


GUx ST 


2 


17 


7 




8.5 


3.5 


GU X SV 


4 


45 


41 


not significant 


11.3 


10.3 


GU X GR 


3 


40 


36 


not significant 


13.3 


12.0 


GUxBA 


10 


423 


381 


2.2 >.05 


42.3 


38.1 


SVx ST 


1 


1 


2 


.... 


1.0 


2.0 


SVx GU 


9 


412 


20 


significant 


45.8 


2.2 


SV X GR 


7 


331 


315 


not significant 


47.3 


45.0 


SVx BA 


6 


282 


224 


6.64 =.01 


47.0 


37.3 


GRx ST 


2 


3 











GRXGU 


7 


358 


3 


significant 


51.1 


0.43 


GRx SV 


7 


594 


530 


3.64 >.05 


84.9 


75.7 


GRXBA 


6 


256 


210 


4.54 >.01 


42.7 


35.0 


BAX ST 


1 


1 





.... 






BAxGU 


10 


370 


164 


significant 


37.0 


16.4 


BAx SV 


4 


16 


26 


2.5 >.05 


4.0 


6.5 


BA X GR 


2 


5 


29 


significant 


2.5 


14.5 



Table 



total nun 


iber ot mbr 


ed hybrid ottspnng i 


rom crosses in laoie 


s % J axiu / 






F, 


Fs 




9 $ 




$ 9 ^ (5 


99 $ $ 


99 $ $ 


PRX ST 




Fertile 


Fertile 


Fertile 


STx PR 




Fertile 


Fertile 


Fertile 


SVxGR 




Fertile 


Fertile 


Fertile 


GRXSV 




Fertile 


Fertile 


Fertile 


SVxGU 




222:88 


29:29 


275:218* 1 


BAxSV 




217:144 


None 




BAxGU 




9:6 


None 




GUxBA 




3:2 


None 





GUxBA 




32:24i 


None 




* Sample count fi-om hybi 


-id stock. 








t Original parents one ma 


ted pair. 









Heed and Krishnamurthy : Cardini Group of Drosophila 



163 



Table 9 
Backcrosses of 23 9 9 per cross except where marked 







NT r 


Inbred Offspring 




N( 


) of 


Inbred Offspring 


Backcross 


Oifspring 

? c^ 


No. of Fertile Vials 


Backcross 


Offs 


pring 


No. of Fertile Viah 


?/^Xcf 


No. of V.als 


?/cf Xd" 


? 


d 


No. of Vials 


GU/STxST 










GU/SVxSV 


Ill 


95 


0/5 


ST/GUxST 










BA/SVxSV 


310 


196 


1/6 


*ST/SVxST 


90 


53 


0/2 


fGR/STxGR 


10 


10 


1/1 


SV/STxST 


221 


82 


1/6 


GR/GUxGR 


207 


92 


0/6 


fGR/STxST 


173 


71 


1/5 


JGU/GRxGR 


106 


62 


0/3 


ST/BAxST 


86 


44 


0/4 


GR/BAxGR 


235 


174 


1/7 


GU/STxGU 










ST/BAxBA 


80 


21 


0/2 


ST/GUxGU 










GU/BAxBA 


194 


185 


V5 


GU/SVxGU 


202 


80 


2/3 


BA/SVxBA 


298 


242 


2/6 


GR/GUxGU 


229 


55 


1/2 


GR/BAxBA 


432 


259 


3/6 


GU/BAxGU 


233 


118 


1/4 


§BA/GRxBA 


255 


207 


2/6 


SV/STxSV 


89 


46 


1/4 










f 12?? 
i 18?? 
















§ 9 ?? 

















most of the low number hybrids are inferred to be due to sexual isolation. The 
two extremes in genetic relationship agree well with the geography of the islands 
(notice that St. Vincent and Grenada are "bridged" by the Grenadines). 

In regard to sexual activity another inference may be made. BA males have 
a strong sexual drive, for they consistently give a high number of hybrids with 
other island females and produced the highest number of offspring in homogamic 
mass matings. The high sexual activity may be of a density-dependent nature 
since BA homogamic pair matings produced the lowest number of offspring com- 
pared to the other four homogamic matings (Table 7) . 

Some very obvious unequal sex ratios may be seen in the F^ hybrids (Tables 
4, 5, 6 and 7). The most consistent abnormal ratios are from the mating of GU 
males with other island females and from the mating of BA females with other 
island males. Tables 10 and 11 show this relationship. The data are extracted 
from the tables mentioned above. The reciprocal crosses of these matings usually 
produced a normal sex ratio. 

Concerning the crosses with GU males, Table 10 shows a decrease in hybrid 
males with more or less increased distance from the island of Guadeloupe. The 
cline in sex ratio is illustrated in relation to the geography of the islands in Table 
12. The data in the right hand column are taken from the backcross tests (Table 
9). The reciprocal crosses, hybrid GU females to males of other islands, gave a 
1 : 1 ratio with BA and SV males and a 2: 1 ratio with GR males. 

There are several factors to be considered here: the cline in phenotype, the sex 
ratio cline and the geographic position of the islands. Barbados is the most iso- 
lated island (92 miles from St. Lucia) and contains the darkest individuals. 
However it does not have as severe an abnormal sex ratio as the St. Vincent and 
Grenada strains. The St. Vincent and Grenada strains are phenotypically iden- 



164 



The University of Texas Publication 



Table 10 
Sex ratio cline of Guadeloupe males with females of other islands from five sets of crosses 



Cross 

9 $ 



Females 



Males 



Ratio 



GUxGU 




567 


391 


1.4:1* 






917 


740 


1.2:1* 






378 


314 


1.2:11 




Total 


1,862 


1,445 


1.3:1* 


BAX GU 




937 


215 


4.4: 1 






1,548 


329 


4.7:1 






370 


164 


2.3: If 






362 


113 


3.2:1 






137 


18 


7.6:1 




Total 


3,354 


839 


4.0:1 


SLxGU 




50 


12 


4.2:1 


SVxGU 




405 


48 


8.4:1 






232 


36 


6.4:1 






412 


20 


20.6: If 






34 


3 


11.3:1 






119 


2 


58.5:1 




Total 


1,202 


109 


11.0:1 


GRxGU 




828 


5 


165.6:1 






768 


10 


76.8:1 






358 


3 


118.9:lf 






45 





.... 






90 


2 


45 :1 




Total 


2,089 


20 


104.5:1 



* Deviation from 1 :1 ratio highly significant in homogamic matings. 
7 Totals from pair matings. 

tical and produce a highly fertile F2; however, crosses of the strains to GU males 
show a significant difference in sex ratio. 

The nature of the lethality in the hybrid males has not yet been determined. 
Egg counts were not made, but it is assumed that the cross-lethal effect acts 
between the zygote and pupal stages since the control crosses of GU x GU gave 
a much higher proportion of males. That is, any male meiotic disturbance such 
as that leading to the "sex-ratio" effect analyzed by Sturtevant and Dob- 
zhansky (1936) in D. pseudoobscura is ruled out. Also there is no reason to be- 
lieve that a cytoplasmic factor is causing unequal sex ratios as reported by Malo- 
golowkin (1958) for D. willistoni and paulistorum. Our unequal ratios are de- 
pendent on the genotype of the male, at least, and probably also of the female. 

The backcrosses of the hybrid females to GU males show that even though the 
progeny have predominantly GU chromosomes the sex ratio cline is still in evi- 
dence. This gives some indication that the cross-lethal effect is an unfavorable 
interaction between a factor (or factors) on the other island X-chromosomes 
with the GU Y-chromosome. 



Heed and Krishnamurthy: Cardini Group of Drosophila 



165 



The only cross involving GU males that gave sufficient numbers in the F^ for 
comparison with the above data is that of SV x GU (Table 8) . The hybrid mating 
produced a ratio of 222 females : 88 males or 28.4 percent males. This is the same 
proportion of males produced in the backcross of GU/SV X GU. Thus the cross 
lethal effect is acting in a consistent fashion regardless of the proportion of GU 
autosomes in the hybrids. 

The F3 generation gave a normal sex ratio indicating that the cross lethal effect 
had been nullified due either to more recombination of the original SV X chromo- 
some by crossing over or due to selection for the GU X chromosome or most of it. 

Table 11 
Sex ratio cline of Barbados females with males of other islands from five sets of crosses 



Cioss 

9 $ 



Females 



Males 



Ratio 



BAx ST 




47 


19 


2.5:1 






39 


15 


2.6:1 






1 





....t . 






5 


1 


5 :1 






15 


8 


1.9:1 




Total 


107 


43 


2.5:1 


BAx GU 




937 


215 


4.4:1 






1,548 


329 


4.7:1 






370 


164 


2.3: If 






362 


113 


3.2:1 






137 


18 


7.6:1 




Total 


3,354 


839 


4.0:1 


BAx SL 




105 


99 


1.1:1$ 


BAxBA 




746 ^ : 


741 


1.0:1 






1,501 


1,278 


1.2:1* 






142 


151 


1 :l.li 




Total 


2,389 


2,170 


1.1:1* 


BAx SV 




133 


167 


1 :1.3t 






13 


61 


1 :4.7 






16 


26 


1 :1.6it 






45 


83 


1 :1.8 






30 


67 


1 :2.2 




Total 


237 


404 


1 :1.7 


BAxGR 




9 


41 


1 :4.6 






10 


101 


1 :10.0 






5 


29 


1 :5.8f 






77 


141 


1 :1.8 






35 


51 


1 :1.5$ 




Total 


136 


363 


1 :2.7 



* Deviation from 1 :1 ratio highly significant in honiogamic niatings. 

f Totals from pair matings. 

+ Deviation from 1 : 1 ratio not significant in heterogamic matings. 



166 The University of Texas Publication 

Table 12 
Sex ratios obtained in crosses with Guadeloupe (GU) males 



? d 


Degrees N. 
latitude 


Miles through 

connecting 

islands 


Percent d d 

from all 

parental crosses 


Percent dd 
from hybrid 

?9xGUcf(^ 


GUxGU 


16° 15" 





43.7 




SL X GU 


13° 55" 


140 


19.4 




BAxGU 


13° 10" 


255 


20.0 


33.6 


SVxGU 


13° 15" 


190 


8.3 


28.4 


GRXGU 


12° 5" 


280 


0.9 


19.4 



A sample count from the Y^ hybrid stock gave 44.2 percent males, which is more 
similar to the proportion of males regularly produced within the GU strain 
(43.7%) than within the SV strain (47.2%). There was also strong selection 
toward the GU phenotype after the second generation. The Fg hybrid progeny 
could not be distinguished from the GU phenotype. Contamination of this stock 
by GU individuals is ruled out since biochemical studies on enzyme activity now 
in progress show that the stock is of hybrid origin. 

Table 13 shows the sex ratios from each pair mating involving GU males. 
Three of the seven homogamic pairs produced significantly more females than 

Table 13 
Pair matings of Guadeloupe males with females of other islands 





GU X GU 


BAXGU 


SVXGU 


GRXGU 




Pairs 


?? 


dd 


9? 


dd 


99 


dd 


99 


dd 




1 


47 


50 


20 


3 


20 





52 







2 


58 


37* 


30 


8 


119 





50 







3 


35 


46 


34 


24 


106 


5 


144 


3 




4 


68 


44* 


4 





18 


3 


42 







5 


68 


35* 


114 


51 


14 


8 


54 







6 


72 


60 


28 


3 


4 





4 







7 


30 


42 


57 


18 


65 


1 


12 







8 






26 


4 


16 


2 








9 






40 


43 


50 


1 








10 






17 


10 




•• 


•• 






Total 


378 


314 


370 


164 


412 


20 


358 


3 





* Deviation from 1:1 ratio significant in homogamic matings. 

males. This would not be emphasized if it were not for the fact that the two mass 
homogamic matings of the GU strain also produced significantly more females 
than males. The tendency toward abnormal ratios is thus already present in the 
GU strain and the effect is increased with crosses to the other island females. The 
GU females mated with other island males produced mostly normal sex ratios 
(except with ST males) in mass matings and with each pair mating. Therefore 
it does not appear as though the GU X-chromosomes carry any lethal or semi- 
lethal factors. 

A tentative conclusion from the data at hand is that the cross-lethal effect 



Heed and Krishnamurthy : Cardini Group of Drosophila 167 

involving GU males is due to one or several factors on the other island X-chromo- 
somes interacting abnormally with the GU Y-chromosome. 

Matings with BA females to males of the other islands (Table 11) show an 
interesting sex ratio cline in that more females than males are produced with 
islands far to the north of Barbados; an equal number are produced with SL 
males (slightly north of Barbados), and more males than females are produced 
at the same latitude (St. Vincent) and south of Barbados (Grenada) (Table 14). 

Table 14 
Proportion of male offspring in crosses involving females from Barbados 







Miles through 


Percent d (f from 






Degrees N. 


connecting 


all parental 




9 cf 


latitude 


islands 


crosses 




BAx ST 


18° 20" 


555 


28.7 




BAxGU 


16° 15" 


255 


20.0 




BAx SL 


13° 55" 


92 


48.5 




BAx BA 


13° 10" 





47.6 




BAx SV 


13° 15" 


100 


63.0 




BAxGR 


12° 5" 


180 


72.7 





The strains from Barbados and St. Lucia have given equal effects in sex ratio 
among themselves and with GU males. In this respect BA has closer affinities to 
SL than either SV or GR. Again it is seen that SV and GR differ from one 
another, this time in respect to BA females. The difference is highly significant 
using the totals from Table 11 in a 2 x 2 contingency table. The nature of the 
reverse in sex ratio is not yet known. 

Backcrosses of hybrid BA females to males of BA and to males of other islands 
(Table 9) show that the cline, including the reversal in sex ratio, is nullified 
when the chromosomes are predominantly of either BA or the other parental 
type. In the former case most of the crosses approximate a 1 : 1 ratio. In the latter 
case most of the crosses approximate a 2: 1 ratio. Too few hybrids were produced 
in pair matings with BA females to show the result for each pair. 

CYTOLOGY 

The number of strains used in the cytological analysis is more inclusive than 
is reported for the hybridization tests. Only the matings that produced new or 
otherwise unpredictable information on the ability of two strains to cross will be 
reported in this section along with the cytological results. 

Figures 2 and 3 show the somatic metaphase chromosomes. They may be 
grouped into five distinct haploid types. 

Type 1. Two V's, one smaller than the other, one long rod with a proximal 
satellite (X chromosome) and a dot chromosome. Common to: SK, 
GU, MA, SL, BA, SV, and GR. 
Type 2. Two V's, one smaller than the other, one long rod with proximal 
satellite (X chromosome) and a small rod. Found only in Puerto 
Rico, 



168 The University of Texas Publication 







dll 

a. ' V\w /; D. c. 




^ / ^« g. 



-f^ 



X 

I 

i 



V'Cc 




^ 



x|^ 



X 

I. m. n. 

Fig. 2. Camera lucida drawings of the neurocyte chromosomes of the dunni subgroup (a-1) 
and of belladunni (m-n). Island strains are as follows: Puerto Rico (a-c), St. Thomas (d, $ ; e, 
6 ), Guadeloupe (f), St. Kitts (g), Martinique (h), St. Lucia (i), St. Vincent (j), Grenada (k), 
Barbados (1) ; Jamaica belladunni (m, early metaphase; n, late metaphase). 

Type 3. One large V, one J (autosome), one J (X chromosome) and a small 

rod. The Y is a dark staining heterochromatic rod. Found only in 

St. Thomas. 

Type 4. Two V's, one smaller than the other, and two rods, one stains darker 

than the other. The lighter staining rod is the X chromosome. Found 

only in D. belladunni from Jamaica. 

Type 5. Two V's, one rod (X chromosome) and a dot. Common to D. acuti- 

labella from Florida, Cuba, Jamaica and Haiti. The configuration 

agrees with Stalker's description of acutilabella from St. Petersburg, 

Fla. (1953). 

The salivary chromosomes of all the strains listed above were examined. They 

showed five arms and a dot radiating from the chromocenter. One of the arms is 

lightly staining in good preparations and bears a large puff at the distal end. 

Comparison of male and female salivaries suggests that this is the X chromosome. 

The dot, which is partly incorporated in the chromocenter, represents the fourth 



Heed and Krishnamurthy : Cardini Group of Drosophila 169 

chromosome. It is the same size and carries the same number of bands in all cases 
indicating that its size difference in metaphase configurations is due to shifts m 
heterochromatic material. 

The other four strands represent the arms of the second and third metacentric 
chromosomes. All arms are distinguishable from one another by the character- 
istic appearance of the chromatin bands. 

At least eight larval salivary gland preparations were examined from the 
original stocks. No inversions were found in the intrastrain preparations except 
in the stock from Martinique which was heterozygous for inversion 2RB in the 
right arm of the second chromosome. 

At least eight larvae, when they were available, were examined from most of 
the inter-island crosses. Five different inversions were found, one on each long 
arm; they have been designated as Xe, 2LB, 2RB, 3LB complex, and 3RB. For 
discussion, the strains used in these crosses may be divided into three groups: ( 1 ) 
D. acutilabella from Jamaica, Cuba, Haiti, St. Petersburg and Floral City, Flor- 
ida; (2) D. belladunni from Jamaica; (3) the dunni subgroup from the Lesser 
Antilles and Puerto Rico. The inversion relations between them are shown dia- 
grammatically in Fig. 8. 

Initially, D. belladunni would not cross with any of the other two groups. 
However, further testing has shown that belladunni females will cross with 
males of BA, GR, GU, PR, MA and ST, the order indicating the relative ease 






^ 



a 




»• 



X 



Fig. 3. Camera lucida drawings of the neurocyte chromosomes of the strains of Drosophila 
acutilabella; Jamaica (a, anaphase), St. Petersburg, Fla. (b), Floral City, Fla. (c), Cuba (d), 
Haiti (e). 



170 



The University of Texas Publication 



with which the crosses can be made. The inversions have not yet been worked 
out. 

The various strains of acutilabella crossed well among themselves and only 
one inversion was found; whenever the Haiti strain was one of the parents the 
progeny showed a complex inversion (3LB complex) in the left arm of the third 
chromosome (Fig. 5), 

The five acutilabella strains would cross only with the Guadeloupe and St. 
Vincent strains of the dunni subgroup although they were tested with all of 
them (Table 15). The hybrid larvae examined for inversion differences were 
with the Jamaican strain of acutilabella crossed to Guadeloupe and St. Vincent. 
In both cases an inversion in the X-chromosome was found, Xe (Fig. 6) . In addi- 

Table 15 

Hybrids obtained from crosses of D. acutilabella from Jamaica (J), Hispaniola (H), Cuba (C), 

St. Petersburg, Fla. (F) and Floral City, Fla. (Fl) ; D. belladunni from Jamaica (Jb) ; 

and two strains of the dunni subgroup, Guadeloupe (GU) and St. Vincent (SV). 



i 


GU 


SV 

$0^ 


Jb 


J 


H 

9d 


c 


F 

?<3" 


Fl 


GU 









6:0 


1:0 


2:0 


3:0 


1:0 


SV 









2:0 











3:0 


Jb 


0* 





143:99 

















J 


2:1 


7:1 







. . 








H 


1:0 


2:0 











. . 


. . 


C 


2:1 


5:0 





. . 


. . 








F 





3:0 













. . 


Fl 





2:0 
















Some larvae were produced in a recent re-test; these are now being studied. 




Proximal 



Distal 

Fig. 4. Camera lucida drawing of the proximal inversion 3RB present in the progeny of dunni 
(Puerto Rico) X dunni (Guadeloupe). 



Heed and Krishnamurthy: Cardini Group of Drosophila 



171 



tion, an inversion in the second chromosome, 2LB, was present in the Jamaica- 
Guadeloupe hybrids (Fig. 7). These are interspecific crosses and pairing was 

poor. 

Crosses within the dunni subgroup showed three inversions, two of which have 
already been mentioned. The inversion 2RB, heterozygous in Martinique, ap- 
peared in some of the hybrid larvae in every cross of Martinique to other islands. 
The inversion 2LB, previously mentioned in the flcw/z7aZ?^//ff-Guadeloupe cross, 
appears in all hybrid larvae of PR, GU, MA, SL, and BA crossed to SV and GR. 
The third inversion within the dunni subgroup appears in all hybrid larvae of 
GU, MA, SL, BA and SV crossed to PR and ST. This inversion is 3RB (Fig. 4). 

Table 16 hsts all the inversions actually observed except the complex inversion 
3LB, homozygous in the Haiti strain of acutilabella. The degree of pairing of 
homologous arms is also indicated. 

The inversion differences of the inter-island crosses (Fig. 8) divide the dunni 
subgroup into three distinct units: a northern unit (PR and ST), a middle unit 
(GU, MA, SL and BA) and a southern unit (SV and GR). This agrees with the 
genetic tests in that PR and ST are interfertile and SV and GR are interf ertile. 
The fertility of the GU X SV hybrids is an exception to this and shows an unex- 
pected relationship between the two islands. Further confirmation of this rela- 
tionship is the fact that the strains from these two islands are the only ones in the 
Lesser Antilles that give at least a few hybrids with the five strains of acutilabella. 
Figure 8 illustrates the inter-island relationships by the inversions found in the 
hybrids. 



^■f Distal 




Proximal 



Fig. 5. Camera lucida drawing of the complex inversion 3LB present in the progeny of 
acutilabella (Haiti) X acutilabella (Jamaica). 



172 



The University of Texas Publication 




Fig. 6. Camera lucida drawing of the salivary chromosomes of a female hybrid from the cross 
dunni (St. Vincent) X acutilabella (Jamaica). The inversion Xe is shown and the numerous 
regions of poor pairing (arrows) . 



DISCUSSION AND SUMMARY 

A glance at Table 9 showing the results of the backcrosses in the dunni sub- 
group indicates that in the majority of the cases introgressive hybridization be- 
tween island populations is theoretically possible. Direct fertile hybridization is 
possible between the two widely separated islands of St. Vincent and Guade- 
loupe, as well as the adjacent islands of Puerto Rico and St. Thomas, and of 
Grenada and St. Vincent. 

However the genetic differences between the island populations are very 
noticeable. Most of the inter-island F^ hybrids show a variable phenotype indi- 
cating the original types are heterozygous for factors controlling the abdominal 
pattern. This heterozygosity does not seem to depend on inter-island migration 
since each of the three main inversion groups (PR, ST; GU, MA, SV, BA; SV, 
GR) are homozygous for their own inversions. As far as our samples are ade- 



Heed and Krishnamurthy: Cardini Group of Drosophila 



173 



quate (each island stock is derived from many females) this indicates a distinct 
isolation at least between the three main groups. 

There is of course the possibility that selection acting on the segregates from 
the hybrids is favoring the inversion already established on the island that is 
receiving the migrants. The Fe hybrids of SV X GU are phenotypically very con- 
stant and similar to the Guadeloupe parent. The fixation to one parental type m 
later hybrid generations (here only inbreeding and not backcrossing was al- 
lowed) supports Stebbins' (1959) recent statement that in animals selection 
after hybridization would be expected to act upon a specific adaptive type rather 
than any variety of types (hybrid swarm). Identification of the hybrid chromo- 
somes is now in progress. 

The increase in severity of abnormal sex ratios with distance in two series of 
inter-island crosses shows that differently integrated gene systems are being 
formed characteristic of each island population. Most impressive is the fact that 
St. Vincent and Grenada, although phenotypically identical, behave significantly 
different in crosses to males of Guadeloupe in the one case and to females of 
Barbados in the other. For other well documented cases of cross-lethals and semi- 
lethals see Crow (1942) and Patterson and Griffen (1944) . 




-:-J W Chromosome 



Fig. 7. Camera lucida drawing of the left arm of the II chromosome with the basal inversion 
2LB present in the progeny of dunni (Guadeloupe) X acutilabella (Jamaica). 



1 74 The University of Texas Publication 

Table 16 

Inversion differences and degree of pairing in the salivary chromosomes found in hybrids of 
inter-island crosses in the dunni subgroup and with D. acutilabella from Jamaica (J) 





PR 


ST 


GU 


MA 


SL 


BA 


SV 


GR 


PR 




none 
good 


3RB 

good 


3RB 

2RB-some 

poor 


3RB 

poor 


3RB 

poor 


3RB 
2LB 

poor 




ST 






3RB 

good 


3RB 

2RB-some 

poor 




3RB 

good 






GU 






.... 


2RB-some 
good 


none 
good 


none 
good 


2LB 

good 




MA .... .... .... ..;. 


SL 








2RB-few 

good 




none 
good 




2LB 

good 


BA 


.... 


.... 


.... 


2RB 

good 




.... 


2LB 
good 




SV 








2LB 

2RB-few 

good 








none 
good 


GR 














none 
good 




J 






Xe 

2LB 

poor 








Xe 
poor 





The cytological picture in the dunni subgroup is rather conservative in com- 
parison to the genetic differentiation. Martinique has the only population carry- 
ing an inversion in the heterozygous condition. This inversion is restricted to 
Martinique. The other two inversions, 3RB and 2LB, appear only in hybrids in 
crosses of Puerto Rico and St. Thomas to all other islands, and St. Vincent and 
Grenada to all other islands. 

The most noticeable characteristic of the dunni subgroup is the cline in ab- 
dominal pattern. Several types of clines have been reported in Drosophila. They 
have ranged from the temperature viability clines in D. funebris (Timofeeff- 
Ressovsky, 1933) through morphological gradients in D. robusta (Stalker and 
Carson, 1947) to clines in inversion frequencies in D. pseudoobscura (Dob- 
zhansky, 1944), D. robusta (Carson and Stalker, 1947) and others. It is gen- 
erally agreed that clines are the result of adaptation of local populations either 
directly or indirectly to environmental differences throughout the range of the 
organism. For Drosophila and many other organisms these environmental differ- 
ences are not usually obvious because we must think and measure in terms of 
the microenvironment of the organism. This is a field that deserves intense 
investigation. 

In the tropical lowlands the majority of Drosophila species are yellow. In the 



Heed and Krishnamurthy : Cardini Group of Drosophila 



175 



tropical highlands there is a much higher frequency of dark species. Very few 
Drosophila species in the tropics have a continuous range from sea level to even 
five or six thousand feet (Heed, 1957b). Darker insects at higher altitudes are 
thought to be more protected from ultra violet radiations (Kalmus, 1941a). Also 
it has been demonstrated that darker Drosophila are more resistant to desiccation 
(Kalmus, 1941b). Even with this knowledge it would be mere speculation to 
assign a reason for the tremendous increase in black pigment in the dunni sub- 
group in the more southern parts of its range. 

An alternative explanation for the chne in the dunni subgroup is the possi- 
bility that it is due to chance. This does not seem to be the case because of the 
regularity of the chne. The genetic make-up of the original founder population 
on each island was a chance phenomenon but there must be at present a constant 
and step-wise different selection pressure acting on each island population. For 
an excellent theoretical discussion of founder populations on peripheral islands 
seeMayr (1954). 

There are three types of clines exhibited in the cardini group m the West 
Indies. The first two clines, abdominal pattern and sex ratio, are restricted to the 
dunni subgroup. The third type, chromosomal, must include D. belladunni in 
Jamaica in order to estabhsh this as a true cline. The three clines are independent 
of one another although the first two follow mostly the same path. The dunni 
subgroup forms a sequence from north to south of light to dark abdomens but this 



2RB/+ 




Fig. 8. Inversion differences among inter-island hybrids in the dunni subgroup and D. acutila- 
bella. The Martinique strain is heterozygous for inversion 2RB. The island strains are: Grenada 
(GR), St. Vincent (SV), Barbados (BA), St. Lucia (SL), Martinique (MA), Guadeloupe (GU), 
St. Thomas (ST), Puerto Rico (PR). 



176 



The University of Texas Publication 



Table 1 7 
Summary of major Drosophila collecting localities in the West Indies 



Collecting dates 



General localities 



Dec. 31-Jan. 5, 1956 


Barbados 


July 29, 1957 


Barbados 


Jan. 13-19, 1956 


St. Lucia 


Jan. 20-23, 1956 


St. Kitts 


July 24^28, 1957 


St. Vincent 


July 20-23, 1957 


Grenada 


July 31 -Aug. 1, 1957 


Martinique 


Aug. 3-8, 1957 


Guadeloupe 


Aug. 11-13,1957 


St. Thomas 


Jan.30-Feb. 7, 1956 


Puerto Rico 


Aug. 16-19, 1957 


Puerto Rico 


Oct. 11-Nov. 1, 1957 


Puerto Rico 


Feb. 11-18, 1956 


Haiti 


Feb. 21-29, 1956 


Jamaica 


July 1-4, 1957 


Jamaica 


Nov. 5-14, 1957 


Jamaica 


July 19-27, 1958 


Jamaica 



Turner's Hall Wood, Blower's Wood, 
Grants Castle. 

Monkey Hill. 

Castries, Barre de File, Pigeon Island. 

Basseterre, Mt. Misery, Monkey Hill. 

Kingstown, Blue Lagoon, Soufriere 
Mountains. 

St. George, Grand Etang. 

Forte de France, Morne Rouge. 

Matouba, Volcan Soufriere. 

Signal Hill. 

Carib. Natl. Forest at El Yunque and 
Sabana, USDA Exper. Sta. at Rio 
Piedras. 

USDA Exper. Sta. at Mayaguez. 

Mayaguez, Ensenada, Cedra, Maricao, 
El Yunque. 

Kenscoff, Furcy, Petionville. 

Bath, Hermitage Reservoir. 

Montego Bay, Windsor. 

Kingston, Hardware Gap, Mona, Bath. 

Windsor, Ocho Rios, Mt. Diablo, Hard- 
ware Gap, Hermitage Reservoir. 



is not a perfect sequence. Barbados has the darkest form but is not the most 
southern island. Martinique is north of St. Lucia but it has the darker type. 

The sex ratio chne with Guadeloupe males has the sequence: GU>SL>BA 
>SV>GR. This puts the Barbados form in the peculiar position of being the 
more closely related to the form from St. Lucia than the darker types from St. 
Vincent and Grenada. The inversion story also bears this out, since GU, SL and 
BA hybrids are all homozygous. Barbados was probably colonized from St. Lucia. 

The chromosomal cline involves the addition (or subtraction) of heterochro- 
matin on the dot chromosome. The metaphase of all island types south of St. 
Thomas show the usual dot chromosome. The Puerto Rico and St. Thomas forms 
have this chromosome enlarged and D. belladunni in Jamaica owns a hetero- 
chromatic rod in place of the dot. D. belladunni has definite affinities with the 
dunni subgroup (see species description) and thus the cline is probably valid in 
the sense that belladunni did not acquire the extra heterochromatin totally inde- 
pendently (or that the Puerto Rico and St. Thomas forms represent a reduction 
in heterochromatin). 

The three clines represent genetic divergence in close relation to the geo- 
graphic position of the islands. According to Mayr (1942) the presence of clines 
indicates continuities between adjacent populations. He was talking mainly 
about mainland populations, however. Our data show discontinuities and di- 
vergences yet they are still in a regular sequence or cline. The introgressive possi- 



Heed and Krishnamurthy: Cardini Group of Drosophila 1 77 

bilities and the clines in the dunni subgroup probably indicate very recent habi- 
tation of Puerto Rico and the Lesser Antilles instead of present day continuities. 
Barbados gives us a lower time limit for the establishment of its very dark popu- 
lation. Beard (1949) dates Barbados' recent uphft as post- Pleistocene. The habi- 
tation of Barbados was probably much more recent. 

If the colonization of the Lesser Antilles by the dunni subgroup is indeed very 
recent then the affinities of its members are with the Greater Antilles rather than 
South America since the closest relatives are D. acutilabella and D. belladunni. 

D. acutilabella should be taken as the standard salivary chromosome type in 
the cardini group since it crosses more readily with all other members of the 
group. Stalker (1953) reports that acutilabella from Florida will give sterile hy- 
brids with cardini, polymorpha and parthenogenetica and fertile hybrids with 
cardinoides. The present work shows that acutilabella from its entire range will 
give sterile hybrids with the dunni subgroup strains from St. Vincent and Guade- 
loupe. If acutilabella is taken as the standard (Figure 8) there is no other choice 
than to follow the inversion steps from Jamaica to Grenada-St. Vincent and 
proceed north to Puerto Rico. The logical conclusion to this migration pattern 
would be to terminate again in Jamaica with D. belladunni. 

However, there are two reasons for believing that this hypothesis may not be 
correct. The migration from Jamaica to the southern islands of the Lesser An- 
tilles seems to be far more improbable than through Hispaniola to Puerto Rico 
and south. Also we believe that belladunni is the older form on Jamaica (see 
species description), inhabiting that island before acutilabella. The establish- 
ment of the dunni subgroup from the Greater Antilles at this point appears more 
likely than from South America but the dispersal route by which the Lesser 
Antilles were colonized is in doubt. For a discussion of migration patterns and 
chance dispersal of D. willistoni in the West Indies see Dobzhansky (1957). 

The relation between acutilabella and belladunni is complex. They are sym- 
patic in Jamaica and should be treated as sibling species since it is usually im- 
possible to separate females in the field. Both species have affinities with the 
dunni subgroup. The arrangement of palpal bristles, an excellent distinguishing 
character in the cardini group, and the male genitalia of belladunni are similar 
to the dunni subgroup. Tests now in progress indicate that female belladunni will 
produce at least a few hybrids with males of most of the members of the dunni 
subgroup. D. acutilabella is not as morphologically similar to the subgroup but 
has produced sterile hybrids with the members from St. Vincent and Guadeloupe. 

D. acutilabella may also be considered a sibling species of D. cardini in Florida. 
From the notes of a collecting trip with Dr. Marshall R. Wheeler in 1953, we 
agreed that acutilabella and cardini females could not be safely distinguished 
(see also Stalker, 1953). There is at the same time an obvious difference between 
cardini and belladunni. It seems as though acutilabella is quite plastic, "imitat- 
ing" cardini in Florida and belladunni in Jamaica. The fact that acutilabella is 
also polymorphic adds to the confusion but does not invalidate the above observa- 
tions. D. cardini is also found in Jamaica but is rarer than the other two species. 
This relationship deserves further investigation. 



1 7S The University of Texas Publication 

ACKNOWLEDGMENTS 

The authors wish to express their appreciation to Prof. J. W. Boyes of McGill 
University, Montreal, and to Dr. Marvin Wasserman, University of Texas, for 
their help in collecting the material, and to Dr. H. E. Warmke and Dr. George 
Wolcott of the USDA Experiment Stations at Mayaguez and Rio Piedras, Puerto 
Rico, for their kind assistance. 

The collecting trips to the Caribbean region were financed by the National 
Science Foundation (Grant NSF-G-1653 to W. S. Stone and M. R. Wheeler, and 
Grant NSF-G-4999 to M. R. Wheeler) while the Rockefeller Foundation grant 
to the Genetics Foundation of the University of Texas has taken care of the major 
laboratory expense. A National Science Foundation grant (NSF-G6235) to 
William B. Heed has supported the work at the University of Arizona. 

BIBLIOGRAPHY 

Beard, J. S. 1949. The natural vegetation of the windward and leeward islands. Oxford at the 

Clarendon Press. 
Carson, H. L. and H. D. Stalker. 1947. Gene arrangements in natural populations of Drosophila 

robusta Sturtevant. Evolution 1:113-133. 
Crow, J. F. 1942. Cross fertility and isolating mechanisms in the Drosophila mulleri group. 

Univ. Texas Publ. 4228:53-67. 
Cunha, A. B. da. 1949. Genetic analysis of the polymorphism of color pattern in Drosophila 

polymorpha. Evolution 3:239-251. 
Dobzhansky, T. 1944. Chromosomal races in Drosophila pseudoobscura and Drosophila persimilis. 

Carnegie Inst. Wash. Publ. 554:47-144^. 
. 1957. Genetics of natural populations XXVI: Chromosomal variability in island 

and continental populations of Drosophila willistoni from Central America and the West 

Indies. Evolution 11:280-293. 
Heed, W. B. 1957a. A preliminary note on the cardini group of Drosophila in the Lesser Antilles. 

Univ. Texas Publ. 5721:123-124. 
. 1957b. Ecological and distributional notes on the Drosophilidae (Diptera) of El 

Salvador. Univ. Texas Publ. 5721:62-78. 
and M. R. Wheeler. 1957. Thirteen new species in the genus Drosophila from the 



Neotropical Region. Univ. Texas Publ. 5721: 17-38. 
Kalmus, H. 1941a. Physiology and ecology of cuticle colour in insects. Nature 148:428-431. 
. 1941b. Resistance to desiccation of Drosophila mutants affecting body colour. Proc. 

Roy. Soc. London (B) 130:185-201. 
Malogolowkin, C. 1958. Maternally inherited "sex-ratio" conditions in Drosophila willistoni and 

Drosophila paulistorum. Genetics 43:274-286. 
Mayr, E. 1942. Systematics and the origin of species. Columbia Univ. Press, N. Y. 
. 1954. Change of genetic environment and evolution. In Evolution as a process, ed. 

J. Huxley, A. C. Hardy and E. B. Ford. London, Allen and Unwin Ltd. 
Patterson, J. T. and A. B. Griff en. 1944. A genetic mechanism underlying species isolation. Univ. 

Texas Publ. 4445:212-223. 
Stalker, H. D. 1953. Taxonomy and hybridization in the cardini group of Drosophila. Ann. Ent. 

Soc. Amer. 46:343-358. 
and H. L. Carson. 1947. Morphological variation in natural populations of Drosophila 

robusta Sturtevant. Evolution 1:237-248. 



Heed and Krishnamurthy: Cardini Group of Drosophila 179 

Stebbins, G. L. 1959. Proc. Amer. Phil. Soc. 103 (No. 2):231-251. 

Sturtevant, A. H. and T. Dobzhansky. 1936. Geographical distribution and cytology of "sex- 
ratio" in Drosophila pseudoobscura and related species. Genetics 21 : 473-490. 

Timofeeff-Ressovsky, N. W. 1933. Uber die relative Vitalitat von Drosophila melanogaster 
Meigen und Drosophila funebris Fabricius unter verschiedenen Zuchtbedingungen, in 
Zusammenhang mit den Verbreitungsarealen dieser Arten. Arch. Naturgesch. N. F. 2: 
285-290. 

Townsend, J. I. and M. R. Wheeler. 1955. Notes on Puerto Rican Drosophilidae including 
descriptions of two new species of Drosophila. Jour. Agric. Univ. Puerto Rico 39:57-64. 



1 



I 



A Nomenclatural Study of the Genus Drosophila 

MARSHALL R. WHEELER 
Department of Zoology, The University of Texas 

About 1934 Dr. Patterson began studying the local species of Drosophila with 
a view to finding additional species which might serve as experimental animals 
in studies of comparative genetics. By 1938 it was apparent that there were many 
more species in the Austin area than had been supposed, and that many of them 
were undescribed. With the aid of Prof. A. H. Sturtevant the species were studied 
taxonomically as well as genetically, and a new phase of genetics research at the 
University of Texas began. 

In those early days various graduate students assisted Dr. Patterson in the 
field collecting, for example Dean Parker, John Carpenter, Gordon Mainland, 
Robert Wagner, William Baker and myself. The taxonomic results of these col- 
lections were summarized in 1942 in the University of Texas Publication 4213 
in which 46 new species were described. A resurgence of interest in Drosophila 
systematics followed this publication, as is shown from the data presented in 
Figure 1 — a resurgence which has not yet lost its momentum. It is clearly evident 
that Dr. Patterson's influence in this field has been tremendous, and I feel that it 
is a distinct privilege to contribute this article to a volume honoring the man who 
was not only responsible to a considerable degree for the healthy development of 
modern Drosophila systematics. but who was also largely responsible for my own 
interest in it. 

THE GENUS DROSOPHILA 

The famous dipterist, C. F. Fallen, established the genus Drosophila in 1823 
for Musca funehris Fabricius and the following eleven new Swedish species: 
curvipennis, variegata, fenestrarum, transversa, obscura, tristis, fuscula, cine- 
rella, flava, gramium, and glabra. In addition to funebris, which was designated 
as the type species by Zetterstedt in 1847,* only five of Fallen's species are still 
considered as belonging to Drosophila: fenestrarum, transversa, obscura, tristis, 
and flava. 

In the years immediately following Fallen's description of the genus a few new 
species were added sporadically but many of them were, in turn, relegated to 
other genera. In later years, however, and in spite of the narrowing concept of 
the generic limits, the growth of the genus has been remarkable. Figure 1 illus- 
trates the number of species described in the genus by decades since 1823, as well 
as the number still considered to be valid members of the genus (the difference 
between the numbers in each sector) . Relatively few species have been placed in 
Drosophila after being described in other genera; these species have not been 
tabulated. 



* The earlier designation of Drosophila cellaris as the type species, by Curtis in 1833 and by 
Westwood in 1840, was invalid since cellaris was not an originally included species. 



182 



The University of Texas Publication 



1940- 



1950-58 



1930-39, 



1920-29 




1820-29 
1830-39 

1840-49 
1850-59 



1860-69 

1870-79 
1880-89 

1890-99 



1910-19 



1900-09 



Fig. 1. Diagram illustrating the number of species described in Drosophila by decades since 
1823. Numbers within the sectors represent the number of described species less the number no 
longer considered valid members of the genus by reason of generic transfer or synonymy. 

At the time of writing, 920 species have been described in the genus Dro- 
sophila, about 85% of them since 1900. The spectacular peak of 179 species 
named between 1920 and 1929 represents, primarily, the efforts of three men, O. 
Duda, J. R. Malloch, and A. H. Sturtevant. Nearly 40% of the known species 
have been described since 1940, thus reflecting the renewed interest of many 
workers in many lands who are engaged in analyzing the multitude of species 
which inhabit the earth. 

New Names Proposed for Junior Homonyms 
In assembling the catalogue list which follows, instances of both primary and 
secondary homonymy have been found. The Rules of Nomenclature require that 
junior primary homonyms are to be corrected whenever discovered while junior 
secondary homonyms need substitute names only if a true biological homonymy 
is considered to exist. I am therefore proposing new names for all junior primary 
homonyms in the genus known to me, but action regarding most secondary homo- 
nyms is being postponed pending further biological information. Since names 
proposed for "varieties" will ultimately be ruled equivalent to subspecific names, 
nomenclaturally, then another series of primary homonyms will exist; these can 
be corrected when the need arises. 



Wheeler: Nomenclature of Drosophila 183 

Drosophila (Hirtodrosophila) unicolorata Wheeler, nom. no?;. 

For Hirtodrosophila unicolor Malloch 1934 (Ins. Samoa, Pt. 6:293); = Dro- 
sophila unicolor, Patterson and Wheeler 1949, Harrison 1954. 
Not Drosophila unicolor de Meijere 1914 (Tijd. Ent. 57:266) . 
The two species concerned are clearly referable to the genus Drosophila; I am 
therefore rejecting the junior name, unicolor Malloch, and proposing the substi- 
tute name, unicolorata. Malloch stated that although Hirtodrosophila was pro- 
posed as a subgenus of Drosophila, he felt that it was worthy of generic rank. 
Recent workers, however, consider it to be a valid subgenus. 

Drosophila tendata Wheeler, nom. nov. 

For Drosophila dentata Buda 1927 (Arch. Naturg. 91A12 (1925):145; 201). 

Not Drosophila longecrinita var. dentata Duda 1924 (Arch. Naturg. 90A3: 

205; 242) ; = Drosophila dentata, Duda 1926 (Suppl. Ent. 14:65). 

Duda (1924) described dentata as a variety of longecrinita, in the subgenus 
Hirtodrosophila, but elevated it to specific rank in 1926. He created a primary 
homonym by using the name dentata a second time for a wholly different species 
from South America. 

Drosophila peruensis Wheeler, nom, nov. 

For Drosophila maculipennis Duda 1927 (Arch. Naturg. 91A12 (1925) : 167). 

Not Drosophila maculipennis Gimmerthal 1847 (Bull. Soc. imper. nat. Moscou 

20: 199) ; = Diastata nebulosa Fallen (Diastatidae) . 

Although maculipennis Gimmerthal has been removed to the Diastatidae, the 
name is preoccupied in Drosophila; since the species described by Duda came 
from Peru, I am using the name peruensis to replace his junior primary 
homonym. - 

Drosophila meijerei Wheeler, nom. nov. 

For Drosophila nigricolor de Meijere 1911 (Tijd. Ent. 54:399), a new name 

proposed for Drosophila nigra de Meijere, not Grimshaw. 

^ot Drosophila nigricolor Strobl 1898 (1897; Mitt. Nat. f. Steiermark 34:266). 

It is clear that de Meijere was not aware of the prior use of nigricolor when he 
proposed it as a substitute for his nigra, preoccupied by nigra Grimshaw. It 
seems appropriate to propose a new name honoring this eminent dipterist. Al- 
though the evidence is scanty, Drosophila meijerei probably belongs to the sub- 
genus Pholadoris. 

Drosophila neochracea Wheeler, nom. nov. 

For Drosophila ochracea Duda 1927 (Arch. Naturg. 91A12 (1925): 195). 

Not Drosophila ochracea Grimshaw 1901 (Fauna Haw. 3:61). 

A new name is needed to replace this junior primary homonym; to retain the 
general color significance of the name, the modification neochracea is proposed. 

Drosophila adamsi Wheeler, nom. nov. 

For Drosophila quadrimaculata Adams 1905 (Kans. Univ. Sci. Bull. 3:182). 
Not Drosophila quadrimaculata Walker 1856 (Ins. Saund. 1, Dipt.:410); 
= Leucophenga varia (Walker) 1849 as Drosophila varia Walker. 



1 84 The University of Texas Publication 

Primary homonymy exists here in spite of the fact that Walker's species has 
been removed to another genus and is a synonym. 

Leucophenga ornata Wheeler, nom. nov. 

For Drosophila ornatipennis de Meijere 1914 (Tijd. Ent. 57:256); 
= Leucophenga ornatipennis, Sturtevant 1921, Duda 1924. 
Not Drosophila ornatipennis Williston 1896 (Trans. Ent. Soc. London, 1896, 
Pt. 3:407). 

In naming his species de Meijere wrote ''Drosophila (Leucophenga) ornati- 
pennis'', since he considered that breaking up Drosophila into many genera was 
poor taxonomy. He indicated by parentheses, how^ever, those species which 
would belong in Leucophenga if it were recognized. As published, he created a 
junior primary homonym which requires a new name. 

Microdrosophila zetterstedti Wheeler, nom, nov. 

For Drosophila nigriventris Zetterstedt 1847 (Dipt. Scand. 6:2557); 
= Drosophila (Incisurifrons) nigriventris, Duda 1934; = Microdrosophila 
nigriventris, Hackman 1954. 

Not Drosophila nigriventris Macquart 1843 (1842. Mem. Soc. Sci. Arts 
IjlleAXQ) ;— Drosophila (Leucophenga) nigriventris, de Meijere 1908; 
= Leucophenga nigriventris, Sturtevant 1921, Duda 1924. 

These two nominal species were originally described in the same genus, and 
Zetterstedt's species, being junior, requires a new name. 

Drosophila multistriata Duda 1923 (Ann. Mus. Nat. Hung. 20:57). 

Equals Drosophila lineata (de Meijere) 1911, as Stegana lineata (Tijd. Ent. 

54:420). 

Not Drosophila lineata van der Wulp 1886 (Dipt. Midd.-Sumatra Exped. 

iv(9):57). 

Duda (1924) transferred Stegana lineata de Meijere to Drosophila, where it 
became a junior homonym of lineata van der Wulp. There is an available re- 
placement name, however, since Duda has stated that his multistriata is the same 
as lineata de Meijere. If it should be shown later that this synonymy is in error, 
then de Meijere's species will require a new name. 

Drosophila punctatonervosa Frey 1954 (Norw. Sci. Exped. Tristan da Cunha 

26:32). 

Equals Drosophila poeciloptera Duda 1940 (Ann. Mus. Nat. Hung. 33:26). 
Not Drosophila poeciloptera (Duda) 1925. as Paramycodrosophila poecilop- 
tera (Ann. Mus. Nat. Hung. 22:226), which equals Drosophila poecila Burla 
and Pavan 1953 (Rev. Brasil. Biol. 13:311). 

Upon transferring Paramycodrosophila poeciloptera Duda 1925 to the genus 
Drosophila, and thus producing homonymy, Burla and Pavan (1953) unfor- 
tunately proposed a new name (poecila) for the senior homonym {poeciloptera 
Duda 1925) rather than the junior one {poeciloptera Duda 1940). Drosophila 
poecila is thus an absolute synonym of poeciloptera Duda 1925, and the junior 
name of 1940 still requires a substitute. 



Wheeler: Nomenclature of Drosophila 185 

There is an available replacement name: punctatonervosa Frey 1954, which 
judging from the description, is a later name for the same species. If this 
synonymy, however, should later prove to be untrue, then poeciloptera Duda 
1940 will require a new name. 

Secondary Homonyms of Uncertain Validity 
There are three instances of uncorrected secondary homonymy in the cata- 
logue list which follows. In each case there is uncertainty regarding the biological 
reality of the homonymy. The pertinent facts are given below. 

1. Drosophila hypopygialis. Duda described a Tanygastrella hypopygialis 
and later relegated Tanygastrella to the status of a subgenus of Drosophila. That 
specific name has also been used by Malloch in Drosophila. Until the position of 
Tanygastrella in Drosophila is verified, however, I am not willing to propose a 
new name for the junior homonym. 

2. Drosophila metallescens. Malloch described a species with this specific 
name in Spinulophila which he considered to be a genus but which was origi- 
nally proposed as a subgenus of Drosophila. Recent authors consider it to be ap- 
proximately equivalent to the immigrans species group of the subgenus Dro- 
sophila. Malloch's species is, therefore, clearly a Drosophila, and the name is a 
junior homonym of metallescens de Meijere. I consider, however, that Malloch's 
species is a probable synonym of monochaeta Sturtevant; hence a new name is 
not required for it unless, and until, it should be shown to be a distinct species. 

3. Drosophila obscuricornis. Duda transferred Stegana obscuricornis de Mei- 
jere into Drosophila where it becomes a junior homonym. Without a re-examina- 
tion of the type of de Meijere's species, however, I beheve that its position in 
Drosophila is to be questioned. If it is shown to be a true Drosophila, it should 
then be given a new name. 

Annotated List of Species, Subspecies and Varieties 
No one can deny that Drosophila is a very large genus. In the list which fol- 
lows, about 750 presumably valid species are enumerated. In view of the fact that 
collecting has been relatively sparse in many areas of the world, it wdll not be 
surprising if the actual number of species reaches 1500. It will be even greater if 
the phenomenon of nearly identical sibling-species is as widespread as it now 
appears to be. It is for this reason, primarily, that this annotated list of species, 
subspecies and varietal names has been assembled. More than 1000 names have 
already been used in the genus, so that the creation of new names for the multi- 
tude of undescribed species is becoming a difficult task. To assist in the avoidance 
of homonymy the hst has been arranged to show which names are unavailable 
for any other species or subspecies in the genus, and which names might be used 
again for some other species. Confusion can best be prevented, however, by 
avoiding the use of any of the names in the list for any future species or sub- 
species of Drosophila. 

In preparing the list I have made extensive use of the earlier catalogue of 
Sturtevant (1921) and of the Hst of vahd names published by Patterson and 
Wheeler (1949). I have attempted to bring these lists up to date and correct the 



186 The University of Texas Publication 

errors as far as possible. The manner of listing the five major categories of names 
is as follows: 

1. Species originally described in the genus Drosophila. The name is in bold- 
face type followed by the author and date without intervening punctuation. 
These names may not be used again in the genus. 

2. Species transferred to the genus Drosophila. The name is in bold-face, but 
with the author's name in parentheses, followed by the original generic refer- 
ence. If the species is still considered to be a Drosophila, its name is not available 
for any other species. A number of species, having been transferred into Dro- 
sophila, were later removed from it again; only in special cases are they included 
in the list. Names of such temporary members of the genus are available for 
future use. 

3. Names of subspecies. According to the Rules names of subspecies have 
equal standing with names of species as regards priority in homonymy. These 
names are in bold-face, with a statement regarding their status as subspecies. 
They may not be used again in the genus. 

4. Names originally proposed as "varieties.'' There are 27 such names in the 
list; they are in italics, with a notation as to their varietal status. It seems prob- 
able that the new version of the Rules, which is to be published in the near 
future, will place varietal names on an equal status with those of subspecies, 
nomenclaturally; they will then be subject to the same rules governing homo- 
nymy as those now applicable to specific and subspecific names. It would seem 
prudent for Drosophila taxonomists to avoid the re-use of these varietal names 
for any new forms in the future. 

5. Names without status in nomenclature. Also shown in italics are certain 
manuscript names, nomina nuda, invalid spellings, etc., which do not preoccupy 
for purposes of homonymy. There are several hundred such names in the litera- 
ture; a few have been included in the list when, in the writer's opinion, they 
might have some bearing on nomenclatural problems. 

The following abbreviations are employed: species (sp.), subspecies (subsp.), 
variety (var.), synonym (syn.), new name (n. n.), probable (prob.), possible 
(poss.), preoccupied, i.e., a homonym (preocc), considered (consid.), erroneous 
subsequent spelling (err. subseq. spelling), nomen nudum (nom. nud.), species 
incerta (sp. inc.), personal communication (pers. comm.). 

ANNOTATED LIST OF NAMES IN THE GENUS DROSOPHILA 

abbreviata de Meijere 1911:400. To Leuco- acuta Sturtevant 1927:370. 

phenga. acvitilabella Stalker 1953:345. 

aberrans Lamb 1914:334. Type of subg. Di- aciitissima Okada 1956:139. 

chaetophora. adamsi Wheeler 1959, antea. N. n. for quadri- 
abregolineata Duda 1925:214. maculata Adams, not Walker. 

abron Burla 1954a: 170. addisoni Pavan 1950:4. 

abureBurla 1954a: 168. adspersa Mik 1886:328. Syn. of repleta Wol- 
acanthoptera Wheeler 1949b: 171. Type of laston. 

subg. Sordophila. adusta Loew 1862:231. To Parascaptomyza. 

aceti Kollar 1851:205-6. Prob. syn. of fune- adyukru Burla 1954a: 138. 

bris (Fabricius) . afer Tan, Hsu, & Sheng 1949: 200. 

acuminata Collin 1952:199. affinis Sturtevant 1916:334. 



Wheeler: Nomenclature of Drosophila 



187 



agamse Burla 1954a: 129. 

agbo Burla 1954a: 104. 

akabo Burla 1954a: 109. 

akai Burla 1954a: 173. 

akaju Burla 1954a: 155. 

alabamensis Sturtevant 1918b: 38. 

alafumosa Patterson & Mainland 1943:187. 

alagitans Patterson & Mainland 1943:194. 

albescens Frota-Pessoa 1954:281. 

albicans Frota-Pessoa 1954:282. Poss. syn. of 

albirostris Sturtevant. 
albiceps de Meijere 1914:258. To Leuco- 

phenga. 
albicincta de Meijere 1908:156. To Leuco- 

phenga. 
albicornis de Meijere 1915a: 58. (the invalid 
original spelling, abicornis, was emended by 
de Meijere). To Mycodrosophila. 
albifrontata Malloch 1934a: 301. 
albilabris Zetterstedt 1860:6425. (as first pub- 
lished, was attributed to "Roth, in litteris'). 
To Amiota. 
albincisa de Meijere 1911:409. 
albipes Walker 1 852: 41 0. Sp. inc. 
albirostris Sturtevant 1921:78. 
albofasciata Macquart 1851:277. To Leuco- 

phenga. 
alboguttata Wahlberg 1838:22. To Amiota. 
albolimbata Duda 1924a:216, & 1924b:256. 
albomarginata Duda 1927:173. 
albomicans Duda 1924a: 209. & 1924b: 245. 
Nom. nud. in Duda 1923:47. Syn. of rmsuta 
Lamb, 
albonotata de Meijere 1911:408. 
albopunctata Becker 1900:64. To Chymomv- 

za. 
alboralis Momma & Takada 1954:98. 
albostriata Malloch 1924b: 352. 
albovittata Duda 1926a: 83, 87. N. n. for sul- 
furigaster Duda, as an invalid replacement; 
syn of nasuta Lamb. 
aldrichi Patterson & Crow 1940:25L 
alexandrei Cordeiro 1951:1. 
alfari Sturtevant 1921:75. 
algonqviin Sturtevant & Dobzhansky 1936: 

575. 
alladian Burla 1954a: 175. 
alpina Burla 1948:274. 
alternata de Meijere 1911:402. 
alternolineata Duda 1925:213. 
altiplanica Brncic & Santibanez 1957:69. 
amabalis de Meijere 1911:405. To Mycodro- 
sophila. 
ambigua Pomini 1940:157. 
americana Spencer 1938:169, as subsp. of vi- 
rilis; now consid. sp. 



amoena Loew 1862:230. To Chymomyza. 
ampelophila Loew 1862:231. Syn. of melano- 

gaster Meigen. 
amplipennis Malloch 1934b: 442. 
analis Macquart 1843:415. Sp. inc. 
ananassae Doleschall 1858:128. 
ancep Patterson & Mainland 1944:39. 
andalusiaca Strobl 1906:372. 
andina Dobzhansky & Pavan 1943:59. 
angularis Okada 1956: 128. 
angusta de Meijere 191 5a: 57. 
angustibucca Duda 1925:218. 
angustipennis de Meijere 1911:413. To Dias- 

tatidae. 
annularis Sturtevant 1916:327. N. n. for an- 

nulata Williston, not Fallen. 
annulata (Fallen 1813:250, in Notiphila). To 
Drosophila by Zetterstedt in 1847; later 
moved to Periscelidae. 
annulata Williston 1896:409. Preocc; n. n. is 

annularis Sturtevant. 
annulimana Duda 1927:117. 
annulipes Duda 1924a:209. 221, & 1924b: 

250. Nom. nud. in Duda 1923:58. 
anomalipes Grimshaw 1901:62. 
anuda Curran 1936:43. 
anyi Burla 1954a: 145. 
apectinata Duda 1931:194. 
apicata Thomson 1869:596. To Scaptomyza. 
apicifera Adams 1905:185. To Leucophenga. 
appendiculata Malloch 1934b: 441. Type of 

subg. Chusqueophila. 
approximata Zetterstedt 1847:2557. Syn. of 

fasciata Meigen (Duda 1935a). 
aracea Heed & Wheeler 1957:36. 
araicas Pavan & Nacrur 1950:264. 
arapuan da Cunha & L Pavan 1947:36. 
ararama Pavan & da Cunha 1947:28. 
arassari da Cunha & Frota-Pessoa 1947: 32. 
araucana Brncic 195 7a: 82. 
arauna Pavan & Nacrur 1950:268. 
argentata de Meijere 1914:258. To Leuco- 
phenga. 
argenteifrons Wheeler 1954:50. 
argentina de Meijere 1924:46. To Leuco- 
phenga. 
arizonensis Patt3rson & Wheeler 1942:96. 
asozana Okada 1956:87. 
astioidea Duda 1923:42. 
aterrima Duda 1940:28, 48. 
athabasca Sturtevant & Dobzhansky 1936:576. 
atkinsoni (Miller 1921:302, in Leucophenga). 

Syn. of funebris (Fabricius). 
atra Walker 1852:412. Sp. inc. 
atrala Burla & Pavan 1953:307. 
atropyga Duda 1924a: 215. as var. of montium. 



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auraria Peng 1937:23. 

aurea Patterson & Mainland 1944:46. 

aureata Wheeler 1957:83. 

australica Duda 1923:59. Poss. syn. of obsoleta 

Malloch. 
austrorepleta Dobzhansky & Pavan 1943:50. 

Syn. of repleta Wollaston. 
austrosaltans Spassky 1957:57. 
azteca Sturtevant & Dobzhansky 1936:577. 
baeomyia Wheeler 1949a: 145. Syn. of lati- 

fasciaeformis Duda. 
balneorum Sturtevant 1927:369. 
balteata Bergroth 1894:75. Syn. of melano- 

gaster Meigen. 
bandeirantorum Dobzhansky & Pavan 1943: 

30. 
bangi Burla 1954a: 143. 
baoleBurla 1954a: 187. 
basdeni Wheeler 1957:94. 
baseogrisea Duda 1924a: 221, & 1924b: 25 7. 
basilaris Adams 1905:184. To Leucophenga. 
bellula Bergroth 1894:75. To Leucophenga. 
bellula Williston 1896:410. Preocc; n. n. is 

pulchella Sturtevant. 
berryi Cockerell 1923:332. Fossil sp. 
betari Dobzhansky & Pavan 1943:48. 
biarmipes Malloch 1 924c : 64. 
bicolor de Meijere 191 1 :399. To Lissocephala. 
bicoloripes Malloch 1926:31. To Chymomyza. 
bifasciata Pomini 1940:163. {bifasciata & bi- 

lineata w^ere both original spellings; bilin- 

eata being preocc, bifasciata is an available 

replacement name), 
bifilum Frota-Pessoa 1954:284. 
bifurca Patterson & Wheeler 1942:85. 
bilimbata Bezzi 1928:159. Syn. of nasuta 

Lamb. 
bilineata Williston 1896:409. To Zygothrica. 
bilineata Pomini 1940:155. Preocc; replace- 
ment name is bifasciata Pomini. 
blmaculata Loew 1865:183. To Leucophenga. 
binotata de Meijere 1914:257. 
biopaca Sturtevant 1942:37. Syn. of sturte- 

vanti Duda. 
bipectinata Duda 1923:52. 
bipunctata Patterson & Mainland 1943:194. 
biradiata Duda 1923:50. 
bistriata de Meijere 1911:397. To Zaprionus, 

subg. Phorticella. 
bizonata Kikkawa & Peng 1938:532. 
bocainensis Pavan & da Cunha 1947: 18. 
bocainoides Carson 1954:150. 
boletina Duda 1927:202. 
boliviana Duda 1927:198. 
boliviensis Duda 1927:215, as var. of kertes- 

zina Duda. 



borealis Patterson 1952:20. 
brachynephros Okada 1956:126. 
brevicarinata Patterson & WTieeler 1942:88. 
brevicornis Duda 1935a: 76. 
brevis Walker 1852:411. Sp. inc. 
briegeri Pavan & Breuer 1954:459. 
bromeliae Sturtevant 1921:72. 
bromelioides Pavan & da Cunha 1947:7. 
brouni Hutton 1901:91. Officially suppressed 

in favor of immigrans Sturtevant. 
brunetti Chaudhuri & Mukherjee 1941:216. 
brunnea de Meijere 1911:401. 
brunneipalpa Dobzhansky & Pavan 1943:53. 
brunneipennis Malloch 1923:617. 
bryani Malloch 1934a: 3 10. 
busckii Coquillett 1901:18. (Invalid original 

spelling, buskii, was a typographical error). 

Type of subg. Dorsilopha. 
buzzatii Patterson & Wheeler 1942:97. 
calceolata Duda 1926a: 94, 105. 
calif ornica Sturtevant 1923:9. 
caliginosa Lamb 1914:341. 
calloptera Schiner 1868:239. 
camargoi Dobzhansky & Pavan 1950:6. 
camaronensis Brncic 1957a: 95. 
cameraria Haliday 1833:174. 
campestris Burla 1950b: 9. Prob. syn. of cro- 

cina Patterson & Mainland. 
canalinea Patterson & Mainland 1944:50. 
canalinioides Wheeler 1957:92. 
canapalpa Patterson & Mainland 1944:40. 
cancellata Mather 1955:550. 
canescens Duda 1927:210. 
capnoptera Patterson & Mainland 1944:47. 
caponei Pavan & da Cunha 1947:4. 
capricorni Dobzhansky & Pavan 1943:14. 
carbonaria Patterson & Wheeler 1942:103. 
cardini Sturtevant 1916:336. 
cardinoides Dobzhansky & Pavan 1943:21. 
caribea Sturtevant 1916:335. Syn. of ananas- 

sae Doleschall. 
carinata Grimshaw 1901:70. 
carinata Duda 1923:41. Preocc; n. n. is lati- 

frontata Frota-Pessoa. 
carsoni Wheeler 1957:95. 
castanea Patterson & Mainland 1944:51. 
caxiensis Cordeiro 1952:304. 
cellaris (Linne 1758:597, in Musca). Sp. inc.; 

consid. Drosophila^ especially D. funebris, by 

older authors; prob. to Phoridae (see Schiner 

1864:278, and Duda 1935a: 84). 
centralis Patterson & Mainland 1944:57, as 

subsp. of pallidipennis. 
chagrinensis Stalker & Spencer 1939:111. 
cheda Tan, Hsu & Sheng 1949: 199. 
chinoi Okada 1956:162. 



Wheeler: Nomenclature of Drosophila 



189 



cilifemur Villeneuve 1923:28. Syn. of immi- 
grans Sturtevant. 

cilitarsus Hering 1940:293. 

cincta de Meijere 1911:395. To Leucophenga. 

cinctifrons de Meijere i. litt. of Duda 1924a: 
227. Invalid citation; sp. is Chymomyza 
cinctifrons de Meijere 1924:47. 

cinerea Patterson & Wheeler 1942: 71 . 

cinerella Fallen 1823:7. Becker 1926:43 lists 
''Drosophila cinerella Meigen 1830" as syn. 
of Discocerina plumosa (Fallen 1823) of the 
Ephydridae, meaning cinerella Fallen as 
understood by Meigen; to Scaptomyza (Bas- 
den, pers. comm.). 

circumdata Duda 1926a: 82, 84. Prob. to 
Chaetodrosophilella. 

clarkii Hutton 1901:91. Syn. of funebris (Fa- 
bricius). 

clunicrus Duda 1923:51. 

coflf eata Williston 1 896 : 409. 

coflfeina Schiner 1868:238. 

cognata Grimshaw 1901:69. 

colocasiae Duda 1924b: 252. 

colorata Walker 1849:1110. 

comoe Burla 1954a: 205. 

compressiceps Duda 1923:55. 

compressifrons of Hennig 1941:151. Err. sub- 
seq. spelling for compressiceps. 

confusa Staeger 1844:16. 

congesta Zetterstedt 1847:2558. To Micro- 
drosophila. 

conspicua Grimshaw 1901:59. 

converga Heed & Wheeler 1957:22. 

convergens de Meijere 1911:400. To Orthoste- 
gana (Hendel 1914); to Stegana (Sturte- 
vant 1921); to Oxyphortica (Duda 1923). 

convexa Malloch 1934a: 303. 

coracina Kikkawa & Peng 1938:523. 

cordata Sturtevant 1942:34. 

costata Zetterstedt 1838:776. To Chymomyza. 

crassa Patterson & Mainland 1944:52. 

crassifemur Grimshaw 1901:66. 

crocina Patterson & Mainland 1944:34. 

crockeri Curran 1936:44. 

crucigera Grimshaw 1902:86. 

cubana Townsend 1954:339, as subsp. of tropi- 
calis. 

curvapex Frota-Pessoa 1954:296. 

curvicapillata Duda 1923:49. 

curviceps Okada & Kurokawa 1957:8. 

curvinervis Duda 1924a: 204, as var. of longe- 
crinita Duda. 

curvipennis Fallen 1823:4. To Protostegana. 

cuzcoica Duda 1927:120. 

daruma Okada 1956:155. 
debilis Walker 1849: 1109. Sp. inc. 



decemguttata Walker 1852:411. To Diastati- 
dae. 

decemseriata Hendel 1936:98. 

decipiens Duda 1923:55. 

deflecta Malloch 1924d:36. 

deflexa Duda 1924a: 222. 

deltaneuron Bryan 1938:40. 

denieri Blanchard 1938:362. 

dentata Duda 1924a: 205, as var. of longecri- 
nita; raised to sp. by Duda 1926a: 65, 69. 
(Nom. nud. in Duda 1923:42). 

dentata Duda 1927:201. Preocc; n. n. is ten- 
data Wheeler. 

denticeps Okada & Sasakawa 1956:26. 

diama Burla 1954a: 192. 

dibi Burla 1954a: 126. 

dilacerata Becker 1919:208. Consid. Scapto- 
myza by Duda 1927, but descr. & wing fig. 
indicate Drosophila. 

dimidiata Loew 1862:230. To Mycodrosophila. 

dispar Mather 1955:570. 

distincta Egger 1862:780. To Chymomyza. 

divisa Duda 1927:187. {diversa is an err. sub- 
seq. spelling in Patterson & Wheeler 1949). 

dobzhanskii Patterson 1943:82. 

dorsalis Walker 1865:128. 

dorsata Duda 1924a:207, 220, & 1924b:248. 
(Nom. nud. in Duda 1923:56). 

dorsivitta Walker 1 86 1 : 330. Sp. inc. 

dreyfusi Dobzhansky & Pavan 1943:61. 

dubia Sturtevant 1921:73. To Clastopteromyia. 

dudai Malloch 1934b: 444. 

dumuya Burla 1954a: 185. 

duncani Sturtevant 191 8a: 446. 

dunni Townsend & Wheeler 1955:61. - 

dyula Burla 1954a: 178. 

dyaramankana Burla 1954a: 196. 

earlei Sturtevant 1916:329. 

elliptica Sturtevant 1942:35. 

elongata Sturtevant 1927:372. 

emarginata Sturtevant 1942:36. 

emulata Chaudhuri & Mukherjee 1941:216. 

enderbii Hutton 1902:174. To Ephydridae. 

enigma Malloch 1927:6. 

equinoxialis Dobzhansky 1946:209. 

errans Malloch 1933:21. N. n. for similis 
Lamb, not Williston; syn. of ananassae 
Doleschall. 

erythrophthalma (Panzer 1794:24, in Mus- 
ca). Sp. inc., poss. not Drosophila (Duda 
1935a). 

euronotus Patterson & Ward 1952:158. 

excepta Malloch 1934a: 308. 

exeita Giglio-Tos 1893:14. To Ephydridae. 

exigua Grimshaw 1901:72. 

ezoana Takada & Okada 1958: 134. 



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facialba Heed & Wheeler 1957: 19. 

facialis Adams 1905:183. 

fasciata Meigen 1830:84. Consid. by some au- 
thors as prior name (page preference) for 
melanogaster Meigen. 

fasciata Dufour 1839:49. Preocc, but consid. 
syn. of testacea von Roser by Duda 1935a. 

fasciola Williston 1896:410. 

fascioloides Dobzhansky & Pavan 1943:42. 

fenestrarvim Fallen 1823:6. 

fenestrata de Meijere of Duda 1923:36. In- 
valid citation from museum label; valid 
name is Zaprionus (Phorticella) fenestrata 
(Duda) 1923. 

ferruginea Becker 1919:209. 

ficusphila Kikkawa & Peng 1938:531. 

fima Burla 1954a: 165. 

finigutta Walker 1859: 126. To Lauxaniidae. 

finitima Lamb 1914:340. 

flava Fallen 1823:7. Often consid. Scaptomyza 
but Collin (1953:150) consid. it Drosophila. 
fenestrarum species group. 

flaveola (Meigen 1830:66, in Notiphila). To 
Scaptomyza ace. to many authors; to Hy- 
drellia (Ephydridae) by Duda 1934. 

flaviceps Grimshaw 1901:63. 

flavipennis Zetterstedt 1838:777. To Scapto- 
myza. 

flavipes (Meigen 1830:108, in Opomyza). 
Consid. Drosophila by some older authors; 
Hennig (1937:31) makes it syn. of Phyllo- 
myza securicornis Fallen in Milichiidae. 

flaviseta Adams 1905:184. To Leucophenga. 

flavissima Duda 1929:419. 

flavohalterata Duda 1925:198. 

flavohirta Malloch 1924b: 354. 

flavolineata Duda 1927:157. 

flavomontana Patterson 1952:21. 

flavopilosa Frey 1919:14. 

flavopinicola Wheeler 1954:47. 

flavorepleta Patterson & Pavan 1952:114, as 
subsp. of fulvimacula. 

flexa Loew 1865:182. 

florae Sturtevant 1916:339. 

floricola Sturtevant 1942:42. Type of subg. 
Phloridosa. 

forcipata Collin 1952:198. Poss. syn. of anda- 
lusiaca Strobl. 

formosana Duda 1926b: 50, as var. of immi- 
grant Sturtevant (as ^'tripunctata Becker"); 
raised to sp. by Duda 1926a: 83. 

formosana Sturtevant 1927:368, as var. of im- 
migrans Sturtevant. Preoc; syn. of formo- 
sana Duda. 

fraburu Burla 1954a: 183. 



fracticosta Lamb 1914:329. To Mycodro- 
sophila. 

fragilis Wheeler 1949b: 191. 

framire Burla 1954a: 128. 

frolovae Wheeler 1949b: 175. 

frontalis Williston 1896:413. To Leucophenga. 

frontata de Meijere 1916:204. To Microdro- 
sophila. 

fronto Walker 1852:410. Sp. inc. 

fruhstorferi Duda 1924a: 211, 218. & 1924b: 
255. 

fuliginea Patterson & Wheeler 1942:80. Prob. 
syn. of californica Sturtevant. 

fulvalineata Patterson & Wheeler 1942: 106. 

fulvimacula Patterson & Mainland 1944:42. 

fulvimaculoides Wasserman & Wilson 1957: 
137.' 

fumipennis Duda 1925:220. 

fumosa Pavan & da Cunha 1947: 14. 

fundomaculata Duda 1925:209. 

funebris (Fabricius 1787:345. in Musca) . 
Type of gen. & subg. Drosophila. 

fungicola Villeneuve 1921:158. Syn. of uni- 
striata Strobl. 

fusca Coquillett 1900:264. Sp. inc. 

fuscimana Zetterstedt 1838:776. To Chymo- 
myza. 

fuscipennis Duda 1927:198. 

fuscithorax Malloch 1924b: 353. 

fuscoamoeba Bryan 1934:438. 

fuscohalterata Duda 1925:197. 

fuscolineata Duda 1925:213. 

fuscovittata Harrison 1954:106. 

fuscula Fallen 1823:7. To Diastatidae. 

gasici Brncic 195 7a: 92. 

gaucha Jaeger & Salzano 1953:205. 

gibberosa Patterson & Mainland 1943: 195. 

gibbinsi Aubertin 1937:169. 

gibbosa de Meijere 1914:264. To Leuco- 
phenga. 

gigantea Thomson 1869:596. Poss. Leuco- 
phenga (Kahl 1917:392); poss. Curtonotum 
(Malloch 1934b:437); true Drosophila (Sa- 
brosky, pers. comm.). 

gigas Duda 1925:216. 

gilva Burla 1956:263. 

glabra Fallen 1823:8. To Camillidae. 

glabrifrons Duda 1925:196. 

gracilipes Duda 1940:27, 39, as var. of finitima 
Lamb; prob. syn. of latifasciaeformis Duda 
(Burla 1954a). 

gracilis (Duda 1924a: 192, & 1924b: 253, in 
Tanygastrella) . To Drosophila by Duda 
1926a: 99, with Tanygastrella as subgen. of 
Drosophila. 

graminum Fallen 1823:8. To Scaptomyza. 



Wheeler: Nomenclature of Drosophila 



191 



grandis Kikkawa & Peng 1938:543. 

gratiosa de Meijere 1911:404. To Mycodro- 

sophila. 
grimshawi Oldenherg 1914:23. N. n. for varie- 

gata Grimshaw, not Fallen. 
grischiina Burla 1950a: 620. Syn. of confusa 

Staeger. 
grisea Patterson & Wheeler 1942: 72. 
griseicoUis Becker 1919:209. 
griseola Zetterstedt 1847:2562. To Scapto- 

myza. 
griseolineata Duda 1927:161. 
grossipalpis Lamb 1914:328. To Leuco- 

phenga. 
guaraja King 1947:48. 

guaramunu Dobzhansky & Pavan 1943:39. 
guarani Dobzhansky & Pavan 1943:36. 
guarii Dobzhansky & Pavan 1943:37. 
guinensis Duda 1924a: 209. Err. subseq. spell- 
ing for novoguineensis Duda. 
guttifera Walker 1849:1110. 
guttiventris de Meijere 1908:331. N. n. for 

maculiventris de Meijere, not van der Wulp. 

To Leucophenga. 
guyenoti Burla 1948:277. Poss. syn. of deflexa 

Duda. 
haleakalae Grimshaw 1901:64. 
hamatofila Patterson & Wheeler 1942:91. 
hawaiiensis Grimshaw 1901:60. 
helvetica Burla 1948:276. 

heterobristalis Tan, Hsu, & Sheng 1949:204. 
hexastigma Patterson & Mainland 1944: 43. 
hexastriata Tan, Hsu, & Sheng 1949:201. 
hirsuta Duda 1926a: 94, 97. Proposed as n. sp. 

or var., but cited, p. 97, as var. of fenes- 

Lrarum. 
hirticornis de Meijere 1914:261. 
hirtipes Lamb 1914:337. 
hirtiscutellata Sturtevant 1927:372. 
histrio Meigen 1830:85. 
histrioides Okada & Kurokawa 1957:4. 
hoeckeri Brncic 1957a: 76. Prob. syn. of nigri- 

cruria Patterson & Mainland. 
hoozani Duda 1923:54. 
huilliehe Brncic 195 7a: 85. 
humeralis Grimshaw 1901:64. 
hyalipennis Duda 1927:119. 
hydei Sturtevant 1921:101. 
hy decides Patterson & Wheeler 1942:84. 
hypocausta Osten-Sacken 1882:245. 
hypopygialis (Duda 1924a: 192, & 1924b:254, 

in Tanygastrella) . To Drosophila by Duda 

1926a: 96, with Tanygastrella as subg. of 

Drosophila. 
hypopygialis Malloch 1934a: 307. 
icteroscuta Wheeler 1949b: 184. 



iki Bryan 1934:439, as var. of nigra Grimshaw. 

illata Walker 1860:168. Sp. inc. 

illota Williston 1896:415. 

imeretensis Sokolov 1948:1007. 

immatura Walker 1849:1108. Sp. inc. 

immigrans Sturtevant 1921:83. 

imparata Walker 1859:126. Syn. of ananassae 
Doleschall (de Meijere). 

impvidica Duda 1927:196. 

inaequalis Grimshaw 1901:69. 

inca Dobzhansky & Pavan 1943:44. 

incana Meigen 1830:86. To Scaptomyza. 

inconspicua de Meijere 1914:262. 

infuscata Grimshaw 1901:63. 

ingrata Haliday 1833:174. Syn. of obscura 
Fallen (Duda 1935a). 

innocua (Malloch 1934a: 294, in Hirtodro- 
sophila) . 

innubila Spencer 1943:94. 

inornata Malloch 1923:617. 

insulana Schiner 1868:240. To Leucophenga. 

insularis Dobzhansky 1957:41. 

intermedia Duda 1927:125, 151, as var. of 
adusta in Scaptomyza which was consid. 
subg. of Drosophila. To Parascaptomyza. 

interrvipta Duda 1923:45. 

inversa Walker 1861:331. To Clastopteromyia. 

invicta (Walker 1857:130, in Helomyza). Con- 
sid. Drosophila by some older authors; to 
Leucophenga (Czerny) ; to T richiaspiphen- 
ga (Duda); to Paraleucophenga Hendel 
(Hennig 1941:150) whose type species, tri- 
seta Hendel, is syn. of invicta. 

iri Burla 1954a: 180. 

iroko Burla 1954a: 171. 

iroquois Sturtevant & Dobzhansky 1936:576, 
as subsp. of affinis. 

itambacuriensis da Cunha 1955:119, as subsp. 
of neocardini. 

jacobsoni Duda 1926a: 65, as var. of latifrons 
Duda. 

johni Pokorny 1896:63. To Mycodrosophila. 

jordanensis Frota-Pessoa 1945:473. 

jucunda Lamb 1914:339. 

kallima Wheeler 1957:87. 

kauluai Bryan 1934:439. 

kerteszina Duda 1925:222. 

kikkawai Burla 1954b: 47. 

kirki Harrison 1959:303. 

komaii Kikkawa & Peng 1938: 525. 

krugi Pavan & Breuer 1954:462. 

kulango Burla 1954a: 172. 

kuntzei Duda 1924a:218. 

kuoni Burla 1954a: 190. 

kuscheli Brncic 1957b: 394. 

kweichowensis Tan, Hsu, & Sheng 1949:205. 



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lacertosa Okada 1956: 158. 

lacicola Patterson 1944:102. 

lacteoguttata Portschinsky 1892:226. To Ami- 
ota. 

laeta Zetterstedt 1847:2553, as var. of trans- 
versa Fallen. 

Iambi Duda 1940:48. N. n. for pallipes Lamb, 
not Dufour. 

lamellitarsis Duda 1936:350. 

lanaiensis Grimshaw 1901:60. 

lata Becker 1907:54. To Amiota. subg. Para- 
phortica. 

latebuccata Duda 1927:203. 

latecarinata Duda 1927:170. 

lateralis Walker 1860:169. 

latestriata Becker 1908:157. Syn. of unistriata 
Strobl (Duda 1924c). 

latifascia de Meijere 1914:261. {latifasciata is 
err. subseq. spelling). 

latifasciaeformis Duda 1940:22. 

latifrons Adams 1905:182. 

latifrons Duda 1926a: 64. Preocc; in validly 
proposed as n. sp. to include carinata & asti- 
oidea as vars.; n. n. is latifrontata Frota-Pes- 
soa. 

latifrontata Frota-Pessoa 1945:480. N. n. for 
latifrons Duda, not Adams (and for carinata 
Duda, not Grimshaw; see Frota-Pessoa 1945, 
op. cit. ) . Type of subg. Hirtodrosophila. 

lativittata Malloch 1923:618. 

lativittata Malloch 1924d:36. Preocc; n. n. is 
palustris Spencer. 

lebanonensis Wheeler 1949a: 143. Prob. syn. 
of victoria Sturtevant. 

leonis Patterson & Wheeler 1942:82. 

levigata Burla 1956:261. 

levis Mather 1955:561. Syn. of bryani Mal- 
loch. 

limbata von Roser 1840:62. 

limbata Williston 1896:414. Preocc.; n. n. is 
nebulosa Sturtevant. 

limbinervis Duda 1925:215. 

limbipennis de Meijere 1908:156. To Leuco- 
phenga. 

limbiventris Duda 1925:212. 

limensis Pavan & Patterson 1947:26. 

limpiensis Mainland 1941:160, as subsp. of 
macrospina. 

linearepleta Patterson & Wheeler 1942:79. 

linearis Walker 1852:411. Sp. inc., prob. not 
Drosophila. 

lineata van der Wulp 1886:57. 

lineata (de Meijere 1911:420, in Stegana) . 
Preocc; replacement name is multistriata 
Duda (see remarks in text). 

lineolata de Meijere 1914:254. 



litorella (Meigen 1838:374, in Hydrellia). Sp. 

inc., prob. not Drosophila (Duda 1935a). 
littoralis Meigen 1830:87. 
lividinervis Duda 1923:53. 
lividipennis Duda 1924a: 21 6. Err. subseq. 

spelling for lividinervis Duda. 
longala Patterson & Wheeler 1942:71. 
longecrinita Duda 1924a: 204, & 1924b: 242. 

(Nom. nud. in Duda 1923:42). 
longicornis Patterson & Wheeler 1942:90. 
longifrons Duda 1923:48. 
longiseta Grimshaw 1901:68. 
longitarsis Duda 1931:195. 
lueida Segiiy 1938:351. 
lugens Duda 1926a: 71, 7Q. 
lugubrina Duda 1924a: 224. Syn. of littoralis 

Meigen. 
lugubripennis Duda 1927:199. 
lulumahu Bryan 1938:39. 
lundstroemi Duda 1935a: 72. 
lurida Walker 1860:169. 
lutea Kikkawa & Peng 1938:533. 
luteipes Sturtevant 1921:74, as var. of splen- 

dida; to Clastopteromyia. 
lutzii Sturtevant 1916:340. 
macropolia Patterson & Mainland 1944:54. 
macroptera Patterson & Wheeler 1942:105. 
macrospina Stalker & Spencer 1939: 110. 
macularis Villeneuve 1921:157. Syn. of picta 

Zetterstedt. 
macula ta Dufour 1839:49. To Leucophenga. 
maculifrons Duda 1927:122. 
maculinotata Okada 1956:145. 
maculipennis Gimmerthal 1847:199. To Dias- 

tatidae. 
maculipennis Duda 1927:167. Preocc; n. n. is 

peruensis Wheeler. 
maculiventris van der Wulp 1897:142, Con- 

sid. syn. of repleta Wollaston. 
maculiventris de Meijere 1908:155. Preocc; 

n. n. is guttiventris de Meijere. To Leuco- 
phenga. 
maculosa Coquillett 1895:317, in Johnson, D. 

W., 1895. To Leucophenga. 
maculosa Mather 1955:560. Preocc; n. n. is 

novamaculosa Mather. 
magnarcus Frota-Pessoa 1951:407. 
magnabadia Patterson & Mainland 1943: 196. 
magnafumosa Stalker & Spencer 1939:112. 
magnaquinaria Wheeler 1954:48. 
magnipectinata Okada 1956:113. 
mahican Sturtevant & Dobzhansky 1936:576, 

as subsp. of athabasca. 
mainlandi Patterson 1943:147. 
makinoi Okada 1956:135. 
mallochi Frota-Pessoa 1946: 155. N. n. for lati- 



Wheeler: Nomenclature of Drosophila 



193 



vittata Malloch 1924, not Malloch 1923; syn. 

of palustris Spencer. 
mangabeirai Malogolowkin 1951:432. 
manonoensis Harrison 1954:101. 
mansiira Adams 1905:185. To Leucophenga. 
maracaya Wheeler 1957:88. 
marginata Duda 1924a: 209, & 1924b: 244. 

(Norn. nud. in Duda 1923:46). 
marginella Zetterstedt 1838:777. To Diasta- 

tidae. 
marjoryae Harrison 1954:105. 
marmoria Hutton 1901:91. Sp. inc., poss. is 

hydei Sturtevant. 
martensis Wasserman & Wilson 1957:151. 
mauiensis Grimshaw 1901:67. 
maura de Meijere 1911:406. To Pararhinoleu- 

cophenga. 
mbettie Burla 1954a: 149. 
mediocris Frota-Pessoa 1954:275. 
mediodelta Heed & Wheeler 1957:21. 
mediodiffusa Heed & Wheeler 1957:27. 
medioimpressa Frota-Pessoa 1954:300. 
medionotata Frota-Pessoa 1954:288. 
medioobscurata Duda 1925:217. 
medioparva Heed & Wheeler 1957:28. 
mediopicta Frota-Pessoa 1954:290. 
mediopictoides Heed & Wheeler 1957:24. 
mediopunctata Dobzhansky & Pavan 1943:26. 
mediosignata Dobzhansky & Pavan 1943:24. 
mediostriata Duda 1925:223. 
mediovittata Frota-Pessoa 1954:280. 
megaspis Bezzi 1908b: 191. 
meigeni Duda 1935a: 74, 90, as var. of obscura; 

consid. sp. by Basden 1954:617. 
meijerei Wheeler 1959, antea. N. n. for nigri- 

color de Meijere, not Strobl. 
meitanensis Tan, Hsu, & Sheng 1949:204. 
melanderi Sturtevant 1916:337. 
melanica Sturtevant 1916:332. 
melanissima Sturtevant 1916:333. 
melanogasler Meigen 1830:85. Type of subg. 

Sophophora. 
melanogaster Macquart 1843:415. Preocc; 

syn. of fenestrarum Fallen (Duda). 
melanopalpa Patterson & Wheeler 1942:77. 
melanoptera Duda 1927:158. 
melanosoma Grimshaw 1901:68. 
melanospila Walker 1859:126. To Lauxan- 

idae 
melanura Miller 1944:86. 
mellea Becker 1919:208. 
mercatorum Patterson & Wheeler 1942:93. 
meridiana Patterson & Wheeler 1942:99. 
mesophragniatica Duda 1927:205. 
mesostigma Frota-Pessoa 1954:269. 



metallescens de Meijere 1914:265. To Lisso- 

cephala. 
metallescens (Malloch 1934a: 312, in Spinulo- 

phila). Prob. syn. of monochaeta Sturtevant. 
metallica Sturtevant 1921:73. To Clastop- 

teromyia. 
metzii Sturtevant 1921:78. 
mexicana Macquart 1843:416. Sp. inc. 
mexicana (Wheeler 1949b: 164, in Paramyco- 

drosophila) . Preocc; n. n. is mexicoa 

Wheeler. 
mexicoa Wheeler 1954:54. N. n. for mexicana 

(Wheeler), not Macquart. 
microlabis Segiiy 1938:351. 
micromelanica Patterson 1941:394. 
miki Duda 1924a: 21 3. 
minuta Walker 1852:412. Sp. inc. 
minuta Duda 1926a: 66, 70, as var. of dentata 

Duda 1924. 
miranda Dobzhansky 1935:377. 
mirim Dobzhansky & Pavan 1943:62. Syn. of 

latifasciaeformis Duda. 
mitis Curran 1936:43. 
modesta Sturtevant 1916:338. Syn. of tripunc- 

tata Loew. 
mojavensis Patterson & Crow 1940:251, as 

subsp. of mulleri; later consid. sp. 
mojii Pavan 1950:19. 
mokonfim Burla 1954a: 131. 
molokaiensis Grimshaw 1901:67. 
monochaeta Sturtevant 1927:368. 
montana Stone, Griffen, & Patterson 1941 : 1 72. 
monticola Grimshaw 1901:69. 
montiiim de Meijere 1916:205. 
morena Frota-Pessoa 1954:283. 
moriwakii Okada & Kurokawa 1957:9. 
moronu Burla 1954a: 134. 
mourensis da Cunha 1955:119, as subsp. of 

neocardini. 
mvilleri Sturtevant 1921:101. 
multipunclata Loew 1866:50. Syn. of gutti- 

fera Walker. 
multispina Okada 1956:143. 
mviltistriata Duda 1923:57. 
munda Spencer 1942:58. 

mutabilis Adams 1905:187. To Leucophenga. 
mutandis Tan, Hsu, & Sheng 1949: 198. 
mycetophaga Malloch 1924b: 351. 
nana Williston 1896:416. To Clastopteromyia. 
nannoptera Wheeler 1949b: 177. 
narinosa Frota-Pessoa 1945:476. N. n. for na- 

salis Duda. not Grimshaw. 
narragansett Sturtevant & Dobzhansky 1936: 

577. 
nasalis Grimshaw 1901:66. 



194 



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nasalis Duda 1925:194. Preocc; n. n. is nari- 

nosa Frota-Pessoa. 
nasuta Lamb 1914:346. 
nebulosa Sturtevant 1916:327. N. n. for lim- 

bata Williston, not von Roser. 
neocardini Streisinger 1946:111. 
neochracea Wheeler 1949. antea. N. n. for 

ochracea Duda, not Grimshaw. 
neocordata Magalhaes 1956:275. 
neoelliptica Pavan & Magalhaes 1950:13. 
neoguaramunii Frydenberg 1956:57. 
neomorpha Heed & Wheeler 1957:33. 
neorepleta Patterson & Wheeler 1942: 78. 
neosaltans Pavan & Magalhaes 1950: 16. 
neozelandica Harrison 1959:290. 
nepalensis Okada 1955:388. 
nicholgoni Malloch 1927:4. 
nigerrima Lamb 1914:331. To Mycodro- 

sophila. 
nigra Grimshaw 1901:62. 
nigra de Meijere 1908:153. Preocc; n. n. was 

nigricolor de Meijere, also preocc.; n. n. is 

meijerei Wheeler. 
nigra Duda 1926a: 65, 68, as var. of latifrons 

Duda. 
nigriceps Meigen 1838:378. Consid. syn. of 

obscura Fallen. 
nigricincta Frota-Pessoa 1954:289. 
nigricolor Strobl 1898b: 266. 
nigricolor de Meijere 1911:399. N. n. for 

nigra de Meijere, not Grimshaw; preocc; 

n. n. is meijerei Wheeler. 
nigricosta Malloch 1926:30. To 'Neotanygas- 

trella. 
nigricruria Patterson & Mainland 1943:136. 
nigrifemur Duda 1927:192. 
nigrifrons Malloch 1934a: 304. 
nigrimana Meigen 1830:87. To Chymomyza. 
nigrita Haliday of authors. Err. subseq. spell- 
ing for ingrata Haliday, possibly intended as 

an emendation. 
nigrithorax Strobl 1893b: 132. as var. of fenes- 

trarum. 
nigriventris Macquart 1843:416. To Leuco- 

phenga. 
nigriventris Zetterstedt 1847:2557. Preocc; to 

Microdrosophila with n. n. Microdrosophila 

zetterstedti Wheeler. 
nigrobrunnea Lamb 1914:332. To Mycodro- 

sophila. 
nigrodunni Heed & Wheeler 1957:35. 
nigrofemorata Duda 1926a: 72, 96, 110. 
nigrohalterata Duda 1925: 195. 
nigrohydei Patterson & Wheeler 1942:84. 
nigroniaculata Kikkawa & Peng 1938:537. 
nigronielanica Patterson & Wheeler 1942: 100. 



nigropunctata van der Wulp 1892:216. Con- 
sid. syn. of repleta Wollaston. 
nigrosparsa Strobl 1898b: 267. Type of subg. 

Spinodrosophila. 
nigrospiracula Patterson & Wheeler 1942:81. 
nigrovittata Malloch 1924b: 352. To Dettop- 

somyia. 
nikananu Burla 1954a: 160. 
nipponica Kikkawa & Peng 1938:531. 
nitens Buzzati-Traverso 1943:2. Syn. of rufi- 

frons Loew. 
nitidapex Bigot 1891:279. Sp. inc.. prob. not 

Drosophila. 
nitidithorax Malloch 1927:5. 
nitidiventris Macquart 1835:551. Consid. syn. 

of fenestrarum Fallen. 
niveopunctata Dufour 1846b: 318. Nom. nud. 

ace to Oldenberg 1914:25 ff. 
nixifrons Tan, Hsu. & Sheng 1949:202. 
nodosa Duda 1926a: 94, 103. 
nokogiri Okada 1956:84. 
notabilis Lamb 1914:329. Prob. to Hypselo- 

thyrea. 
novamaculosa Mather 1956:65. N. n. for 

maculosa Mather, not Coquillett. 
novamexicana Patterson 1941a: 535. as subsp. 

of virilis, later raised to sp. 
novemaristata Dobzhansky & Pavan 1943:55. 
novoguineensis Duda 1923:46. Err. subseq. 

spellings are novoguinensis and guinensis. 
novopaca Mather 1956:65. N. n. for opaca 

Mather, not Williston. 
mibihma Wheeler 1949b: 188. 
mitrita Duda 1935b: 33. 

obesa Loew 1872:102. To Rhinoleucophenga. 
obscura Fallen 1823:6. 
obscurata de Meijere 1911:410. 
obscuricolor Duda 1927:190. 
obscuricornis Grimshaw 1901:71. 
obscuricornis (de Meijere 191 5b: 94 in Ste- 

gana) . Consid. Drosophila by Duda 1924a 

&b. 
obscurifrons Grimshaw 1901:72. 
obscuripennis Loew 1865:183. To Leuco- 

phenga. 
obscuroides Pomini 1940:149. Syn. of obscura 

Fallen. 
obsoleta Malloch 1923:616. Consid. by Mal- 
loch (1925) as prob. same as "australis 

Duda." prob. meaning australica Duda 1923; 

if true, Malloch's name is prior (Dec. 14 vs. 

Dec. 24). 
occidentalis Spencer 1942:60. 
ochracea Grimshaw^ 1901:61. 
ochracea Duda 1927:195. Preocc; n. n. is 72^0- 

chracea Wheeler. 



Wheeler: Nomenclature of Drosophila 



195 



ochracella Hendel 1936:98. Prob. to Zygothri- 

ca (Burla 1956). 
ochrifrons Duda 1924a:223, & 1924b:258. 
oenopota (Scopoli 1763:337, in Musca) . Sp. 

inc., prob. not Drosophila (Duda 1935a). 
ohioensis Spencer 1940b: 303, as subsp. of 

macrospina. 
olaae Grimshaw 1901:66. 
oWenbergi Duda 1924a: 204. 
omogoensis Okada 1956:82. 
onca Dobzhansky & Pa van 1943:40. 
onychophora Duda 1927:208. 
opaca Williston 1896:41 1. To Clastopteromyia. 
opaca Mather 1955:558. Preocc; n. n. is novo- 

paca Mather. 
opisthomelaina Nolte & Stoch of Nolte 1958: 

519. Manuscript name based on prior use in 

the unofficial Drosophila Information Serv- 
ice. Syn. of yakuba Burla. 
oralis Duda 1923:44. 

orbitalis Sturtevant 1916:336. To Zygothrica. 
orbospiracula Patterson & Wheeler 1942:70. 
ordinaria Coquillett 1904:190. 
orkui Brncic & Santibanez 1957:69. 
ornatifrons Duda 1927:162. 
ornatipennis Williston 1896:407. 
ornatipennis de Meijere 1914:256. Preocc. ; to 

Leucophenga, with n. n. Leucophenga or- 

nata Wheeler. 
osornina Brncic 195 7a: 97. 
pachea Patterson & Wheeler 1942:87. 
pallida Zetterstedt 1847:2571. To Parascapto- 

myza (Basden 1957:209). 
pallida Williston 1896:415. Preocc; n. n. is 

willistoni Sturtevant. 
pallidipennis Dobzhansky & Pa van 1943:32. 
pallipes Dufour 1846a: 323. Sp. inc., prob. to 

Diastatidae. 
pallipes Lamb 1914:342. Preocc; n. n. is 

Iambi Duda. 
palpalis Adams 1905:185. To Leucophenga. 
palustris Spencer 1942:63. N. n. for lativittata 

Malloch 1924, not 1923. 
panamensis Malloch 1926:28. 
para Pavan & Burla 1950:22. 
parabocainensis Carson 1954:149. 
paracanalinea Wheeler 1957:93. 
parachrogaster Patterson & Mainland 1943: 

197. 
paradoxa Lamb 1918:159. To Clastoptero- 
myia. 
paraguayensis Duda 1927:185. 
paragvittata Thompson 1957:98. 
paralevigata Burla 1956:261. 
paramediostriata Townsend & Wheeler 1955: 

62. 



paramelanica Patterson 1942b: 12, as subsp. of 

melanica; should be consid. sp. 
paranaensis de Barros 1950:266. 
parapunctipennis Duda 1924a:205. Nom. nud. 

in Duda 1923:44 as var. of punctipennis. 
pararepleta Dobzhansky & Pavan 1943:52. 

Now consid. subsp. of mercatorum. 
parasaltans Magalhaes 1956:276. 
paravibrissina Duda 1924a:218, & 1924b:248. 
parenli Villeneuve 1921:157. Syn. of littoralis 

Meigen. 
parlhenogenetica Stalker 1953:347. 
parva Grimshaw 1901:65. 
paitersoni Pipkin 1956:251. 
paucilineata Burla 1957:38. 
pavicipimclata Grimshaw 1901:62. 
paulista Dobzhansky & Pavan 1943:10. Syn. 

of willistoni Sturtevant. 
paulistorum Dobzhansky & Pavan 1949:301. 
pavani Brncic 195 7a: 88. 
pectinipes Duda 1927:177. Manuscript name, 

prob. not intended for use in nomenclature. 
pengi Okada & Kurokawa 1957:11. Equals 

"melanissima'' of Kikkawa & Peng 1938. 
peninsularis Patterson & Wheeler 1942:92. 
perkinsi Grimshaw 1901:59. 
persimilis Dobzhansky & Epling 1944:7. 
peruensis Wheeler 1959, antea. N. n. for 

maculipennis Duda, not Gimmerthal. 
peruviana Duda 1927:204. 
phalerata Meigen 1830:83. 
picta Zetterstedt 1847:2567: (As first pub- 

blished, was credited to "Staeg. in litteris'"). 
picticornis Grimshaw 1901:57. 
piclifrons Duda 1927:182 
pictipennis Kertesz 1901:421. 
pictipes de Meiiere 1911:411. To Styloptera 

(Duda 1924a: 192); to Dettopsomyia (Duda 

1926a:61). 
pictivenlrls Duda 1925:211. 
pieUila de Meiiere 1911:412. To Paramycodro- 

sophila. 
pilicrus Duda 1926a: 71, 74. 
pilifacies Malloch 1926:29. 
pilimana Grimshaw 1901:61. 
pilosiila Becker 1908:156. Duda (1935a) con- 
sid. syn. of fasciata Meigen. 
pinguis Walker 1865:128. To Lauxaniidae. 
pinicola Sturtevant 1942:40. 
plagiata Bezzi 1908a: 197. 
platilarsus Frota-Pessoa 1954:276. 
pleviralis Williston 1896:411. Has been con- 
sid. Mycodrosophila^ but is prob. Drosophila, 

subgen. Hirtodrosophila. 
pleurofasciata Duda 1924a:213. Syn. of picta 

Zetterstedt. 



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pleurovittata Harrison 1954:108. 

plumosa Grimshaw 1901:72. 

plurilineata Villeneuve 1911:83. Syn. of 
busckii Coquillett. 

poecila Burla & Pavan 1953:311. N. n. for, and 
syn. of, Paramycodrosophila poeciloptera 
Duda 1925; see remarks antea. 

poecilithorax Malloch 1925:87. 

poecilogastra Duda 1926a: 65, as var. of lati- 
frons Duda. 

poeciloptera (Duda 1925:226, in Paramyco- 
drosophila). 

poeciloptera Duda 1940:26, 34. Preocc; re- 
placement name is punctatonervosa Frey. 

poeyi Sturt3vant 1921:76. To Zygothrica. 

pokornyi Duda 1924a: 218. 

polita Grimshaw 1901:71. 

poUinosa Williston 1896:414. To Ephydridae. 

poUinospadix Patterson & Mainland 1944:55. 

polychaeta Patterson & Wheeler 1942: 102. 

polymorpha Dobzhansky & Pavan 1943:19. 

polypori Malloch 1924b: 351. 

ponderosa Patterson & Mainland 1943: 198. 

prashadi Brunetti 1923:303. 

preciosa de Meijere 1911:410. To Pictostyl- 
optera (Duda 1924a: 192); to Dettopsomyia 
(Duda 1926a: 61). 

procardinoides Frydenberg 1956:60. 

procnemis Williston 1896:412. To Chymo- 
myza. 

prognatha Sturtevant 1916: 340. 

projectans Sturtevant 1916:342. To Mycodro- 
sophila. 

prorepleia Duda 1925:210, as var. of repleta 
Wollaston. 

prosaltans Duda 1927:164. 

prosimilis Duda 1927:194, as a form, not va- 
riety, of similis Williston ("/c/z bezeichne 
eine Form vorldufig ah prosimilis"); later 
treated as sp. by Dobzhansky & Pavan 1943: 
23. 

proxima Adams 1905:186. To Leucophenga. 

pruinifacies Frota-Pessoa 1954:292. 

pruinosa Duda 1940:27, 41 

pseudomelanica Sturtevant 1916:333. Is prob. 
putrida Sturtevant, melanistic form. 

pseudoobscura Frolova 1929:212. 

pseudosaltans Magalhaes 1956:273. 

pseudotakahashii Mather 1957:222. 

pugionata de Meijere 1915a: 56. 

pulchella Sturtevant 1916:327. N. n. for bel- 
lula Williston, not Bergroth. 

pulchra Schiner 1868:239. To Leucophenga 
(Basden, pers. comm.). 

pulchrella Tan, Hsu, & Sheng 1949:198. 



pulla Pavan & da Cunha 1947:10 Syn. of 
guarajd King. 

puUata Tan, Hsu, & Sheng 1949:200. 

pulvera Duda 1927:181. {pulverea is an in- 
valid orig. spelling). 

pumilio de Meijere 1908: 153. 

punalua Bryan 1934:438. 

punctatonervosa Frey 1954:32. Prob. syn. of 
poeciloptera Duda 1940; the latter being 
preocc, this is an available replacement 
name. 

puncticeps Okada 1956:94. 

punctipennis (van der Wulp 1886:56, in Dis- 
comyza) . Syn. of lurida Walker. 

punctipennis Duda 1940:24, 29, as var. of bi- 
color; syn. of Lissocephala unipuncta Mal- 
loch (Burla 1954a). 

punctiscutata Lamb 1914:333. Poss. to Neo- 
tanygastrella (Burla 1954a). 

piinctvdata Loew 1862:232. Syn. of repleta 
Wollaston. 

pusilla Grimshaw 1901:70. 

pusio Duda 1923:50. 

putrida Sturtevant 1916:339. 

pygmaea Duda 1926a: 94, 102. 

pygmaea Duda 1927:125, as var. of repleta 
Wollaston. 

quadrata Sturtevant 1916:341. To Microdro- 
sophila. 

quadrilineata de Meijere 1911:396. To Chae- 
todrosophilella. 

quadrimaculata Walker 1852:410. To Leuco- 
phenga. 

quadrimaculata Adams 1905:182. Preocc; n. 
n. is adamsi Wheeler. 

quadripunctata de Meijere 1908:154. To Leu- 
cophenga. 

quadriradiata Duda 1923:46. 

quadriseriata Duda 1924a:211, & 1924b:255. 

quadrivittata Okada 1956:83. 

quadrum (Wiedemann 1830:507, in Trypeta). 

quinaria Loew 1865:182. 

quinqueannulata Frey 1917:31. 

racemova Patterson & Mainland 1944:44. 

ramsdeni Sturtevant 1916:328. 

ramulosa Burla 1956:266. 

reamurii Dufour 1845:201. Sp. inc. 

rectangularis Sturtevant 1942:38. 

remota Walker 1849:1111. Prob. not Dro- 
sophila. 

repleta Wollaston 1858:117. 

reticulata Wheeler 1957:101. 

rioensis Patterson 1943:152, as subsp. of meri- 
diana. 

ritae Patterson & Wheeler 1942:87. 

robusta Sturtevant 1916:331. 



Wheeler: Nomenclature of Drosophila 



197 



rostrata Duda 1925:219. 

ruberrima de Meijere 1911:403. 

rubidifrons Patterson & Mainland 1944:48. 

rubra Sturtevant 1927:369. 

rubrifrons Patterson & Wheeler 1942:107. 

rubrostriata Becker 1908:155. Syn. of busckii 
Coquillett. 

rudis Walker 1860:168. To Leucophenga. 

rufa Kikkawa & Peng 1938:529. 

ruficeps von Roser 1840:62. Sp. inc., poss. to 
Scaptomyza. 

rufifrons Loew 1873:50. 

rufipes Meigen 1830:87. Sp. inc., prob. to 
Scaptomyza. 

rufviloventer Lamb 1914:344. 

saba Burla 1954a: 141. 

sadleria Bryan 1938:41. 

salatigae de Meijere 1914:260. To Leuco- 
phenga. 

saltans Sturtevant 1916:328. 

samoaensis Harrison 1954:103. 

sanyi Burla 1954a: 113. 

scaptomyzoptera Duda 1935a: 95. 

schildiMalloch 1924a: 10. 

schmidti Duda 1924a: 21 3. 

scioptera Duda 1927:179. 

scutellaris Duda 1929:420. 

scutellata Duda 1926a: 65, 70, as var. of den- 
tata Duda 1924. 

scvitellimargo Duda 1924a: 206, & 1924b: 243. 
Nom. nud. in Duda 1923:43; consid. by 
Duda 1926a as var. of hrunnea de Meijere. 

segiiyi Smart 1945:56. N. n. for subobscura 
Seguy, not Collin. 

sellata Sturtevant 1942:39. Consid. syn. of pro- 
saltans Duda but consid. prob. valid sp. 
by Magalhaes & Bjornberg 1957:448; prob. 
syn. of saltans St. 

semialba Duda 1925:208. 

semiatra de Meijere 1914:265. 

semiatricornis Duda 1934:63, and 1935a: 69, as 
var. of graminum in Scaptomyza which was 
consid. subg. of Drosophila. To Scaptomyza. 

seminigra Duda 1926a: 65, 68, as var. of lati- 
frons Duda; consid. sp. by Malloch 1934a. 

Seminole Sturtevant & Dobzhansky 1936:577. 

senilis Duda 1926a: 91. Type of subg. Macro- 
palpus. 

senufo Burla 1954a: 137. 

separata de Meijere 1911:406. 

serenensis Brncic 1957a: 78. 

sericea Lamb 1914:326. To Leucophenga. 

serrata Malloch 1927:6. 

setapex Patterson & Mainland 1944:55. 

setifemur Malloch 1924b: 351. 

setiger Grimshaw 1901:64. 



setosa Villeneuve 1921:158. Syn. of testacea 

von Roser. 
setosa Dobzhansky & Pavan 1943:46. Preocc; 

syn. of hydei Sturtevant. 
setula Heed & Wheeler 1957: 18. 
sexlineata Duda 1940:26, 36, as var. of 

quadrimaculata Adams. 
sexpunctata Seguy 1938:352. 
sexvittata Okada 1956:78. 
sharpi Grimshaw 1901:65. 
sigmoides Loew 1872:103. Type of subg. 

Siphlodora. 
signata Duda 1923:48. 
silvaia de Meijere 1916:206. 
silvestris Basden 1954:618. 
similis Williston 1896:415. 
similis Lamb 1914:347. Preocc; n. n. is errans 

Malloch; syn. of ananassae Doleschall. 
simplex de Meijere 1914:266. 
simulans Sturtevant 1919:153. 
singularis Duda 1924a: 220, & 1924b: 249. 

Nom. nud. in Duda 1923:56. 
sogo Burla 1954a: 198. 
solennis Walker 1860:168. 
sordida Zetterstedt 1838:777. To Scaptomyza. 
sordidapex Grimshaw 1901:63. 
sordidula Kikkawa & Peng 1938:539. 
soror Schiner 1868:240. Not Drosophilidae 

(Basden, pers. comm.). 
sororia Williston 1896:408. 
spadicifrons Patterson & Mainland 1944:49. 
spenceri Patterson 1943:160. 
sphaerocera Thomson 1869:596. To Heleomy- 

zidae (Malloch 1934b:437); to Drosophili- 
dae, genus uncertain (Sabrosky, pers. 

comm.). 
spinatermina Heed & Wheeler 1957:30. 
spinicauda Malloch 1926:30. 
spinipes Lamb 1914:336. 
spino femora Patterson & Wheeler 1942:104. 

Poss. syn. of nasuta Lamb. 
splendida Williston 1896:412. To Clastoptero- 

"^via. 
spurca Zetterstedt 1847:2550. Syn. of tristis 

Fallen (Frydenberg 1955:110). 
stackelbergi Duda 1935a: 96. 
stalkeri Wheeler 1954:52. 

sternopleuralis Okada & Kurokawa 1957:6. 
sticta Wheeler 1957:96. 
stonei Pipkin 1956:254. 
striaticeps Duda 1923:58. 
strigifrons de Meijere 1914:264. 
strigiventris Duda 1927:184. 
sturtevanti Duda 1927:167. 
subacuticornis Duda 1924a: 207, & 1924b: 244. 
subbadia Patterson & Mainland 1943:198. 



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subfasciata de Meijere 1914:257. 

subflavohalterata Buria 1956:264. 

subfunebris Stalker & Spencer 1939:108. 

subgilva Burla 1956:263. 

subinfumata Duda 1925:221. 

suhlineata Duda 1926a: 65, 69, as var. of lati- 

frons Duda. 
submacroptera Patterson & Mainland 1943: 

186. 
submelanica Patterson 1942a: 10. Manuscript 

name for melanica Sturtevant. 
subnitida Malloch 1927:5. 
subobscura Collin 1936:25. 
subobscura Segiiy 1938:352. Preocc; n. n. is 

seguyi Smart. 
suboccidentalis Spencer 1942:61. 
subpalustris Spencer 1942:64. 
subpollinosa de Meijere 1914:263. To Leuco- 

phenga. 
subquinaria Spencer 1942:59. 
subsaltans Magalhaes 1956:277. 
subsigmoides Patterson & Mainland 1944:26. 

Syn. of flexa Loew. 
subsplendens Duda 1934:70. Descr. in Scapto- 

myza which was consid. subg. of Drosophila. 

To Scaptomyza. 
subtilis Kikkawa & Peng 1938:541. 
subviridis Patterson & Mainland 1943:140. 
sucinea Patterson & Mainland 1944:31. 
suflfusca Spencer 1943:92. 
sulcata Sturtevant 1916:330. Syn. oi color ata 

Walker. 
sulfurigaster Duda 1923:48. Prob. syn. of 

nasuta Lamb. 
suma Burla 1954a: 200. 
sumatrensis Duda 1926a: 73, 79. 
superba Sturtevant 1916:342. To Clastoptero- 

myia. 
suruku Burla 1954a: 107. 
siituralis Wheeler 1957:100. 
suzukii (Matsumura 1931:366, in Leuco- 

phenga) . 
sydneyensis Malloch 1927:5. 
takahashii Sturtevant 1927:371. 
tarsalis Walker 1852:412. 
tarsata Schiner 1868:240. 
teclifrons de Meijere 1914:263. To Oxysty- 

loptera. 
tendata Wheeler 1959, antea. N. n. for dentata 

Duda 1927, not 1924. 
tenebrosa Spencer 1943:93. 
tenuicauda Okada 1956:141. 
tenuipes (Walker 1849:1112, in Diastata) . 
terminalis Loew 1863:32. To Scaptomyza. 
testacea von Roser 1840:62. 



texana Patterson 1940:219, as subsp. of virilis; 

now consid. subsp. of americana Spencer. 
thienemanni Duda 1931:196. 
thoracis Williston 1896:411. 
tibialis Wheeler 1957:100. 
tibudu Burla 1954a: 194. 
tigrina Buzzati-Traverso 1943:8. Syn. of buz- 

zatii Patterson & Wheeler. 
tjibodas de Meijere 1916:205. 
tolteca Patterson & Mainland 1944:32. 
torrei Sturtevant 1921:86. 
tranquilla Spencer 1943:200. 
transversa Fallen 1823:6. 
trapeza Heed & Wheeler 1957:25. 
trapezina Duda 1923:41. 
triangula Wheeler 1949b: 192. 
triangulifer Lamb 1914:343. 
triangulina Duda 1927:186. 
trichiaspis Duda 1940:23, 31. 
trifasciata de Meijere 1916:206. 
trifiloides Wheeler 1957:81. 
trifilum Frota-Pessoa 1954:292. 
trilimbata Bezzi 1928:158. 
tripunctata Loew 1862:231. 
triseta de Meijere 1911:402. 
trispina Wheeler 1949b: 180. 
tristani Sturtevant 1921:75. 
tristipennis Duda 1924a:215, & 1924b:247. 

Nom. nud. in Duda 1923:53. 
tristipes Duda 1924a: 220, & 1924b: 25 7. 
tristis Fallen 1823:7. 
tristriata Heed & Wheeler 1957:31. 
trivittata Strobl 1893a: 282. 
tropicalis Burla & da Cunha 1949:302. 
Isigana Burla & Gloor 1952: 164. 
tuchaua Pavan 1950:26. 

tumiditarsus Tan, Hsu, & Sheng 1949:205. 
uebe Burla 1954a: 148. 
umbripennis Hendel 1936:99. 
undulata Grimshaw 1901:58. 
ungarensis de Meijere 1911:407. 
unicolor de Meijere 1914:266. 
unicolor (Malloch 1934a: 293, in Hirtodro- 

sophila) . Preocc; n. n. is unicolorata Wheel- 
er. 
unicolorata Wheeler 1959, antea. N. n. for 

unicolor (Malloch), not de Meijere. 
unimaculata Strobl 1893a: 281. 
uninubes Patterson & Mainland 1943:201. 
unipectinata Duda 1924a:215, & 1924b:246. 
unipunctata Patterson & Mainland 1943:182. 
unispina Okada 1956:129. 
unistriata Strobl 1898a: 580 (usual citation is 

1900:636; see references). 
upoluae Malloch 1934a: 305. 



Wheeler: Nomenclature of Drosophila 199 

ussurica Duda 1935a: 74, 98, as var. of trivit- vina Burla 1954a: 112. 

tata Strobl. viracochi Brncic & Santibanez 1957:70. 

ustulatadeMeijere 1908: 157. virgata Tan, Hsu, & Sheng 1949:203. Poss. 

uvarum Rondani 1875:86. Consid. syn. of s,yn. oi annulipes Buda {Okada) . 

melanogaster Meigen. virginea Meigen 1830:84. Syn. of fenestra- 

valida Walker 1858:232. To Lauxaniidae. rum Fallen. 

varia Walker 1849:1109. To Leucophenga. virilis Sturtevant 1916:330. 

variegata Fallen 1823:5. To Amiota, subg. vittata Coquillett 1895:318. To Parascapto- 

Phortica. myza. 

variegata Grimshaw 1901.57. Preocc; n. n. is vittatifrons Williston 1896:408. To Zygoth- 

grimshawi Oldenberg. rica. 

varifrons Grimshaw 1901:71. wheeleri Patterson & Alexander 1952:129. 

variopicta Becker 1908:156. Consid. syn. of willistoni Sturtevant 1916:327. N. n. for pal- 

fenestrarum Fallen, but Basden (pers. lida Williston, not Zetterstedt. 

comm.) consid. it valid sp. willowsi Curran 1936:42. 

varipes Macquart 1835:550. Sp. inc., prob. to xanthogaster Duda 1924a: 21 7, & 1924b: 248. 

Camillidae (Duda 1935a). xanthopyga Duda 1924a:215, as var. of mon- 

versicolor Mather 1955:573. Syn. of buzzatii tium deMeijere. 

Patterson & Wheeler. xanthosoma Grimshaw 1901:68. 

verticis Williston 1896:413. Sp. inc. yakuba Burla 1954a: 161. 

vibrissina Duda 1924a:219. Syn. of confusa yucatanensis Spencer 1940a: 160, as subsp. of 

Staeger. hydei Sturtevant. 

victoria Sturtevant 1942:33. Type of subg. zebrina Bezzi 1928: 157. 

Pholadoris. z-notata Bryan 1934:437. 

SUMMARY 

Eight new names are proposed to replace rejected junior homonyms in the 
genus Drosophila, and an annotated list of all known species, subspecies and 
varieties is presented. There are 1,024 names in the list, credited to 119 authors. 
At the time of writing, 920 species have been described in the genus, not includ- 
ing the 16 subspecies and 27 varieties. Of the 920 proposed species, 1 1 1 have been 
transferred to other genera, about 15 more should probably be transferred, about 
70 are names of synonyms (including those listed as possible or probable syno- 
nyms), and 25 are new names proposed for rejected homonyms. Of the 23 species 
listed as transfers from other genera, only 13 remain as valid Drosophila species. 
There have been some changes from subspecific to specific status, and vice versa, 
and it is evident that several of the proposed varieties should probably be con- 
sidered as valid species. Only a single fossil species has been described. 

The list contains, therefore, the names of approximately 750 presumably 
biologically valid species. Only two women have been honored by having species 
named for them, while 87 men have been so honored. The single largest con- 
tributor to the list is Dr. O. Duda who was responsible for 16% of the proposed 
names, while 14.5% are due to the efforts of Prof. Patterson, his students and 
colleagues at the University of Texas. 

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Genetic Studies of Irradiated Natural Populations of Drosophila. 

III. Experimental Populations of Drosophila ananassae Derived 
from Irradiated Natural Populations 

THOMAS G. GREGg' 

Genetics Foundation, Department of Zoology 
University of Texas 



INTRODUCTION 

With the advent of large scale nuclear weapons tests in the Marshall Islands 
it became feasible to study the effects of radiation on natural populations of 
Drosophila. The first studies of this type were reported by Stone et al. (1957). 
and by Stone and Wilson (1958). In these studies, populations of Drosophila 
ananassae from some of the Marshall Islands and Eastern Carolines, which had 
been subjected to varying amounts of direct and fallout irradiation, were ana- 
lyzed with respect to radiation effects. 

The present study is an outgrowth of these earlier studies, and is designed to 
determine the changes in the evolutionary fitness of certain of the Marshall 
Island populations of Drosophila ananassae as measured by their response to 
different test conditions in the laboratory. Changes in viability, fertility, and 
fecundity of the populations were used as measures of changes in evolutionary 
fitness. Viability is defined as the percentage of eggs that develop into adults, 
fertility as the percentage of matings yielding offspring, and fecundity as the 
average number of eggs laid per day per female. 

Offspring from flies that were collected in the summers of 1956 and 1957 were 
subjected to two different types of experimental conditions subsequent to each 
summer's collection. One type of condition involved maintaining the offspring of 
the collected flies in population cages containing from 2,000 to 10,000 adults and 
in which severe competition prevailed. The second type of condition involved 
maintaining the offspring of the collected flies in groups of pair matings in such a 
way that brother-sister inbreeding did not occur. Thus the pair-mated popula- 
tions resembled the breeding structure of an "isolate" of a human population. 
The viability, fertility, and fecundity of both the population cages and pair- 
mated populations were checked periodically. Changes in these factors were 
observed and comparisons were made with populations which had received 
different amounts of radiation. Rates of change in the cage populations were 
compared to rates of change in the pair-mated populations and changes in both 
types of experimental populations were compared to changes in the original 
populations remaining in the islands. 

1 Present address: NIH Postdoctoral Fellow, Department of G2netics, University of Wisconsin, 
Madison, Wisconsin. 



208 The University of Texas Publication 

PROCEDURE 

In the population cage tests flies from the Bikini and Majuro Atolls were used. 
The Bikini flies were used for the experimental population since the Bikini Atoll 
received more radiation than any of the other Atolls and the Majuro flies were 
used for the control population since the Majuro Atoll received very little 
radiation. Population cages were started in the following way. After each year's 
collection, females that had been fertilized in nature were raised in individual 
culture vials. F^ from these females were transferred to half pint bottles, taking 
care to prevent the more fertile females from contributing disproportionate 
numbers of offspring. Several thousand of the Fo were then transferred to the 
population cages. The population cages used in this experiment were essentially 
boxes screened on four sides with glass tops and wooden bottoms containing 15 
evenly spaced holes. Each hole was fitted with a large cork which had a food cup 
fastened on top. (For a detailed description of the population cage, see Wright 
and Dobzhansky, 1946.) The 15 food cups were changed at the rate of seven per 
week so that no cup remained in a cage more than 15 days. Duplicate cages were 
set up both years so that two cages of Majuro flies and two of Bikini were tested 
each year. With duplicate cages each cage could serve as a control for the other 
and also as insurance against failure of the experiment through the loss or con- 
tamination of one cage. 

In the tests involving groups of pair matings, flies from the Bikini and 
Rongerik Atolls were used in 1956 and flies from Bikini and Rongelap in 1957. 
The substitution of Rongelap for Rongerik in 1957 was the result of an inade- 
quate collection at Rongerik in that year. The Bikini Atoll, as previously men- 
tioned, received a large amount of radiation and Rongerik and Rongelap. by 
comparison, received intermediate amounts. Each pair-mated population was 
started by making 500 F^ pair matings using the progeny from the flies collected 
on Bikini, Rongelap, or Rongerik. Usually between one-half and one-fourth of the 
500 matings were fertile. A few progeny were taken from each fertile mating and 
used to make 500 matings for the next generation. Matings were made in such a 
way that brother-sister inbreeding did not occur. This procedure was followed in 
establishing each succeeding generation. Strictly speaking, there was no control 
for these particular tests but that did not prevent comparisons from being made 
between the populations in this type test and also between the populations in 
these tests and populations in other tests. 

Both the cage populations and the pair-mated populations were maintained at 
77 degrees F.. which approximates the mean daily temperature in the Marshall 
Islands. Each experimental population was sampled at the end of three months 
and again at the end of six months. 

To sample the population cages two food cups were introduced into each cage 
and allowed to remain 48 hours. They were then removed and the contents were 
distributed among several half pint bottles containing supplementary food. 
Larval competition was reduced in this manner in order to retain as many 
genotypes as possible. A number of offspring, selected at random from the half 
pint bottles were then put up in pair matings. About 150 such matings were 
made from which 60 were selected at random for further use. From the offspring 



Gregg: Experimental Populations 209 

of each of the 60 pair matings four brother-sister pair matings were made. A 
similar number of offspring from each of the pair matings were crossbred to 
offspring of the other 59 pair matings. These crossbred matings were also put up 
in pairs and were made in such a way that no two groups of siblings were mated 
to each other. In practice only about 40 of the original 60 pair matings produced 
enough offspring to make both kinds of matings. Consequently the offspring from 
half of the unprohfic parents were inbred in brother-sister matings while off- 
spring from the other half were crossbred in order to have the unprolific parents 
contribute equally to the inbred and crossbred matings. The inbred and crossbred 
matings, when analyzed, showed the reproductive capacities of the population at 
their worst and best respectively. 

From the above procedure 120-160 inbred matings and a like number of cross- 
bred matings were obtained. All inbred and crossbred matings were made with 
flies that had been aged for six to seven days. After the matings had been made 
the flies were given 24 hours in which to mate and then were transferred to 
fresh food every 24 hours for four days. The eggs laid each day were counted and 
the number recorded so that when the aduhs emerged the percentage of eggs 
that developed into viable adults could be determined. First day egg counts are 
often omitted from the data so as to be sure that egg counts were made on 
inseminated females only. The matings were not discarded after the four day 
egg counting period but were kept for an additional ten days without being 
transferred. This additional ten day period allow^ed many matings to produce 
offspring when they had produced none during the four day egg laying period. 
Thus a more accurate estimate of the fertility of the population could be made. 

To sample the pair-mated populations 60 fertile vials were selected at random 
from the population. There 60 were inbred and crossbred in the same way as 
the 60 fertile vials derived from the population cages and analyzed in the same 
way. The remaining fertile vials were used to continue the population, 

RESULTS 

The data from the crossbred and inbred test matings from the various popula- 
tions are shown in Table 1 through 4 and in Figures 1 through 12. In Tables 1 
and 2 the first column shows the total number of matings on which egg counts 
were made, and the second column shows the percentages of the matings which 
produced no offspring. The third column gives the number of matings that 
produced offspring. Columns four and five deal with the number of lethals in 
the population. 

The fourth column gives the percentage of matings in each test in which 
less than 80 per cent of the eggs developed. Theoretically, in an inbred mating, 
if both members were heterozygous for a recessive lethal which they had re- 
ceived from their parents, 25 per cent of the eggs produced by this mating, 
neglecting other factors, would be homozygous lethal and would not develop. 
Thus such a mating would fall into the 75 per cent egg development class. 
However, if this mating falls in the 75 per cent egg development class it cannot 
be distinguished, in the present experiment, whether a single recessive lethal 
has become homozygous or whether several genes have become homozygous and 



210 



The University of Texas Publication 



KEY TO TABLES 1 AND 2 

Column 1 gives the total number of matings made in each test for the various populations. 

Column 2 gives the percentage of the matings in column 1 which were sterile. 

Column 3 gives the number of matings in column 1 which were fertile. 

Column 4 gives the percentage of the matings shown in column 3 that carry at least one lethal 

or lethal equivalent — that is, fall in an egg development class below 80 per cent. 
Column 5 gives the percentage of parent matings that carry a lethal as determined by the fact 

that at least one of the four brother-sister matings made from the parent mating carries a 

lethal. 
Column 6 gives the total number of days on which eggs were counted in each test. 
Column 7 gives the total number of eggs laid in each test. 
Column 8 gives the averago number of eggs laid per day per female and is obtained by dividing 

column 7 by column 6. 
Column 9 gives the actual per cent of the eggs given in column 7 that developed into adults. 
Column 10 gives the mean per cent of egg development per female and is based on angular 

transformations. 

Table 1. 1956 Tests. 



Type of population 
and type of test 



10 



Bikini pair-mated 
Sample 1 inbred 
Sample 2 inbred 
Sample 1 crossbred 
Sample 2 crossbred 

Rongerik pair-mated 
Sample 1 inbred 
Sample 2 inbred 
Sample 1 crossbred 
Sample 2 crossbred 

Bikini cage 1 

Sample 1 inbred 
Sample 2 inbred 
Sample 1 crossbred 
Sample 2 crossbred 

Bikini cage 2 

Sample 1 inbred 
Sample 2 inbred 
Sample 1 crossbred 
Sample 2 crossbred 

Majuro cage 1 
Sample 1 inbred 
Sample 2 inbred 
Sample 1 crossbred 
Sample 2 crossbred 

Majuro cage 2 
Sample 1 inbred 
Sample 1 crossbred 



157 


12.7 


136 


77.5 




464 


13243 


28.5 


60.1 


63 


101 


31.7 


69 


74.0 


92.1 


226 


4735 


21.0 


64.2 


66 


168 


14.7 


141 


44.6 




389 


11148 


28.7 


77.3 


79 


121 


22.3 


86 


46.5 




239 


4830 


20.2 


75.0 


79 


205 


30.7 


132 


75.0 


92.0 


342 


9107 


26.6 


64.0 


66 


99 


32.3 


58 


76.0 


90.9 


135 


4501 


33.3 


60.5 


60 


188 


25.0 


112 


39.3 




274 


7645 


27.9 


79.5 


84 


136 


44.9 


61 


47.5 




161 


3505 


21.8 


75.3 


80 


156 


7.0 


139 


85.6 


100 


362 


7567 


20.9 


57.4 


57 


119 


16.7 


95 


82.0 


100 


216 


7585 


35.1 


59.7 


57 


156 


8.3 


99 


54.4 




376 


4617 


12.3 


72.0 


77 


121 


7.4 


104 


36.5 




274 


10243 


37.4 


81.7 


83 


191 


13.6 


159 


73.0 


97.9 


365 


10957 


30.0 


64.2 


66 


164 


22.0 


102 


71.6 


90.2 


270 


6600 


24.4 


60.9 


64 


169 


23.7 


124 


37.8 




309 


7834 


25.4 


82.8 


80 


130 


13.1 


79 


53.2 




205 


3665 


17.9 


70.3 


75 


160 


27.5 


116 


67.2 


95.0 


316 


8166 


25.8 


64.9 


69 


126 


20.6 


88 


76.1 


97.1 


219 


7929 


36.2 


61.0 


64 


182 


27.5 


99 


35.4 




291 


4849 


16.7 


78.0 


84 


118 


14.4 


94 


24.5 




245 


8134 


33.2 


87.8 


91 


161 


15.0 


131 


84.8 


93.4 


340 


9476 


27.9 


64.8 


65 


120 


15.0 


102 


18.6 




272 


7245 


26.6 


89.1 


91 



that the cumulative effect of these several genes, when all are present and 
homozygous, is lethal. The latter case is called a lethal equivalent. It sometimes 
happens however that a recessive lethal, and especially a lethal equivalent, may 
be "leaky" or not completely lethal even when homozygous. In that case a 
mating in which each fly was heterozygous for a "leaky" lethal would fall in an 
egg development class somewhat higher than 75 per cent, perhaps as high as 78 
or even 80 per cent. Further, it was shown by Stone et al. (1957) that even with 



Gregg: Experimental Populations 



211 



extensive crossbreeding of different populations the viability of many matings 
fell as low as 80 per cent in egg development. Thus it can be assumed that there 
is a general level of subvital factors in all of these populations that normally 
reduces viability as much as 20 per cent. Such subvital factors are often reces- 
sive lethals that are not completely recessive and, as the name implies, are not 
fully viable when present as a heterozygote. Each subvital may then reduce the 
viability by a few per cent. If there are normally enough subvitals present to 
reduce the viability by 20 per cent, then the reduction in viability of any mating 
by more than 20 per cent could easily enough be due to the presence of a reces- 
sive lethal acting as a subvital that is in excess of the normal number of sub- 

KEY TO FIGURES 1 THROUGH 12 

The abscissa in the following figures is divided into five egg development classes. The classes 
batween 1 per cent and 19 per cent are plotted at 10 per cent, those between 20 and 39 per cent 
are plotted at 30 per cent, those between 4 and 59 per cent are plotted at 50 per cent, those 
between 60 and 79 per cent are plotted at 70 per cent, and the classes between 60 and 100 per cent 
are plotted at 90 per cent. 

The ordinate shows the percentage of matings in the particular tests that fell in each of the 
five egg d2vclopment classes. 

The points of these graphs would be the midpoints of the tops of a five bar histogram if the 
histogram were drawn in. 

Matings falling in the 80-100 per cent class are considered normal, those in the 60-80 per 
cent class are considered to have one or two lethals or lethal equivalents, 40-60 per cent class to 
have two or three, the 20-40 per cent class to have four or five, and the 1-20 per cent class has six 
or more lethals or lethal equivalents. 




70 90 10 30 

7o EGG DEVELOPMENT 



212 The University of Texas Publication 

Table 2. 1957 Tests. 



Type of population 






















and type of test 
























1 


2 


3 


4 


5 


6 


7 


8 


9 


10 


Bikini pair-mated 






















Sample 1 inbred 


148 


20.9 


109 


87.2 


97.5 


328 


10887 


33.2 


48.3 


48 


Sample 2 inbred 


150 


10.7 


117 


81.2 


97.6 


239 


4592 


19.2 


54.0 


57 


Sample 1 crossbred 


149 


20.8 


108 


36.1 




271 


6662 


24.6 


77.5 


81 


Sample 2 crossbred 


138 


14.5 


108 


38.9 




279 


4763 


17.1 


79.2 


82 


Rongelap pair-mated 






















Sample 1 inbred 


157 


10.6 


108 


76.9 


95.0 


308 


6303 


20.5 


58.6 


61 


Sample 2 inbred 


146 


25.3 


87 


49.4 


73.5 


259 


2643 


10.2 


73.3 


74 


Sample 1 crossbred 


142 


28.9 


89 


56.1 




251 


3414 


13.6 


70.3 


75 


Sample 2 crossbred 


143 


21.7 


94 


31.9 




238 


3943 


16.6 


82.7 


87 


Bikini cage 1 






















Sample 2 inbred 


149 


7.3 


126 


83.3 


100 


309 


8176 


26.4 


59.4 


63 


Sample 1 crossbred 


168 


16.7 


102 


47.1 




282 


3605 


12.8 


74.5 


36 


Sample 2 crossbred 


133 


12.0 


111 


18.0 




298 


6255 


20.9 


86.2 


89 


Bikini cage 2 






















Sample 1 inbred 


148 


17.6 


101 


89.1 


92.3 


268 


3826 


14.3 


56.7 


55 


Sample 2 inbred 


142 


16.2 


95 


85.3 


100 


219 


3026 


13.8 


52.2 


57 


Sample 1 crossbred 


142 


29.6 


82 


54.9 




234 


2733 


11.7 


73.8 


77 


Sample 2 crossbred 


124 


22.6 


83 


45.7 




199 


2777 


14.0 


77.2 


85 


Majuro cage 1 






















Sample 1 inbred 


110 


17.3 


55 


70.9 


77.8 


176 


2940 


16.7 


63.4 


63 


Sample 2 inbred 


128 


36.9 


58 


84.4 


89.7 


155 


1543 


10.0 


58.5 


62 


Sample 1 crossbred 


128 


36.0 


56 


66.1 




135 


2495 


18.5 


62.6 


67 


Sample 2 crossbred 


127 


32.3 


65 


40.0 




151 


1446 


9.6 


76.8 


83 



vitals present. In that case the mating is said to carry a recessive lethal or lethal 
equivalent, even though both flies in the mating are not heterozygous for it. 
Lowered viability caused by a subvital lethal would be noticed more frequently 
in the crossbred matings than in the inbred matings because in the inbred mat- 
ings such an effect would usually be masked by a more drastic reduction in 
viability due to the recessive lethals becoming homozygous. Therefore, the 
dividing line between the matings considered to be lethal free and those carrying 
a lethal has been placed (Stone et al. 1957) at 80 per cent, somewhat arbitrarily, 
but not without reason. The percentage of matings carrying at least one lethal is 
shown in column four. The analysis is further complicated by the fact that 
several of these various lethal factors may be present at once. The presence in a 
mating of the normal level of subvital factors, plus a recessive lethal, may reduce 
the viability to as low as 60 per cent. The presence of two lethals will not reduce 
the viability twice as much as one lethal because there will be some duplication 
of "killing" by each lethal. Thus each additional lethal has less effect. To aid in 
the analysis, five egg development classes were established: 80-100 per cent was 
considered normal, 60-80 per cent was considered to have one or two lethals or 
lethal equivalents, 40-60 per cent to have two or three, 20-40 per cent to have 
four or five, and 1-20 per cent to have six to eight or more. These egg develop- 
ment classes, although subject to error, are nevertheless a consistent standard 
whereby viability levels in different populations can be compared. The number 
of matings in various egg development classes for the various experimental popu- 
lations for the two years are given in Tables 3 and 4. These data are plotted for 
ease of comparison in Figures 2 to 12. Figures 1 and 2 give data from Stone et al. 
(1957) and from Stone and Wilson (1958). They show similar plots from the 



Gregg: Experimental Populations 



213 



Bikini and Majuro populations on the islands in 1956 and 1957 and have been 
included for comparison. 

Returning to the explanation of Tables 1 and 2, the fifth column shows the 
percentage of parent matings carrying at least one lethal. The frequency of 
these lethals in the P^ generation can be determined by making brother-sister 
inbred matings among the F,. If one of the four brother-sister matings falls in an 
egg development class lower than 80 per cent, at least one of the parents is 
heterozygous for a recessive lethal. It should be noted that even if all four of the 
Fi brother-sister matings fall in egg development classes above 80 per cent, there 
is still a 32 per cent chance that the parent mating carried a lethal which was 
not present in both flies in any of the brother-sister matings and consequently 
was not detected. Also significant is the fact that when fewer than four brother- 
sister matings are fertile, there is a greater chance of "missing" lethals which 
were present in the parent mating. Consequently the figures in column five 
underestimate the actual number of lethals present. Columns six, seven and eight 
show the total number of days on which eggs were counted, the total number of 
eggs laid, and the average number of eggs laid per day per female, respectively. 
The latter column shows the fecundity of the various populations and is ob- 
tained by dividing column seven by column six. Column nine gives the actual 
percentage of eggs that developed into adults and column ten gives the mean 
percentage of egg development per female. The latter column was calculated by 
converting the percentage development of each mating to the corresponding 
angular transformation, finding the angular transformation mean, and recon- 
verting this to per cent. The use of the angular transformations placed more 



||30 



Sample I 

1956 O— O Mean = 63% 

1957 •-• Mean = 48% 



FIGURE 3. 
BIKINI PAIR-MATED POPULATIONS INBRED 




Sample I 

1956 O O Mean = 79% 

1957 • • Mean = 81% 




Sample 2 

1956 0— O Mean = 66' 
I957#— #Mean=57 
39 



Sample 2 

1956 O O Mean = 79% 

1957 • • Mean = 82 % 



30 



70 90 10 30 

% EGG DEVELOPMENT 




FIGURE 4. 
BIKINI PAIR-MATED POPULATIONS CROSSBRED 



214 The University of Texas Publication 

Table 3. 1955 Tests. 
Numbers of Matings in the Various Egg Development Classes for the Various Tests 





1-9% 


10-19% 


20-29% 


30-39% 40-49% 50-59% 


60-69% 70-79% 80-89% 90-100% 


Bikini pair-mated 






















Sample 1 inbred 





3 


4 


12 


19 


27 


22 


20 


13 


17 


Sample 2 inbred 








7 


3 


8 


8 


9 


18 


9 


7 


Sample 1 crossbred 








2 


6 


2 


10 


17 


26 


40 


37 


Sample 2 crossbred 





2 





3 


4 


9 


10 


14 


16 


28 


Rongerik pair-mated 






















Sample 1 inbred 





3 


8 


9 


15 


23 


21 


18 


15 


19 


Sample 2 inbred 


1 


1 


5 


6 


7 


9 


7 


7 


9 


6 


Sample 1 crossbred 








1 


1 


6 


3 


17 


16 


31 


37 


Sample 2 crossbred 





1 





1 


1 


6 


10 


10 


15 


17 


Bikini cage 1 






















Sample 1 inbred 


3 


4 


6 


11 


25 


29 


26 


15 


14 


6 


Sample 2 inbred 


1 


1 


8 


8 


21 


11 


16 


11 


14 


3 


Sample 1 crossbred 


1 


1 


1 


6 


7 


7 


14 


19 


21 


26 


Sample 2 crossbred 














4 


7 


13 


14 


31 


35 


Bikini cage 2 






















Sample 1 inbred 





4 


4 


16 


15 


22 


30 


25 


17 


26 


Sample 2 inbred 





2 


4 


12 


12 


23 


11 


9 


15 


13 


Sample 1 crossbred 








1 


2 


6 


8 


9 


24 


38 


43 


Sample 2 crossbred 





1 


1 


1 


9 


8 


10 


12 


17 


20 


Majuro cage 1 






















Sample 1 inbred 





3 


8 


13 


10 


11 


15 


18 


10 


28 


Sample 2 inbred 








4 


8 


15 


14 


14 


12 


9 


12 


Sample 1 crossbred 


1 


2 


1 


2 


4 


8 


7 


10 


22 


42 


Sample 2 crossbred 





1 


1 








6 


5 


10 


9 


62 


Majuro cage 2 






















Sample 1 inbred 








6 


8 


32 


27 


19 


19 


9 


11 


Sample 2 inbred 






















Sample 1 crossbred 








1 


1 


1 


3 


7 


6 


14 


70 


Sample 2 crossbred 























Stress on the intermediate percentage values usually obtained from larger num- 
ber of eggs and less stress on high and low percentages usually obtained from 
smaller numbers of eggs. Therefore the errors which are frequently accumulated 
when percentages based on different number of eggs are averaged were reduced. 

The reason for including column ten was that in addition to the overall per 
cent of egg development shown in column nine, an important factor in the 
reproductive capacity of a population is the percentage of females in the popula- 
tion that fall in high egg development classes. That is, a population in which 50 
per cent of the females fall into egg development classes below 40 per cent is not 
very efficient reproductively, while a population in which 90 per cent of the 
females fall into egg development classes above 80 per cent is considerably more 
efficient. Thus the mean percentage of egg development of the individuals in a 
population shows a slightly different picture of the reproductive capacities of the 
population than the overall actual percentage of viability. The mean percentages 
shown in column ten are the same means that appear in Figures 3 to 12, since 
the figures are based on the percentage development of the individual matings 
rather than on the actual percentage development. 

In 1956 one Majuro population cage became contaminated with Drosophila 
melanogaster . This contamination was insignificant at the time of the first sample 
but became so severe that a second sample was not taken. In 1957 both Majuro 
cages were contaminated four weeks after they were started. In order to save 



Gregg: Experimental Populations 



215 



them, the flies were trapped out of the cages, etherized, and the melanogaster dis- 
carded. The food cups were put in half pint bottles to hatch and the emerging 
adults were separated and the ananassae returned to the cages. This procedure 
cleared up one cage which soon became as densely populated as the others. The 
second cage, apparently not entirely cleared of melanogaster, was recontami- 
nated and lost. The number of flies in the first cage was reduced to two or three 
thousand at the time the flies were reintroduced. Thus this cage was run through 
a population bottleneck that the others were not subjected to. This may have 
made some difference and should be kept in mind when inspecting the data. Both 
the second sample of the 1956 Majuro cage and the 1957 Majuro cage which was 
lost are omitted from the tables. Also omitted from the tables is the first sample 
of Bikini cage 1 in 1957. This sample was spoiled by bad food. 

The fertihty of the matings in these experiments showed considerable varia- 
tion as can be seen from Tables 1 and 2. The only consistent pattern in this varia- 
tion was that, as in other tests (Stone et al. 1957), the Bikini flies were con- 
sistently more fertile than any of the other populations. This high fertility was 
maintained in both types of tests. Although there is usually a difference in fer- 
tility between crossbred and inbred matings in this case the averages of both types 
of matings showed about the same degree of fertility, the crossbred being more 
fertile sometimes and the inbred at others. This variability in fertility was shown 
when second samples were compared to first samples; at times the second sample 
was more fertile and at other times the first sample was more so. Also, there was 
no evidence to indicate an improved fertihty from one year to the next. 

Such variability among the various matings is not surprising in view of the 



o 

< 

— CO 

id 

ig,o 

LJ UJ 



Sample 



1956 O OMean=66%Rongerik 




OS 

Q. 

UJQ 



RONGERIK FIGURE 5 

AND PAIR -MATED POPULATIONS INBRED 
RONGELAP 



RONGERIK FIGURES 

AND PAIR-MATED POPULATIONS CROSSBRED 

RONGELAP 



Sample 

I9560— OMean=84%Ronqerik 

1957 9— O Mean = 75 % Rongelap 




Sample 2 

9560-OMean = 80%Rongerik 
957#-#Mean = 87 7o Rongelap 



52.5 




50 



90 10 30 

EG 3 DEVELOPIVIE/JT 



70 90 



216 The University of Texas Publication 

Table 4. 1957 Tests 
Numbers of Matings in the Various Egg Development Classes for the Various Tests 





1-9% 


10-19% 


20-29% 


30-39% 


40-49 % 


50-59% 


60-69% 


70-79% 


80-89% 


90-100% 


Bikini pair-mated 






















Sample 1 inbred 


3 


11 


12 


16 


15 


15 


15 


8 


11 


3 


Sample 2 inbred 


1 


3 


5 


21 


18 


22 


10 


15 


17 


5 


Sample 1 crossbred 





2 


6 


3 


2 


7 


3 


17 


34 


34 


Sample 2 crossbred 





1 


1 


5 


5 


4 


8 


18 


32 


34 


Rongelap pair mated 






















Sample 1 inbred 





1 


6 


16 


15 


21 


18 


5 


4 


12 


Sample 2 inbred 





1 


2 


4 


6 


7 


10 


17 


15 


25 


Sample 1 crossbred 





2 


2 


2 


6 


11 


11 


16 


19 


20 


Sample 2 crossbred 








6 


2 


2 


3 


8 


15 


22 


42 


Bikini cage 1 






















Sample 1 inbred 






















Sample 2 inbred 





2 


4 


16 


14 


21 


25 


24 


12 


9 


Sample 1 crossbred 


2 


2 


4 


3 


3 


6 


15 


12 


27 


28 


Sample 2 crossbred 








1 





1 


2 


6 


11 


33 


57 


Bikini cage 2 






















Sample 1 inbred 





3 


11 


12 


16 


19 


15 


14 


7 


4 


Sample 2 inbred 





5 


13 


7 


5 


23 


9 


18 


10 


5 


Sample 1 crossbred 





1 


2 





3 


11 


14 


15 


14 


22 


Sample 2 crossbred 





1 


2 


4 


1 


5 


12 


14 


14 


30 


Maiuro cage 1 






















Sample 1 inbred 





1 


2 


10 


7 


5 


11 


3 


7 


9 


Sample 2 inbred 





1 


1 


7 


6 


13 


10 


11 


5 


4 


Sample 1 crossbred 








6 


2 


5 


8 


10 


7 


13 


7 


Sample 2 crossbred 





2 


4 


2 


2 


3 


3 


11 


14 


24 



fact that ananassae is a poor laboratory stock. In 1956 there was a considerable 
difference between the cages and the pair-mated populations in average per- 
centage of fertihty. Such a difference was not present in 1957. Also, in 1956 the 
pair-mated populations were themselves much more variable in fertility. This 
may have been due to the fact that the population size was somewhat smaller 
and more variable in size in 1956 than in 1957. The considerable increase in 
sterility between the first and second samples of the 1956 pair-mated populations 
probably accounts, at least in part, for the noticeable overall difference between 
the cages and pair-mated populations in 1956. 

A few consistent differences were apparent in the percentage of lethals present 
in the various populations. However, none of these differences were very large. 
A strong case can be made for assuming that all of the F^ matings, as shown by 
inbreeding tests, carried at least one lethal. As mentioned previously, even if all 
four bother-sister inbred matings from each of the parent matings were fertile 
and in egg development classes above 80 per cent, 32 per cent of the lethals in the 
parent matings would have been missed. Actually, enough sterility occurred so 
that almost always fewer than four and sometimes only one inbred mating was 
fertile. Consequently, considerably more than 32 per cent of the lethals carried 
by the parent matings were missed. 

There was a consistent tendency for lethals to be eliminated between the time 
of the first sample and the time of the second. This indicates that there was some 
selection against the lethals in these tests. One should also notice, however, that 
the level of lethals in these tests is higher than the level of lethals found in the 
island populations at the time of the 1956 and 1957 collections. This suggests that 



Gregg: Experimental Populations 



217 



there was an initial increase in the number of lethals in the experimental popula- 
tions. This was perhaps due to some inbreeding for a few generations which 
lasted until the population size had built up. 

Viability is closely related to the number of lethals in a population although it 
is affected by other factors as well. In these experiments the mean viability was 
divided into five classes and is plotted in Figures 1 to 12. The percentage of indi- 
viduals in each test whose egg development fell between 80 and 1 00 per cent is 
plotted at 90 per cent. The percentage of individuals in each test whose egg devel- 
opment fell between 60 and 80 per cent is plotted at 70 per cent, and so on. It can 
be seen that the points on the graphs would be the midpoints of the tops of five 
histogram bars if the histograms were drawn in. These graphs give a m.ore accu- 
rate picture of the viability and the reproductive capacities of the various popula- 
tions than one gets by merely determining the number of flies that carry at least 
one lethal. 

The differences in viability between the various populations are also shown in 
Tables 1 and 2. It can be seen in each case, with the exception of Majuro cage 1 
sample 1, 1957, that the inbred matings were significantly less viable than the 
crossbred matings. The Majuro cage had been reduced in numbers to about 
2,000 from the melanogaster contamination only a few generations before sample 
1 was taken. This probably accounts for the similarity in viability between the 
crossbred and inbred matings in that particular sample. 

The differences between the viabilities of the crossbred matings are larger than 
the moderate differences between the inbred matings. The single exception to the 
small differences between the first and second samples in the inbred matings is 




70 90 10 30 

% EGG DEVELOPMENT 



218 



The University of Texas Publication 



50-, 



Sample I 

1956 O — OMean=57 7o 



1957 Bad Food 



Sample I 

1956 O — O Mean = 667© 
" 1957 # — #Mean =55% 



30- 



LU 
O 

LU 
Q- 10 



FIGURE 9 
BIKINI CAGES I INBRED 



FIGURE 10 
BIKINI CAGES 2 INBRED 



Sample 2 

1956 O — OMean = 577c 

1957 #— # Mean =63 7o 




Sample 2 

I9560— O Meon = 647o 

1957 • • Mean = 57 % 




90 10 30 

7o EGG DEVELOPMENT 



the Rongelap 1957 population. It must be noted that this population, at the time 
of the first sample, showed much lower viability than the corresponding parent 
population collected on the island three months earlier. Therefore, the consider- 
able increase in viability between the time of the first sample and the time of the 
second sample merely restored the viability to a range comparable to that in the 
other populations. 

It can also be seen from Tables 1 and 2 that fecundity, as measured by average 
egg production, is variable. This, as is the case with fertility, is not surprising in 
light of the fact that ananassae is a poor laboratory stock. There is often a de- 
crease in fecundity between the first and second samples. 



DISCUSSION 

This experiment marks the first time that natural populations which received 
large amounts of radiation were tested under experimental conditions in the 
laboratory. 

Wallace (1950), and Wallace and King (1951) tested in population cages 
various populations of Drosophila melanogaster w^hich had been irradiated in the 
laboratory. In their tests they started with populations with lethal free second 
chromosomes and only studied the effects of radiation on the second chromosome 
of the populations. One population containing 10.000 flies w^as given a single 
large dose of radiation, one w^as kept as a control, and others of various sizes were 
subjected to chronic gamma radiation. In the heavily irradiated population there 
was an immediate increase in lethal frequency due to the radiation treatment. 



Gregg: Experimental Populations 



219 



The lethal level then fell off rapidly until the lethal level of the irradiated popu- 
lation was only about eight per cent higher than the level in the control popula- 
tion. From this point on, lethals accumulated at the same rate in the control and 
irradiated populations and both populations were apparently approaching equi- 
librium values. The population which received a single large dose of radiation 
apparently incorporated the radiation produced lethal mutations and other radia- 
tion produced variability, as well as spontaneous mutations, into heterotic gene 
combinations which increased the estimated adaptive value of the irradiated 
population over that of the control, for a time at least. The chronically exposed 
populations showed a decrease rather than an increase in adaptive value. 

Such possible benefit from single large doses of radiation would not be expected 
to prevail in a natural population however, since the total variability of the popu- 
lation, including lethal mutations, is already near an equihbrium value rather 
than below it. Indeed, as Stone et al. (1957) showed, the viability of ananassae 
populations on the Bikini and Rongelap Atolls was still seriously impaired 18 
months after the time of irradiation from the thermonuclear device exploded 
March 1, 1954, due partly at least, to an abnormally high level of lethals. 

In these experiments using Drosophila ananassae sufficient markers and in- 
versions were not available to enable one to make a detailed analysis of the effects 
of radiation on a particular chromosome of the population such as Wallace made. 
The complex problem of comparing the fitness of various populations was re- 
duced to comparing the viability, fertility, and fecundity, of the various popula- 
tions and, associated with viability, the lethal level. These tests made no distinc- 
tion between chromosomes, but rather tested the genome as a whole. Sex linked 




220 The University of Texas Publication 

lethals and lethals associated with rearrangements, such as translocations and 
inversions, that were produced by the initial irradiation, would not be expected 
to be present in the populations by the time that these experiments were started. 
Cytological observations confirmed this expectation (Stone et al. 1957). 

The cage populations consisted roughly of 8,000 to 10,000 flies. The size of the 
populations would be expected to be rather constant since the rate of food addition 
was constant. In contrast the pair-mated populations were quite small, consisting 
of at the most 1,000 flies per generation. Further, the size of the pair-mated popu- 
lations was variable. Fluctuations in fertility and fecundity, especially in 1956, 
sometimes made it difficult to get enough flies to make 500 new matings without 
using large numbers of flies from the more fertile matings. From the small popu- 
lation size of the pair-mated populations it might be expected that some of the 
changes in viability, fecundity, and fertility were due in part to genetic drift. 
Also, the smaller populations would be expected to have been somewhat more 
inbred than the larger populations even though direct brother-sister inbreeding 
did not occur. 

From the inherent differences in the two types of populations it might be 
expected that there would have been greater changes in one of the populations. 
In these experiments however, this was not usually so, although occasionally the 
small populations were more variable. Over a longer period of time more per- 
sistent differences might have arisen between the two populations. In this study 
the cages were only maintained for about 15 generations and the pair-mated 
populations for 12. The generation time of the pair-mated populations was ex- 
tended several days by the handling time necessary for making up the matings. 

The importance of the present tests lies in the fact that they serve as part of 
the control for the original experiments of Stone et al. (1957). The purpose of 
the original tests was to determine the effects of radiation on natural populations 
of Drosophila ananassae. The greatest problem which was confronted was that 
there had been no chance to study the populations before the irradiation oc- 
curred. The normal population cycles could not be measured until after the 
genetic damage had been done. It was necessary to study the damaged popula- 
tions first and to try to establish what were normal population fluctuations later. 
The mean viabilities of the Majuro, Bikini, and Rongelap populations, when 
inbred, in 1955 were 56.6, 46.4, and 40.8 per cent respectively. By 1956 they had 
improved to 66.4, 60.3, and 70.9 per cent. The one small experimental pair-mated 
population of Rongelap flies in 1957 was the only test population that, when in- 
bred, approached the improvement shown by the original populations on Bikini 
and Rongelap. The viability of this population changed from 58.6 per cent in the 
first sample to 73.3 per cent in the second sample. It should be noted, however, 
that in addition to its small size, this population was derived from the Rongelap 
population which recovered from a mean viability of 40.8 per cent in 1955 to 78.9 
per cent in 1956. Thus it is not surprising that this pair-mated population at- 
tained a relatively high level of viability. Another point of interest is that the 
lowest viability shown by any population in these experiments, except for the 
small inbred 1957 Bikini pair-mated population, is 57.4 per cent. This lowest 
value is strikingly and significantly higher than the 46.6 per cent and 40.8 per 
cent viabilities encountered in the Bikini and Rongelap populations in 1955. The 



Gregg: Experimental Populations 221 

48.3 value for the first sample, inbred, of the 1957 Bikini pair-mated population 
is the only value not significantly higher than the 46.6 per cent shown by the 
1955 Bikini population on the atoll. This low value may be explained by sup- 
posing, as Stone and Wilson (1958) suggest, that the Bikini population has still 
not completely recovered and that the 1957 Bikini pair-mated population was 
more highly inbred than the 1956 Bikini pair-mated population. 

SUMMARY 

1 . Samples from natural populations of Drosophila ananassae from the Bikini, 
Rongelap, and Rongerik Atolls, which had been subjected to large amounts of 
radiation, and a sample from the Majuro Atoll, which had received a negligible 
amount of radiation by comparison, were maintained in the laboratory in two 
types of experimental populations. 

2. The various experimental populations were tested periodically for changes 
in viability, fertility and fecundity. These factors were used as an index of evo- 
lutionary fitness. The procedure used in testing for changes in these factors is 
discussed. 

3. The only consistent observation as regards the fertility of the populations 
was that the Bikini populations were almost always more fertile than the other 
populations. Stone et al (1957) also found the Bikini population to be more 
fertile than the other populations. 

4. No consistent differences in fecundity were observed between the various 
populations. 

5. Moderate decreases in the level of lethal mutations from the first sample to 
the second sample in most of the populations indicates that there was some selec- 
tion against lethals, especially when the initial level of lethals was high. 

6. That only moderate changes in viability occurred in the various experi- 
mental populations, especially in the inbred tests, is taken as strong evidence 
that the marked improvement of the Bikini and Rongelap populations in the 
Marshall Islands (Stone et al. 1957) between 1955 and 1956, was due to the fact 
that in 1955 both populations were considerably less viable than normal, due to 
the effects of the radiation that they had received 18 months earlier. 

ACKNOWLEDGMENT 

The writer wishes to express appreciation to Professor W. S. Stone for advice 
and consultation during the course of the experiment, and to Mrs. F. D. Wilson 
for consultation and invaluable technical assistance. This work was supported by 
an A. E. C. contract [AT- (40-1) -1323]. 

BIBLIOGRAPHY 

Dobzhansky, Theodosius and Boris Spassky. 1954. Genetics of Natural Populations. XXII. A 
comparison of the concealed variability in Drosophila prosaltans with that in other species. 
Genetics, 39: 472-487. 

Dubinin, N. P. 1946. On lethal mutations in natural populations. Genetics, 31: 21-38. 

Goldschmidt, E. J., J. Wahrman, A. Ledermann-Klein, and R. Weiss. 1955. A two years' survey 
of population dynamics in Drosophila rnelanogaster. Evolution, 9: 353-366. 



222 The University of Texas Publication 

Ives, P. T. 1954. Genetic changes in American populations of Drosophila melanogaster. Proc. 
Nat. Acad. Sci. Wash., 40: 87-92. 

Merrell, David J. 1951. Interspecific competition between Drosophila funebris and Drosophila 
melanogaster. Amer. Nat., 85: 159-169. 

Moore, John A. 1952. Competition between Drosophila melanogaster and Drosophila simulans. 
I. Population cage experiments. Evolution, 6: 407-420. 

Stern, C. and Edward Novitski. 1948. The viability of individuals heterozygous for recessive 
lethals. Science, 108: 538-539. 

Stone, W. S., Mary L. Alexander, and Frances E. Clayton. 1954. Heterosis studies with species 
of Drosophila living in small populations. Univ. of Tex. Publ. 5422: 272-307. 

Stone, W. S., M. R. Wheeler, W. P. Spencer, F. D. Wilson, J. T. Neuenschwander, T. G. Gregg, 
R. L. Seecof, C. L. Ward. 1957. Genetic studies of irradiated natural populations of 
Drosophila. Univ. of Tex. Publ., 5721 : 260-316. 

Stone, W. S., and F. D. Wilson. 1958. Genetic Studies of irradiated natural populations of 
Drosophila II. Proc. Nat. Acad. Sci. 44:565-575. 

Wallace, Bruce. 1950. Genetic changes within populations after X-irradiation. Genetics, 36: 

612-628. 

Wallace, Bruce, and J. C. King. 1951. Genetic changes in populations under irradiation. Amer. 

Nat., 85: 209-222. 

Wallace, Bruce. 1956. Studies on irradiated populations of Drosophila melanogaster. J. Genet., 
54: 280-293. 

Wright, S., and Th. Dobzhansky. 1946. Genetics of Natural Populations. XII. Experimental 
reduction of some of the changes caused by natural selection in certain populations of 
Drosophila pseudoobscura. Genetics, 31: 125-156. 

Wright, Sewell. 1950. Discussion of population genetics and radiation. J. Cell. Comp. Physiol., 35 
(Suppl \): 187-205. 



Genetic Studies of Irradiated Natural Populations 
of Drosophila. IV. 1958 Tests^ 

WILSON S. STONE AND FLORENCE D. WILSON 

Genetics Foundation, Department of Zoology 

University of Texas 

INTRODUCTION 

Since Muller (1927) discovered that X-rays produced mutations and chro- 
mosomal abnormalities, the problem of the effect of widespread irradiation of 
populations has been of great interest to scientists. The discoveries that led thereto 
and the development of the atomic and thermonuclear bombs have made this 
problem of acute interest not only to scientists but to the general human popula- 
tion. To a number of scientists and to the members of the Division of Biology 
and Medicine of the Atomic Energy Commission, it seemed to be imperative that 
natural populations subject to direct and fallout radiations from atomic and ther- 
monuclear explosions should be investigated. We have been studying the genetic 
damage and its decay through time using natural populations of Drosophila 
ananassae. The irradiated populations live in the Pacific Proving Ground on the 
nothern Marshall Islands, and the control populations are from Majuro in the 
southern Marshalls and Ponape in the eastern Caroline Islands. 

In earlier publications (Stone, Wheeler, Spencer, Wilson, Neuenschwander, 
Gregg, Seecof and Ward, 1957; Stone and Wilson, 1958) we have reported in- 
vestigations of the populations collected in the summers of 1955, 1956 and 1957. 
As we indicated and demonstrated in the earlier publications, Drosophila ananas- 
sae, a tropical species found around the world, is the characteristic species of the 
Pacific Islands. It is suited by its temperature tolerance and the superior competi- 
tive ability of its larvae to survive in these climates on the small islands. Ponape 
has several other species but on the low islands, at least, ananassae is the most 
numerous Drosophila species. The islands in the Pacific Proving Ground which 
received direct irradiation and fallout (Bikini) or fallout only (Rongelap. Ron- 
gerik, and Eniwetak in the Rongerik atoll), have populations of ananassae of 
sufficient size to be studied. The thermonuclear device of March 1, 1954, gave 
very heavy radiation and fallout to these areas. Since then, the fallout on Bikini 
island and especially on Rongelap and Rongerik has been much less intense than 
from the 1954 device. In fact, the Division of Biology and Medicine of the AEC 
informs us that probably 99% of the total fallout on Rongelap and Rongerik came 
from the March 1, 1954 test. The large island, Bikini, in the atoll of that name, 
received fallout on several other occasions. Populations of Drosophila have been 
collected from these islands in late July and August of 1955, 1956, 1957 and 1958. 
This paper reports the tests of the ananassae populations collected in 1958. 

1 This work was supported by a contract with the Atomic Energy Commission [AT-(40-l)- 
1323] and by a grant from the Rockefeller Foundation. 



224 The University of Texas Publication 

EXPERIMENTAL 

The populations collected on the several islands were shipped air mail to the 
University of Texas for analysis. Unfortunately, the size of the sample which 
reached Austin for analysis was small this year. On Ponape a typhoon had re- 
duced the fruit to such a low level that only one very small local population was 
sampled. The population on Majuro was low also, as was the one on Rongelap 
where freshly fallen fruit was very difficult to find. The F^ generation from the 
individual females or pairs sent in was not large enough and an Fo from crosses 
of progeny of different females had to be raised in the laboratory to secure enough 
flies for the tests. These were the flies used in tests 1 through 15, Table 1. The 
progeny of crosses 10 through 15 were tested in the crosses 16 through 26. 

In order to assay the genetic and environmental components of variation in 
these populations, we compared the results of crossbreeding (crossing offspring of 
different parents within a population or crossing individuals from different popu- 
lations) and inbreeding (brother X sister matings) . The variables tested are basic 
components in natural selection: fertility, fecundity and viability. The pairs were 
mated five to eight days after emergence to insure their sexual maturity. Fertile 
pairs are those which produced eggs that developed. Viability was measured in 
terms of egg development. After mature pairs were allowed to mate one day, they 
were changed daily for four or five days to fresh food, the number of eggs laid 
on each day was recorded, then the number of adults that developed was deter- 
mined after a suitable interval of 12 to 15 days. After the egg counts the pairs 
were placed on fresh food. These vials were examined at the end of the period to 
determine if any pairs had proven to be fertile although no eggs from the egg 
count period had developed. More detailed accounts of the techniques and meth- 
ods of analyses are to be found in the article by Gregg (1959, this bulletin). 

RESULTS 

The data from the tests of the populations collected in 1958 are given in Table 
1 . The first four columns give the tests for fertility and an estimate of the number 
of pairs in which at least one lethal was present. This could be determined for 
the parents of brother-sister matings but since too few such cultures were tested 
and fertile, the number of pairs with one or more lethals is underestimated. 

The next four columns of Table 1 give the fecundity, measured as the average 
number of eggs laid per day, and the percentage of pairs that showed the presence 
of a lethal or lethal equivalent. If the egg development from a pair was less than 
80%, the pair was scored as having one or more lethals or lethal equivalents. The 
last seven columns give the information on total egg development from a cross and 
the pattern of egg development for each cross. The last five columns, giving the 
pattern of egg development, were used to make the comparisons between the 
results in 1958 and in earlier years shown in Figures 1 through 4. 

The data can be compared between populations, between years, and within 
and between populations and their heterozygotes using sib matings and cross- 
breeding. Environmental effects are always present and it is desirable to dis- 
criminate as much as possible between environmental effects and genetic effects 
on the variables measured. 



Stone and Wilson: Irradiated Natural Populations 



226 



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226 



The University of Texas Publication 



PONAPE (P) 



-dlU^ I 
88.3 87.8 J 

81.5 



55 ' 56 ' 57 



BIKINKB) 



56 ' 57 ' 58 



69.8 



75.4 74.7 



RONGELAP(A) 



55 ' 56 ' 57 ' 58 



J 83 6 



J 

73.5 

J 



RONGERIK(K) 



55 ' 56 ' 57 



Fig. 1. Egg development within populations. The darkened figures are brother X sister matings; 
the plain figures are crosses between offspring of different pairs. These were run at the same time, 
with the progeny of pairs tested both ways when possible. The test years are indicated at the top 
for each population. The histogram shows the relative percentage of females with egg develop- 
ment of 1-19, 20-39, 40-59, 60-79, and 80-100 per cent from left to right. The frequency distri- 
bution of females which gave different effective egg developments gives a better picture of the 
effective productivity of the population or cross. The average per cent development of all eggs 
laid is given under each population histogram. The letter symbol for the particular island is indi- 
cated with the name. 





MXP 


AXM 




MXB 




PX/! 




BXP 




BXA 






56 ' 57 ' 58 


55 ' 56 ' 57 ' 58 


55 ' 56 ' 57 ' 58 


56 ' 57 ' 58 


56 ' 57 ' 58 


55 ' 56 ' 57 ' 58 


100 


- 












90 


- 


^ 




nJ n 

^ 73.6 


jH 


rf 




r 

86.1 






80 
70 


77.1 

. m 

70.1 


81.8 


71.1 


^^•9 83.8 


ii ^ 


^' jfl 
76.5 ^_ 

70.2 






0^ 


67.8 


HI. 




A 

60 




60 


•ttJ 


64.3 




63.9 









58.2 












50 


- 












40 


- 













Fig. 2. Distribution of classes and average egg development for crosses between flies from 
different populations. 



Stone and Wilson: Irradiated Natural Populations 



227 



The Ponape (P) population was not sampled in 1955. Ponape is a larger high 
island with a number of species in competition. Drosophila ananassae populations 
from Ponape are shghtly superior in egg development on inbreeding or cross- 
breeding to the other populations sampled, Figure 1. The crosses and heterozy- 
gotes of Ponape with other strains do not differ very much from other tests 



55 ' 56 ' 57 ' 58 55 ' 56 ' 57 '58 55 ' 56 ' 57 '58 56 ' 57 ' 58 56 ' 57 ' 58 56 57 58 



J 



-J« 



i 



i"'^ 



MPXMP 
1 ,, I 



^m 



Fig. 3. Distribution of classes and average egg development for brother X sister matings of F^ 
from crosses between populations. 



60 



56 57 ■ 58 



J 



y 



J 



56 57 58 



JJ 



56 ' 57 ' 58 



7 3.0 nmll 



55 ' 56 ' 57 ■ 58 



55 56 ' 57 ' 58 



J 



J 



J 



■J 



Fig. 4. Distribution of classes and average egg development for three-way and four-way crosses 
between different F/s. 



228 



The University of Texas Publication 



although there is an indication that flies from Ponape do not cross as readily to the 
other strains as they do among themselves. There may be some sexual isolation 
factors acting to separate these populations. The strains from Majuro (M), 
Bikini (B), and Rongelap (A) have been tested each year. In following the 
crosses of these populations by numbers in Tables 1 and 2, tests 3, 5 and 8 are 
inbreeding tests using brother X sister ma tings, 11, 12 and 15 are crosses between 
these strains, 19, 20 and 21 are brother X sister matings of the F^^ heterozygotes 
between populations, while 25 and 26 are crosses between these heterozygotes, 
termed "three-way" crosses because they involve three strains. Before considering 
Table 2, it is well to compare the several variables measured in 1957 with 1958 
for all tests. Differences in the unweighed averages of all tests in 1957 and in 1958 
showed a decrease in average egg development of 7.3% (72.8 to 65.5%), 2.2% 
increase in sterility (40.1 to 42.3%) and reduction in average number of eggs 
laid per day of 3.6 (27.3 to 23.7) . The simplest assumption is that the laboratory 
environment was less favorable in 1958, especially in view of the consistency of 
the results — for example, the per cent egg development decreased in 26 of 28 
tests. 

The values and their changes for egg development for all stocks and crosses 
tested each year from 1955 to 1958 are given in Table 2. The general trend is for 

Table 2 
Differences in egg development* 



Tests 


1955 


Difference 


1956 


Difference 


1957 


Difference 


1958 


3. 


Majuro (M) 


56.6 


+9.8 


66.4 


—2.7 


63.7 


—13.4 


50.3 


5. 


Bikini (B) 


46.4 


+ 13.9 


60.3 


—2.6 


57.7 


+7.4 


65.1 


8. 


Rongelap (A) 


40.8 


+38.1 


78.9 


—14.4 


64.5 


—3.3 


61.2 




Average P^ inbred 


47.9 


+20.6 


68.5 


—6.5 


62.0 


—3.1 


58.9 


11. 


Ax M 


64.3 


+20.3 


84.6 


—2.8 


81.8 


—4.7 


77.1 


12. 


M X B 


67.8 


+ 7.6 


75.4 


+4.0 


79.4 


—8.3 


71.1 


15. 


Bx A 


70.2 


+6.3 


76.5 


—2.9 


73.6 


0.0 


73.6 




Average P^ crossbred 


67.4 


+ 11.4 


78.8 


—0.5 


78.3 


—4.4 


73.9 




Average Difference, 


















P^ crossbred — P^ inbred 


+ 19.5 




+ 10.3 




+ 16.3 




+ 15.0 


19. 


AM X AM 


59.8 


+ 11.3 


71.1 


—7.1 


64.0 


—6.4 


57.6 


20. 


BA X BA 


52.4 


+ 17.9 


70.3 


—13.3 


57.0 


—15.0 


42.0 


21. 


MB X MB 


62.5 


—3.9 


58.6 


+6.0 


64.6 


— 7.2 


57.4 




Average F^ inbred 


58.2 


+8.5 


66.7 


—4.8 


61.9 


—9.6 


52.3 


25. 


MB X BA 


84.3 


+0.9 


85.2 


—12.8 


72.4 


—7.6 


64.8 


26. 


AM X MB 


89.4 


—5.7 


83.7 


—5.0 


78.7 


—6.9 


71.8 




Average F^ crossbred 


86.9 


—2.4 


84.5 


—8.9 


75.6 


—7.3 


68.3 




Average Difference, 


















F-^ crossbred — F-^ inbred 


+28.7 




+ 17.8 




+ 14.7 




+ 16.0 




Unweighed Average Difference, 
















(P-^ and F^ crossbred) — 


















(P, and F, inbred) 


24.1 




14.1 




15.0 




15.5 



* At the left are given the stocks and their crosses as well as the designations of averages for certain types of crosses. The 
body of the table gives the per cent egg development each year, the differences between j^ears for crosses and for averages 
of crosses. 



Stone and Wilson: Irradiated Natural Populations 229 

increased viability from 1955 to 1956 (9 of 11 tests) and decreased viability from 
1956 to 1957 (9 of 11 tests) and 1957 to 1958 (9 of 11 tests). This might represent 
environmental effects. It is the common experience for investigators to find that 
heterozygotes are influenced adversely less than homozygous combinations. On 
comparing changes in three-way crosses (25 and 26) with this greater stability 
of the heterozygote in mind, it is seen that there is a small reduction in egg de- 
velopment between 1955 and 1956, and larger reductions between 1956 and 1957, 
and 1957 and 1958. This would suggest that the laboratory test environment 
became steadily worse from 1955 to 1958. Figure 5 plots the change in egg de- 
velopment through time using only averages for types of tests. That data in Table 
2 are consistent enough in direction of change to make such a comparison seem 
reasonable, especially since the P^ crosses and inbreeding were done ai the same 
time, and the F, inbred and three-way crosses were done at the same time. Judged 
by these gross comparisons in Table 2 and Figure 5, there was a greater difference 
in genetic viability and variability in 1955 than in later years as shown by the 
greater spread of values and especially by the greater differences in egg develop- 
ment between inbreeding and crossbreeding due to the influence of Bikini and 
Rongelap genotypes. Also the laboratory environment became worse in 1957 
and 1958 although the genetic difference measured between inbred and cross- 
bred tests remained about the same the last three years. We can ascribe the very 
low values of Bikini and Rongelap in 1955 to the heavy, direct radiation and fall- 
out on Bikini and fallout on Rongelap from the powerful thermonuclear device 
of March 1, 1954. (No test values have gone down as low as these P^ inbred in 
1955.) The changes other than the recovery of Bikini and Rongelap might also 
include population cycles in lethal frequencies in these islands which can be 
demonstrated only by more tests. It should be noted that these two irradiated 
populations are two of the three strains tested each year. We have already 
pointed out that the general trend, which also includes tests involving Ponape, is 
in the same direction. 

DISCUSSION 

Analyses of the genetic variations in populations are always difficult. Stone 
et al (1957) tried to measure as many variables as possible. In that pubhcation 
Wheeler and others discussed the ecology and competition within and between 
species; Spencer showed that the pattern and frequency of visible mutations re- 
sembled those analyzed in species such as Drosophila melanogaster , D. simulans, 
D. hydei, and others; Seecof showed that the cytological variation was conserva- 
tive, much as in D. melanogaster, and that the frequencies and types of rearrange- 
ments produced by X-radiation resembled the results obtained with the well 
analyzed D. melanogaster and D. virilis. However, Stone and Wilson (1957 and 
1958) found that the fertihty and especially egg development were inferior to the 
small or thin desert population of D. novamexicana and D. hydei. 

It is desirable to know about the changes in "genetic load" of detrimental 
mutants through fluctuations in population size, selection and mutation (includ- 
ing irradiation in the northern Marshalls). This requires repeated sampling over 
a long period. Some information on the extent of fluctuations that may be ex- 



230 The University of Texas Publication 

pected can be ascertained in laboratory tests. Gregg (1959, this bulletin) reports 
on such tests, using two types of population structure: cages, where each cage 
contained 2000 to 10,000 flies, and pair matings, which consisted of less than 500 
pairs. He discusses the techniques and analyses in detail. The only large improve- 
ments are from low values toward the general population equilibrium. No test 
value of egg development w^as as low as those of the Bikini and Rongelap popula- 
tions of 1955, either from pair matings or cages. This is in agreement with the 
data for all four years when the stocks were tested as soon as possible after being 
brought in from the field. The tests with Ponape crosses show similar relations 
for 1956, 1957 and 1958. The combined tests given in Table 2 and Figure 5 are 
made from the two heavily irradiated populations, Bikini and Rongelap, and the 
control population from Majuro. The greater difference between inbred and 
crossbred for 1955 is due primarily to the very low viability values of popula- 
tions from Bikini and Rongelap. The most probable explanation for these low 
viabilities is that these two populations were still suffering from the increase in 
number of detrimental mutations from fallout from the March 1, 1954 thermo- 
nuclear device. Figure 6 shows a schematic representation of the effect of the 
thermonuclear device of March 1, 1954 and of subsequent test explosions on the 
genetic load of adverse mutations in the Bikini populations. Fluctuations in the 
environment of the islands and in the laboratory tests and the effects of natural 
selection on the populations prevent our finding such a simple representation of 
the population in our tests. Adverse environment causes us to overestimate the 
number of lethals or lethal equivalents, but linkage with the small numbers of 
autosomes in ananassae prevents our detecting the full number of possible ad- 
verse combinations. Nevertheless, comparisons of the differences between the re- 
sults of inbreeding and crossbreeding on the basic factor of evolutionary fitness, 
viability, allow us to assess and compare the accumulated genetic load of detri- 
mental mutations and gene combinations. Crossbreeding keeps detrimental mu- 
tants in the population heterozygous so that only the dominant detrimental com- 
ponent of their action reduces fitness. Inbreeding allows detrimental factors to 
become homozygous and to occur in a larger fraction of the population as hetero- 
zygotes. It is not possible to determine if detrimental effects such as reduction 
in viability are due to lethals or are the result of a cumulative action of less 
detrimental factors which in sum are called lethal equivalents. The dominant 
lethal components of so-called recessive lethals and lethal equivalents (whether 
heterozygous or homozygous) are often, perhaps always, threshold systems which 
vary in effect with the remainder of the genotype and the fluctuations of the en- 
vironment. It is the sum of these factors, good or bad, that determines the evolu- 
tionary fitness of a population in a particular environment. We have previously 
compared (Stone et al, 1957; Stone and Wilson, 1958) these natural populations 
with the extensive studies of Wallace on irradiated laboratory populations, and 
Gregg (1959, this bulletin) has made further comparisons so we will not repeat 
those here. Haldane (1957) has recently published an article pointing out the 
cost of natural selection in replacing a gene in a population by a more effective 
allele. Our tests do not give us any information on the effect of adding such better 
alleles that might have occurred in these populations. 



Stone and Wilson: Irradiated Natural Populations 



231 



100 



90 



80 



TO- 



GO 



50 



40 



1955 



956 



1957 



O-,. 



1958 




3 WAY CROSSES 

A AVERAGE CROSSBRED 

D Pi CROSSES 



0Fi INBRED 

AVERAGE INBRED 

api INBRED 



Fig. 5. Comparison of egg development on inbreeding and outbreeding. The difference between 
the results of inbreeding and outcrossing gives us a measure of the genetic load of detrimental 
mutations. The plots are for the averages of the tests of the Majuro (control) and Bikini and 
Rongelap (irradiated) populations each year. In addition the average of P^ and F^ inbred and P^ 
and F-^ outcrossed is plotted. 



232 



The University of Texas Publication 



FIGURE 
MUTATION 



6-EFFECT 
LEVEL OF 



OF RADIATION ON 
BIKINI POPULATION 



z 
o 

I- 
< 



NORMAL LEVEL 
OFDETRIMENTAl 



MUTATION LEVEL ONE GENERATION AFTER 
MARCH I, THERMONUCLEAR TEST 



INCREMENT ADDED FROM RESIDUAL FALLOUT RADIATIONS 



MUTATIONS IN 
POPULATION 




MAR. I, '54 AUG. '55 AU6.'56 AU6.'57 AUG. '58 

TIME 

Fig. 6. Graphic diagram showing the increase in number of detrimental mutations in the 
Bikini population due to radiations from detonations and fallout and the decay toward a normal 
level for the population. The Rongelap and Rongerik populations would have a similar but smaller 
increase in mutations, particularly in 1956 and 1958. With a generation time of 10-15 days, the 
total number of generations involved between March 1, 1954, and August, 1958, would be about 
108-161. 



SUMMARY 

An attempt has been made to determine the effect of the direct and fallout 
radiations produced by testing atomic and thermonuclear devices on the geno- 
type of populations of Drosophila ananassae living in the area of the Pacific Prov- 
ing Ground. Collections were made in late July and August in 1955, 1956, 1957 
and 1958 from natural populations of ananassae on Bikini, Rongelap, Rongerik 
and Majuro atolls of the Marshall Islands and from Ponape in the eastern 
Caroline Islands the last three years. 

The populations on the island of Bikini received direct and fallout radiations 
from atomic tests and especially from the thermonuclear device of March 1, 1954. 
This test contributed a large fraction of the total fallout on Bikini from all tests, 
and about 99% of the fallout suffered by Rongelap and Rongerik. Majuro and 
Ponape received very little radiation from the tests so ananassae populations from 
these islands served as control populations. This paper reports the results of test- 



Stone and Wilson: Irradiated Natural Populations 233 

ing the population samples obtained in the summer of 1958. The evolutionary 
fitness of the populations, measured as fertility, fecundity (egg production per 
female per day), and especially viability (egg development) was assayed by a 
series of crossbreeding and inbreeding (brother X sister matings) tests. The best 
measure of the genetic load is the comparison between the same genotypes when 
inbred and crossbred. The genetic load was much greater for Bikini and Rongelap 
in 1955 (the difference in viability in terms of egg development was greater that 
year) and has been reduced to about the same level in 1956, 1957 and 1958. This 
supports the concept that the heavy fallout with its long residual radiation from 
the March 1, 1954 thermonuclear device had increased the genetic load of 
detrimental and lethal factors in the two northern Marshall Island populations, 
but that natural selection had returned these populations to the normal range by 
1956 where they remained in 1957 and 1958. 

ACKNOWLEDGMENTS 

The authors wish to express their appreciation for the support of Mr. Ernest 
Wynkoop, A.E.C. resident engineer, and his staff at the Pacific Proving Ground; 
for the cooperation of several civil administrative personnel on Majuro and 
Ponape; for the logistic support of the Air Force and Navy and the kindness of 
many officers and men of the Armed Services. We wish to thank Dr. Marvin 
Wasserman, Dr. Thomas Gregg, Dr. N. B. Krishnamurthy and Mr. Tsueng- 
Hsing Chang for their assistance in collecting and laboratory work. 

REFERENCES 

Gregg, Thomas G. 1959. Genetic studies of irradiated natural populations of Drosophila. III. 
Experimental populations of Drosophila ananassae derived from irradiated natural popula- 
tions. (This bulletin). 

Haldane, J. B. S. 1957. The cost of natural selection. J. Gen. 55:511-524. 

Muller, H. J. 1927. Artificial transmutation of the gene. Science 66:84-87. 

Stone, Wilson S., Marshall R. Wheeler, Warren P. Spencer, Florence D. Wilson, June T. 
Neuenschwander, Thomas G. Gregg, Robert L. Seecof, and Calvin L. Ward. 1957. Genetic 
studies of irradiated natural populations of Drosophila. Univ. of Texas Publ. 5721: 260-316. 

and Florence D. Wilson. 1958. Genetic studies of irradiated natural populations of 

Drosophila. II. 1957 Tests. Proc. Nat. Acad. Sci. 44:565-575. 



Some Values of Endomitosis 

THEOPHILUS S. PAINTER 
The University of Texas, Austin 

In multicellular organisms growth in body size comes about in two ways. 
Most commonly, of course, there is growth in the size of cells followed by cell 
division. But in the larvae of many insects, as has long been known, growth in 
the size of larval organs is due to the enlargement of individual cells which do 
not undergo mitosis. By larval organs is meant those structures which function 
in larval hfe but which undergo lysis when the larva pupates. As we now know, 
the enlargement of cells in larval organs is due to a real growth and rephcation 
of cell organelles without the dissolution of the nuclear envelope, or cytokinesis, 
a process which is known as endomitosis. After very briefly following the develop- 
ment of the concept of endomitosis the writer wishes to call attention to the 
adaptive usefulness of this method of cell growth. 

When von Mohl, more than a century ago, emphasized that the important 
part of the cell is the material enclosed within the cell wall — and to which he 
gave the name of protoplasm — it was soon realized that not everything seen in 
the protoplasm is living, such as starch grains, oil globules and the like, and 
this gave rise to much speculation about the nature of the living protoplasm and, 
indeed, about the nature of life itself. Two general views were widely discussed 
by biologists during the last quarter of the past century. Many held that the cell 
is made up of a number of living units, a unique kind of molecule perhaps, or an 
organized mass of molecules, endowed with the ability to take in and assimilate 
food materials, to grow, to reproduce and to exhibit all other characteristics of 
life. The other view was that life is a property of the cell as a whole and is due 
to the organization and interactions of cell parts or organelles which of them- 
selves are not living. 

In his book, Plasma und Telle, pubhshed in 1907, M. Heidenhain was one of 
the last of the older biologists of this era to urge that cells are made up of special 
and unique living units. Some fifteen years later, a biologist named Jacob], in- 
trigued, as he wrote, by Heidenhain's arguments sought to put this hypothesis 
to a test. For if cells are made up of living units capable of self-duplication, then 
in non-dividing tissue in which there is a wide variation in nuclear and cell size, 
the volumes of such cells should fall into a geometrical series. The liver of mice 
was selected for study, first because there is a considerable variation in nuclear 
size and, second, because normally mitosis does not occur in such tissue. Nuclear 
diameters were measured and from these nuclear volumes computed. Jacob] 
(1925) found that in late embryonic life, liver nuclei all had substantially the 
same volumes and when these were plotted against frequency a single sharply 
peaked curve was seen. In young adult mice, such a plotting of nuclear volumes 
gave a double-peaked curve; the first corresponded to the size class found in em- 
bryos and the second just double the volume of the latter. Old mice showed three 
peaks, and their volumes formed a geometrical series, 2:4:8, etc. From his 



236 The University of Texas Publication 

studies Jacob] concluded that so far as nuclei are concerned it is evident that a 
process involving an ''innere Teilung" of nuclear contents was taking place. At 
the time, Jacob] 's w^ork received little attention but when salivary gland chromo- 
somes came into prominence in the mid-thirties, both Koltzoff and Bridges in- 
voked the concept of ''innere Teilungen'' to explain the large size of salivary 
chromosomes. 

The investigator who gave the first direct morphological evidence about the 
nature of the ''innere Teilung'' was Geitler (1937) who studied somatic nuclei 
of larval organs in the water strider, Gerris lateralis. In larval tissues of young 
males, Geitler observed that the single X-chromosome remained condensed, or 
heteropycnotic, in nuclei. In older males 2 and 4 X-chromosomes appeared in 
the same type but larger nuclei. This led to a careful study of nuclei of the same 
size class and Geitler discovered that prior to the appearance of the larger nuclei, 
all the chromosomes of the nucleus underwent a series of changes closely parallel- 
ing the behavior of chromosomes in ordinary mitosis. There was an exact cor- 
respondence to the interphase, early, and late prophases in larval organs but the 
nuclear wall remained intact and no evidence of any spindle could be found. 
After the chromosomes reached the late prophase, the two chromatids simply 
separated and the nucleus reverted to the interphase condition. Geitler gave the 
name "endomitosis" to this method of replicating the contents of a cell. In 1939, 
Painter and Reindorp showed that endomitosis regularly occurred during the 
growth in volume of nurse-cells in the ovary of the fruit fly, and this accounted 
for the great variations in the appearance of the nuclei in this type of cell. Since 
these early papers, endomitosis has come to be recognized as occurring in all 
species of plants and animals and this brings up the question. What is the selec- 
tive advantage of endomitosis over ordinary mitosis so that this method of cell 
growth is so widely employed in nature? It is this question which has prompted 
me to prepare this discussion. 

The word endomitosis is used for the process in which there is cell growth and 
the replication of chromosome sets and certain other organelles, without the dis- 
solution of the nuclear wall or the formation of a spindle. As a result cells come 
to contain many chromosome sets. Thus, in either salivary gland or nurse-cell 
nuclei, quite commonly as many as 512 or 1024 haploid sets of chromosomes are 
present. We are not concerned here with the question of how the replicated 
chromosomes are associated. 

In order to understand the adaptive value of endomitosis as a method of cell 
growth, it is necessary to consider a number of facts, dealing in general with cell 
chemistry, which have come to light since endomitosis was recognized. 

An organism which shows an extremely rapid growth during larval stages is 
the common fruit fly, Drosophila melanogaster^ and it is in this species that 
endomitosis leads to the presence of nuclei which carry high multiples of the 
original diploid complex. It is very interesting and extremely important to note 
that the basic problem of larval life is just the same as confronts the organism 
during the formation of the egg from which the larva arises. In order for an egg 
to hatch into a larva it must carry within its nucleus and cytoplasm all the food 
materials needed for embryonic development. Similarly, the larva must assemble 
in the various tissues of its body all of the materials required for the formation 



Painter: Some Values of Endomitosis 237 

of the body of the aduh fly. It is for this reason that any facts and concepts which 
shed hght of the basic processes involved in oocyte growth are directly appli- 
cable to the larval stage as well. Further, since the necessary food materials 
stored in either the cytoplasm of the egg, or in larval tissues, consist of proteins, 
nucleic acids and energy-rich compounds of various kinds, any light which can 
be obtained about the synthesis of such materials is pertinent to a discussion of 
the question before us. 

Since the pioneering work of Caspersson, and Brachet, in the early forties, we 
have known that the synthesis of proteins requires the presence of ribose nucleic 
acid (RNA). A vast amount of experimental data, which has accumulated since 
these early studies, has proved that both DNA and RNA are essential for the syn- 
thesis of proteins. For a recent review see Brachet, 1957. 

A second well estabHshed fact is that large quantities of protein are synthesized 
in the nucleus of those cells which produce proteins either for storage of such 
materials in the cytoplasm of a single cell, e.g., oocytes, or for the secretion of 
such materials by exocrine gland cells. Here it should be noted that part of the 
protein synthesized in the nucleus appears in the form of nucleoli because Vin- 
cent (1955) and others have shown that the main constituent of a nucleolus is 
protein in a rather concentrated, dehydrated form. 

A very remarkable example of protein synthesis within the nucleus is seen 
in the lateral pharyngeal glands which secrete the royal jelly of the young adult 
worker bee. Analysis has shown that royal jelly contains about 15 per cent wet 
weight of proteins. At the time when the adult worker bee emerges from pupation, 
the royal jelly gland is in an undeveloped state and the gland cells look much 
Hke any other diploid cell of the bee. But over the first five or six days of hive 
life, the cells of the lateral pharyngeal gland undergo four or five endomitotic 
division cycles so that each nucleus contains from 64 to 128, perhaps more, sets of 
chromosomes. As the gland cells increase in ploidy the number and size of the 
nucleoli increases. In an actively secreting gland cell the nucleus is crowded with 
a high number (up to 80 have been counted in a single nucleus) of large nucleoli 
each of which is rich in RNA. As the gland cell ceases to secrete the royal jelly 
after some eleven days of hive life, the nucleoli shrink in size, lose their RNA 
and finally disappear completely. 

Another very illuminating fact has recently been established by Allfrey and 
Mirsky (1958) who have shown not only that DNA is essential for the produc- 
tion of protein within a nucleus but also that the amount of protein a nucleus 
can synthesize depends on the amount of DNA it contains. These authors studied 
nuclei of mammalian thymocytes, isolated in a sucrose solution. When all of the 
essential amino acids are added to the medium, by the use of radioactive isotopes 
they have shown that protein is synthesized by these nuclei. If the DNA of nuclei 
is slowly removed by the use of DNase, in measure, the ability to synthesize 
proteins falls off. Nuclei depleted of their DNA can regain some of the ability to 
synthesize proteins if DNA from any source is made available to them. Most 
recently, it has been shown that a polyanion, which is not structurally related 
to the nucleic acids, can be substituted for DNA. From this it appears that, in part 
at least, the DNA has a non-specific effect in protein synthesis and functions be- 
cause of the configuration of its constitutent molecules. 



238 The University of Texas Publication 

From the work of Allfrey and Mirsky the conclusion may be drawn that the 
ability of any diploid nucleus to synthesize protein, other things being equal, 
depends to a considerable extent on how much DNA it contains, or more prob- 
ably on the amount of a special kind of DNA it contains. It is tempting to suggest 
that this special sort of DNA is carried by heterochromatic regions but as yet 
there is no direct evidence of this. However, the chemical heterogeneity of DNA 
is now a well established fact. As (diploid) sets of chromosomes are increased 
within a nucleus, by endomitosis or other means, the capacity of such a nucleus 
to synthesize proteins is greatly increased. 

One more factor must be considered; this is the topographical relation between 
exocrine gland cells and the source of precursors for secretory products. Whether 
one considers animals with a closed circulatory system, or an open one, the es- 
sential fact is that the base of the gland cell is directly exposed to a source of 
food. In the pancreas of man, for example, the base of the gland cell lies adjacent 
to a capillary network in the basement membrane, or in the fruit fly the salivary 
gland lies freely in the body cavity and is directly exposed to the haemolymph. 
The distal surface of the gland cell usually forms the gland duct. 

The adaptative value of endomitosis is well illustrated b}'^ the salivary gland of 
Drosophila melanogaster . At the time of hatching, the salivary gland contains 
around 130 cells and since, on the average, some four cells abut on the duct, at 
any given level, the gland is about 30 cells long. As larvae grow, the need for 
more digestive enzymes can be met in one of two ways. The number of secretory 
cells might be increased by mitosis. The alternative would be for the synthetic 
apparatus of the gland cell to be increased without cell division. 

Because of the topographical relation between a gland cell and a source of 
food, it is readily apparent that to increase the number of cells would produce a 
gland of great length. For example, the length of the salivary gland of the fruit 
fly, at the time of hatching, is about 0.3 millimeters. If all gland cells were to 
undergo a single mitotic division, the length of the gland would about double. It 
is well known, of course, both from cytological analyses of the chromosomes, 
nuclear volumes, and other evidence, that salivary chromosomes contain up to 
some 512 to 1024 chromatids which are the product of 6 to 8 endomitotic division 
cycles. If the length of the salivary gland were doubled even 6 times, the gland 
would be 19 millimeters long which would be extremely difficult to house in a 
mature larva with a body length of only 6 to 8 millimeters. In addition the prob- 
lem of resistance to flow of secretion products would be very great in a long thin 
gland. 

Endomitosis allows for a growth of cell volume in three dimensions and at the 
same time preserves the topographical relation needed for secretion. When the 
volume of a cell is doubled each of the three dimensions is not so greatly affected. 
Embryological studies indicate that during larval development there is actually 
about a 20-fold increase in the length of the salivary gland and presumably 
similar increases in the other two dimensions. 

In addition to the problems introduced by excessive gland length, from the 
standpoint of metabolism and synthetic activities mitosis would entail a loss of 
much time. In the case of an amoeba, for example, which normally divides once 
in 24 hours, Mazia (1955) has shown that synthetic activities go on for about 



Painter: Some Values of Endomitosis 239 

18 hours and the rest of the daily cycle is concerned with the separation of repH- 
cated cell parts. Presumably, similar time relations exist in other dividing cells. 
During endomitosis there is no need for a cessation of secretory activity because 
the replication of chromosomes, and other cell elements normally takes place in 
early interphase, a time when other synthetic processes are at a high level. 

From the evidence which has been presented it is abundantly clear that endo- 
mitosis has great adaptive value to an organism in which true growth is restricted 
to a relatively short period of the entire hfe cycle, or in other situations which 
call for the rapid synthesis of proteins, nucleic acid precursors and other ma- 
terials to meet the special needs of an organism. In the fruit fly true growth from 
food outside the organism (gonads excepted) is confined to the larval period 
which ordinarily lasts about 96 hours. The continuous ingestion of food over this 
period requires the continuous production of digestive enzymes to convert such 
food into a form available for assimilation by the soma cells. The replication of 
chromosomes increases the capacity of the nucleus to synthesize nuclear proteins 
and an increase in cytoplasmic volume not only increases the basal area of gland 
cells, for the intake of food, but also adds to the capacity of cytoplasmic or- 
ganelles which synthesize proteins from such food. 

Examples of cases in which large quantities of proteins and other materials 
are required for special purposes are widespreai in nature. Notable are nurse- 
cell mechanisms so prominent in the ovaries of invertebrates and especially 
insects. In order for a zygote to function effectively it must contain within its 
nucleus and cytoplasm all the proteins, nucleic acid precursors and energy-rich 
compounds required for the development of the embryo to the stage when it can 
obtain food from its environment. In the fruit fly this function of reserve food 
synthesis is largely delegated to specialized nurse-cells which serve as unit fac- 
tories for the production of essential materials. As was shown some years ago 
(Pamter and Reindorp, Ic.) the nurse-cells of Drosophila undergo endomitosis 
and reach a degree of ploidy comparable to that of salivary gland cells. Even- 
tually, the contents of nurse-cells are incorporated into the cytoplasm of the 
oocyte. Throughout the whole animal kingdom, it is the rule for young oocytes 
to be intimately associated with one or more nutritive cells. 

Another example in which endopolyploidy plays a very important role is in 
the lateral pharyngeal glands of young adult worker bees. As I show^ed in an 
earlier paper (Painter, 1945) each of the cells which secrete the royal jelly, dur- 
ing the development of this gland, have the synthetic capacity of the gland cells 
increased in two ways. Initially the young diploid gland cell is associated with a 
nutritive nurse-cell which is at least tetraploid in chromosome constitution. 
Eventually the nurse-cell is engulfed by the gland cell and after this there are at 
least four endoreplications of the chromosomes. The high number and the large 
size of nucleoli, in actively secreting cells, attest to the fact that nuclear pro- 
teins are being synthesized in large amounts. 

In conclusion, the writer feels that in spite of the fact that our understanding 
of cell chemistry is still very fragmentary, many cytological phenom.ena. long 
known from the morphological standpoint, can now be profitably considered 
again in the light of our present knowledge and in this sense an understanding 



240 The University of Texas Publication 

of the adaptive values of endomitosis in the economy of organisms opens the way 
for other such explorations. 

REFERENCES 

Allfrey, V. G., and A.E. Mirsky. 1958. Some effects of substituting the deoxyribonucleic acid of 
isolated nuclei with other polyelectrolytes. Science, 128: 1142. 

Brachet, Jean. Biochemical Cytology. Academic Press, New York, 1957. 

Geitler, L. 1937. Die Analyse des Kernbau und der Kernteilung der Wasserlaufer, Gerris. Z. 
Zellforsch., 26. 

Jacob], W. 1925. tJber das rhymthmische Wachstums der Zellen durch Verdoppelung ihres 
Volumens. Roux Arch. Entwmech. Org., 106: 124-192. 

Mazia, D. Materials for the biophysical and biochemical study of cell division. Advances in 
Biological and Medical Physics. Academic Press, New York, 1956. 

Painter, T. S. 1945. Nuclear phenomena associated with secretion in certain gland cells with 
especial reference to the origin of cytoplasmic nucleic acid. J. Exp. ZooL, 100: 528-547. 

Painter, T. S., and E. C. Reindorp. 1939. Endomitosis in the nurse cells of Drosophila melano- 
gaster. Chromosoma 1: 276-283. 

Vincent, W. S. 1955. Structure and chemistry of nucleoli. Intern. Rev. Cytol., 4: 269-298. 



Alexander von Humboldt and the Science 
of the Nineteenth Century' 

ADOLF MEYER-ABICH 

Professor Emeritus of the History of Science, 
The University of Hamburg, Germany 

I 

The year of the death of Alexander von Humboldt (1859) marks a milestone 
in the History of Science. The same year appeared the first edition of Darwin's 
Origin of Species and with it, started by the evolution theory, there developed a 
completely new Biology. That was the origin of those very revolutionary trends 
of our twentieth century science which obtained their most obvious shape during 
the first years of our century with Planck's Quantumphysics and Mendel's— ^z> 
venia uerbof—'' Quantum'' Genetics. By these absolutely new scientific creations 
the so-called classical physics of Newton, with Helmholtz and Lord Kelvin as its 
last significant representatives, and classical biology from Linne to Cuvier, have 
been definitely brought to their final historical perfection. And it is to be the 
prmcipal task of this essay to show that Alexander von Humboldt's scientific 
work has given us a most perfect and splendid synthesis of that eighteenth and 
mneteenth century science which today we call "classical." 

Such a change, not only in centuries but in eras, comprehends not only chang- 
mg ideas, principles and ideals in science, but also practical methods of gaining 
scientific facts and experience such as voyages of exploration, for instance 
Humboldt is a very good example for this, also. His great American voyage illu- 
minates m the same way the character and spirit of the classical epoch of modern 
science, as do also the leading philosophical ideas of that time. The most noble 
way to exercise science in Humboldt's time was to make exploring expeditions 
and research voyages; in our own twentieth century it is the research Institute 
Research voyages are, of course, being made today, but we are making them, so 
to say, m the form of travelling research institutes. To understand clearly the 
essential difference between research travelling during the nineteenth century 
and our own modern method, we have to compare only the arctic expedition of 
INansen with the antarctic expedition of Admiral Byrd. The polar expeditions of 
the latter twentieth century have been steadily-growing research institutes, 
travelling by boat, motor-sled and plane; Nansen travelled by dogsled and by 
skiing. It IS hard for us to imagine that Nansen, only some few thousand miles 
away Irom his home country, Norway, was without any communication with it 
lor more than four years; Nansen was realizing his research expedition in much 

Ausl^aMhl' A'"^'""T'f "" ^IT" ^'^^"'"'^ ^" ^P"^' 1^^^' '' '^' University of Texas, 
Can da' T^e w 'TT i "^ 1 '."'"'" ^"^ ^^^^' ^°^*°"' ^"^ ^* ^^e University of Toronto, 
vers ty of T^^^^^^^^^ translations of German quotations were made by Prof. Silber of the Um- 

versity ot Texas to whom the author expresses his great indebtedness. 



242 The University of Texas Publication 

the same way that Humboldt did it a century ago, whereas Admiral Byrd, only 
a few years later, applied all of the great achievements of our technocracy. So, 
with the changing century, the age of individual and personal scientific travel- 
ling is also definitely gone, and the era of liberalism and personalism in science 
has been replaced by the still steadily-growing teamwork plan of today. 

Just as Humboldt's work during his epoch was outstanding, so also was his 
great American voyage in the history of research voyages. Before him and after 
him there have been a great many scientific travellers of his kind, but none of 
them has left behind the same glorious memory of his doings as did Humboldt. 
The many mountains, rivers, streams, cities, counties and other places which 
bear his name record it. There has to be a reason for this, which w^e will have to 
consider first in order that we may understand better Humboldt's work. The 
principal reason, it seems to me, has been the fact that Humboldt's entire work — 
his practical research voyage as well as his theoretical books — are guided by one 
and the same philosophical and universal idea which he himself described as 
^^Videe (Tune physique du jnonde'' (in a letter to his friend Pictet) or later by the 
single word "Kosmos'' which we may now characterize as the idea of a philos- 
ophy of the earth or of Geography. The second reason for Humboldt's unique 
success has been the historical fact that he represented the very end of his epoch, 
so that he was able to summarize and synthesize the immense amount of ideas, 
facts, and ideals that the ending of any era has to offer. And, finally, the last 
reason for his splendid results has been the very charming personality of Hum- 
boldt himself, being a nobleman of the cosmopolitan education of Goethe's time 
and also being financially completely independent. All of these characters came 
together to shape Humboldt's work and his personality into the outstanding and 
surpassing figure of classical modern science that we know. 

We must examine this in all of its details, for in doing so we are not only ful- 
filling a request of the past history of science but we are also serving the imme- 
diate philosophical necessities of our present time. To clarify that, I will use a 
somewhat risky allegory, which compares the history of science with political 
history. Machiavelli stated somewhere that in political history the neighbor of 
any state is always its enemy but the neighbor of the neighbor is its friend. Just 
the same has always been true for the sequence of historical epochs and genera- 
tions. For any present time has as its antagonistic opponent its immediate prede- 
cessor, whereas the predecessor of the predecessor always has the meaning of a 
congenial collaborator for it. In this way an examination of Humboldt's time, as 
the antagonist of its own successor and as our own predecessor, will, I hope, help 
us in clarifying the darkness of the spiritual horizon before us. 

II 

What distinguishes Humboldt's work fundamentally from all similar under- 
takings of his contemporaries is, as I said above, the fact that not only his whole 
theoretical but also his empirical and experimental research was guided by one 
and the same universal idea which served him constantly as his philosophical 
ideal of knowledge. Humboldt himself defined it as "Tidee d'une physique du 
monde'' or, in one word, as the idea of the "Kosmos." This philosophy of the earth 



Meyer-Abich: Alexander von Humboldt 243 

or geography accompanied him throughout all his life. Before beginning his 
great American voyage he wrote from Madrid in a farewell letter to his friend 
Baron von Moll concerning his main intentions on that exploration voyage: ''auf 
das Zusammenwirken der Krdfte, den Einfluss der unbelebten Schopfung auf die 
belebte Tier- und Pflanzenwelt, auf diese Harmonie sollen stets meine Augen 
gerichtet seinr' And immediately after his return from that long trip he defined 
anew the general task of his voyage and his literary work in his first general and 
most popular hook,'' Ansicht en der Natur' (Views or Aspects of Nature) : ''Ueber- 
blick der Natur im Grossen, Beweis von dem Zusammenwirken der Krdfte, Er- 
neuerung des Genusses, welchen die unmittelbare Ansicht der Tropenldnder dem 
fuhlenden Menschen gewdhrt,''' that is the main purpose of this book. At the end 
of his Hfe, in "Kosmos, Entwurf einer physischen Weltbeschreibung'' (Cosmos, 
a General Survey of the Physical Phenomena of the Universe), the "book of all 
his hfe" as he himself tells us, he states for the last time: ''Das Grundprinzip 
dieses Werkes ist in dem Streben enthalten, die W elterscheinungen als ein 
Naturganzes aufzufassen^ So the book will give us "die denkende Betrachtung 
der durch Empirie gegebenen Erscheinungen als eines Natur ganzen.''^ With good 
reason one of his biographers (J. V. Cams in Bruhns 1872) states that Humboldt 
was always more deeply interested in the mutual correlation of the facts than 
only to know isolated facts. 

This universal philosophical idea to deliver a whole "philosophy of the earth" 
comprehends not only the work which Humboldt himself accompHshed, it marks 
too the still-living tradition of it throughout the history of science which is again 
becoming strong in our own time. Our discourse, then, has now a double task to 
fulfill; first it has to present a sketch showing how Humboldt has realized it by 
his own work, and then we must characterize the trends of his main ideas and 
ideals of knowledge which continue to work as a living tradition in our actual 
situation of science. 

The first and indeed immensely great undertaking to realize this "cosmic" 
philosophy was Humboldt's great American voyage, or his "Voyage aux regions 
equinoctiales du Noveau Continent, fait en 1799, 1800, 1801, 1802, 1803 et 
1804,'' as it is given in the title to his great opus concerning this voyage, con- 
sisting of thirty large volumes and which appeared in Paris from 1808-1834. 
This gigantic work represents a real opus scientificum iberoamericanum, and 
marks the start of modern scientific research in Latin America. Official repre- 
sentatives of the University of Habana, Cuba, conferred sometime a mark of 
honor upon the monument of Humboldt which stands before the University of 
Berlin, which names Humboldt the second discoverer of Cuba (after Columbus) . 
It seems to me that we are not really saying too much to characterize Humboldt 
as the scientific discoverer of Latin America. His opus americanum encompasses 



- "upon the coordination of forces, the influence of inanimate creation on the animate world 
of plants and animals, upon this harmony shall my eyes be constantly directed." 

•^ "The survey of gross natur3, the demonstration of the coordination of forces, the renewal of 
enjoyments which the immediate inspection of tropical lands gives to sensitive men.'" 

^ "The basic principle of this work is expressed in the struggle to comprehend the appearances 
of the world as a totality of nature." So the book will give us "the thoughtful contemplation of 
the whole of nature as given empirically through appearances." 



244 The University of Texas Publication 

all of the sciences of his time: Biology, Geography, Geology, Geophysics, Zoology, 
Archaeology, History, etc., and furthermore, he has created a field of science 
which did not exist until this time, the Geography of Plants. Also it is true that 
his grandiose geographical monographs, particularly those dealing with Mexico 
and Cuba, have for the first time made Geography as a real and exact science. 
Before Humboldt Geography was little more than a collection of more or less 
interesting facts or curiosities, and could scarcely be called a science. For Hum- 
boldt himself, the immense collections of facts and experiences had meaning only 
in providing him with all that empirical experience which is needed to obtain 
a philosophy. But the creation of such a philosophy needs a man whose eyes are 
looking not only at the heavens but whose feet are also strongly rooted in the soil 
because, as Humboldt says (Bruhns 1872): ""Man schadet der Erweiterung der 
Wissenschaft, wenn man sich zu allgemeinen Ideen erheben and dock die ein- 
zelnen Tatsachen nicht kennen lernen will.''''' In the aforementioned Ansichten 
der Natur^ his most favorite book as well as that of the general public, we have 
the best illustration of how he connected ideas with facts. This book consists of a 
series of independent essays of high literary rank written in the most noble Ger- 
man created by Goethe, Humboldt's best friend and, as we later will recognize, 
also his best adviser in science. Each of these essays represents in itself a "micro- 
cosmos" about its special theme (as compared with the great "Kosmos" compre- 
hending the universe as a whole) and each is accompanied by a long series of 
notes which give references to all the facts concerned, correlating them inti- 
mately with the ideas in the corresponding essays. Many of these notes represent 
in themselves small scientific treatises based upon observations and exact meas- 
urements and often consist of many times more pages than the essays themselves. 
But it is not only bibliographically — in the logical structure of his books — that 
we recognize that Humboldt's collecting of facts and experience was always 
guided by his philosophical ideas; we also have the good testimony of the voyage 
itself, testimony which reveals to us at once also the spiritual origin of his cosmic 
idea. Humboldt writes (in Ansichten der Natur) : 'Tn den Wdldern des Amazon- 
enflusses ivie auf dem Rilcken der hohen Anden erkannte ich, wie von einem 
Hauch beseelt von Pol zu Pol nur ein Leben ausgegossen ist in Steinen, Pflanzen 
und Tier en und in des Menschen schwellender Brust. TJeberall ward ich von dem 
Gefilhl durchdrungen, wie mdchtig jene Verhdltnisse in Jena auf mich gewirkt, 
wie ich durch Goethes Natur einsichten gehoben^ gleichsam mit neuen Organen 
ausgerilstet worden warT^ Here we are at the real origin of Humboldt's cosmic 
ideas and philosophy. They are most intimately connected with Goethe's natural 
science and philosophy. Here, too, we are at the starting point to prove and sub- 
stantiate the main thesis of our lecture — that Humboldt represents the culmina- 
tion point and the highest perfection of the natural science of Goethe's time. Only 



^ "Now the expansion of science suffers if one rises to universal idaas and yet is not willing to 
learn the individual facts." 

*5 "In the forests of the Amazon River as upon the ridges of the high Andes I recognized how 
only one life, animated by one breath from pole to pole, is diffused in stones, plants, and 
animals, and in the swelling breast of man. Above all, I was filled with the feeling of how power- 
fully these relationships affect me in Jena, of how I had been elevated and, at the same time, 
equipped with new organons by Goethe's Insights into Nature.'''' 



Meyer-Abich: Alexander von Humboldt 246 

in connection with it do Humboldt's results and ideas show their real sense and 
high value. 

Ill 

The last five years of the eighteenth century Humboldt spent preparing 
spiritually and practically his great American voyage which started June 5th, 
1799, in La Coruna, Spain. Essential for these years were Humboldt's various 
visits' with Goethe in Jena and Weimar during the years 1 794-97 and the follow- 
ing visit in Paris where at this time the best scientists of Europe worked together 
at the Paris Academy and Natural History Museum. In the circle around Goethe, 
Humboldt conceived and evolved his natural philosophy and the general princi- 
ples of his scientific thinking, whereas in Paris he learned the practical methods 
to estabhsh by modern exact measuring scientifically assured facts and experi- 
ence. Upon these fundamental principles are especially based the original scien- 
tific creations of Humboldt's— his morphological geography of plants and his 
physiology. 

Goethe, as is well known, was a morphologist in all his science, even in Anat- 
omy and Botany just as in his doctrine of colors and his essays about philology 
and history. The word "morphology" itself was created by Goethe. The basic 
principles of Goethe's, the so-called "ideahstic morphology" (Naef 1919) have 
also been the fundamental principles upon which Humboldt based his plant geog- 
raphy and all his philosophy of the Kosmos. And we are able to determine exactly 
the meeting point between Goethe's and Humboldt's thinking: it is fixed in the 
first personal meeting of them during the year 1794. In his corresponding diary 
Goethe writes about this meeting: ''Alexander von Humboldt, Idngst erwartet, 
von Bayreuth ankommend, notigte uns ins Allgemeinere der N aturwissenschaft 
. . ." And in his ''Nachtrdgen zur Osteologie'' Goethe points out more precisely 
about the matters they have been discussing, namely: '7c/z trug die Angelegen- 
heiten meines Typus so oft und zudringlich vor, dass man, beinahe ungeduldig, 
zuletzt verlangte, ich solle das in Schriften verfassen, was mir im Geiste, Sinn 
und Geddchtnis so lebendig vorschwebte.''' Following this proposal Goethe wrote, 
in the year 1795, therefore immediately after Humboldt's visit, his famous 
''Erster Entwurf einer allgemeinen Einleitung in die vergleichende Anatomie, 
ausgehend von der OsteologieJ'^ which I consider to be the best and most thor- 
ough treatise of all the biological writings of Goethe. Here he evolves his three 
fundamental principles of Morphology, which — no accident, of course — are just 
the same principles upon which Humboldt based his new science of Plant 
Geography. 

To show that, we must first evolve Goethe's views of these principles. They are 
three: the principle of type in morphology, the compensation principle, and the 



7 "Alexander von Humboldt, long awaited, arriving from Bayreut, forced us into the universal 
aspects of natural science . . ." And in his Supplements to Osteology Goethe points out more 
precisely about the matters they have been discussing, namely: "I expressed this business of my 
Typs so often and so obtrusively that one finally demanded, almost impatiently, that I set down 
in writing that which hovered so vitally before my mind, sense and thought." 

8 "First proposal of a General Introduction to Comparative Anatomy, derived from 
Osteology." 



246 The University of Texas Publication 

principle of holism (as it is called today, after Smuts, to underline its antagonism 
to the principle of mechanism) . The principle of type used by Goethe and Hum- 
boldt has nothing to do with the phylogenetic types of our present Biology based 
upon the Evolution Theory. Goethe's type represents a pure ideal construction 
without any historical meaning, wherefore Naef has named this kind of morphol- 
ogy "idealistic morphology." Indeed, this is all really true Platonism. Goethe 
very often confessed himself to be a Platonist: ''Um sich aus der grenzenlosen 
Vielfachheit, Zerstiickelung und Verwicklung der modernen Naturlehre wieder 
ins Einfache zu retten^ muss man sich immer die Frage vorlegen: wie wiirde sich 
Plato gegen die Natur^ wie sie uns jetzt in ihrer grosseren Mannigfaltigkeit^ bei 
aller griindlichen Einheit, erscheinen mag^ benommen haben?'"''^ Unity in the 
multiplicity — that is indeed the most concise formula to define the platonic idea, 
and it serves equally well to define the type in Goethe's idealistic morphology. 
The ideal geometrical circle, for example, represents the ideal unity in the multi- 
plicity of an infinity of real circles, each one of them somewhat different from all 
the others. And just the same is true of the relation of the ideal type of any 
morphological species of plant or animal to the almost infinite multiplicity of the 
really existing individuals of those species, each of them somewhat different from 
all the others. So any morphological type, too, always represents the ideal unity 
of a real multiplicity. 

We recognize that immediately, if we hear now how Goethe himself defines 
his anatomical type in the above-mentioned treatise: ''Deshalb geschieht hier ein 
Vorschlag zu einem anatomischen Typus, zu einem allgemeinen Bilde, worin 
die Gestalten sdmtlicher Tiere, der Moglichkeit nach, enthalten wdren, und 
wonach man jedes Tier in einer gewissen Ordnung beschriebe. Dieser Typus 
miisste soviel wie moglich in physiologischer RUcksicht aufgestellt sein. Schon 
aus der allgemeinen Idee eines Typus folgt, dass kein einzelnes Tier als ein 
solcher Vergleichskanon aufgestellt werden konne; kein Einzelnes kann Muster 
des Ganzen sein. . . . Die Idee muss ilber dem Ganzen walten und auf eine genet- 
ische Weise das allgemeine Bild abziehen.''^'^ Remember that the term "genetic" 
here means typological-genetic but not phylogenetic. Take, for example, Goethe's 
"metamorphosis" of the leaf into the flower, which illustrates exactly the typo- 
logical genetic development but of course not a real historical phylogenetic evo- 
lution. The best samples for Goethe's ideal types are his "Urpflanze" (original 
plant) and his "Urtier'' (original animal). They represent for him ideal ana- 
tomical models for possible physiological construction of new kinds of really 
possible plants and animals. In a letter written in 1787 from Rome to Mrs. von 



" "In order to preserve oneself in simplicity from the limitless diversity, partitioning and 
complication of the modern theory of nature, one always must pose to himself this question: 
How would Plato have maintained the basic unity of nature had he confronted her, as she now 
appears to us, in far greater multiplicity." 

^0 "A proposal, therefore, is put forward here of an anatomical type, of a universal picture, 
wherein the forms of all animals, according to their possibility, would be included. And accord- 
ing to this type, or picture, each animal can be described in a certain order. This type must be 
based, so far as possible, on physiological considerations. From the universal idea of a type it 
follows that no single animal could be proposed as a paradigm for comparison. No individual 
can be a model for the whole . . . The idea must rule over the totality and, according to a genetic 
method, derive the universal picture." 



Meyer-Abich: Alexander von Humboldt 247 

Stein, Goethe says from his Urpflanze: "Mit dem Modell und dem Schlussel dazu 
kannman alsdann noch Pflanzen ins Unendliche erfinden, die konsequent sein 
mussen, d.h., die, wenn sie auch nicht existieren, dock existieren konnten und 
nicht etwa malerische oder dichterische Schaiten oder Scheine sind, sondern eine 
innerliche Wahrheit und Notwendigkeit haben. Dasselbe Gesetz wird sich auf 
alles ubrige Lebendige anwcnden lassen."^^ In this sense, as typological-physio- 
logical models but not as real historical beings, Goethe understood his Urpflanze 
and Urtier as anatomical types, and as we will see, this is true also of Humboldt's 
"Physiognomische Grundgestalten'' (physiognomic groundforms) in his plant 

geography. 

The second typological-morphological fundamental principle from Goethe 
which Humboldt also applied in his plant geography is the so-called compensa- 
tion principle. It defines the basic postulate which any typological transformation 
—named metamorphosis by Goethe — has to fulfill if it is a possible one and a 
"true" one. Goethe says this about it: "Betrachten wir nach jenem, erst im allge- 
meinsten aufgestellten Typus die verschiedenen Tiere der vollkommensten, die 
wir Sdugetiere nennen, so finden wir, doss der Bildungskreis der Natur zwar 
eingeschrdnkt ist, dabei jedoch, wegen der Menge der Telle und wegen der viel- 
fachen Modifikabilitdt, die Verdnderungen der Gestalt ins Unendliche moglich 
werden. ...Wenn wir die Telle genau kennen und betrachten, so werden wir 
finden, dass die Mannigfaltigkeit der Gestalt daher entspringt, dass diesem oder 
jenem Teil ein Uebergewicht iiber die andern zugestanden ist. ...So sind, zum 
Beispiel, Hals und Extremitdten auf Kosten des Korpers bei der Giraffe begun- 
stigt, dahingegen beim Maulwurf das Umgekehrte stattfindet. ...Bei dieser Be- 
trachtung tritt uns nun gleich das Gesetz entgegen: dass keinem Teil etwas 
zugelegt werden konne, ohne dass einem andern dagegen etwas abgezogen werde, 
und umgekehrt. ...Der Bildungstrieb ist hier in einem zwar beschrdnkten, aber 
dock wohl eingerichteten Reiche zum Beherrscher gesetzt. Die Rubriken seines 
Etats, in welche sein Aufwand zu verteilen ist, sind ihm vorgeschrieben; was er 
auf jedes wenden will, steht ihm, bis auf einen gewissen Grad, frei. Will er der 
einen mehr zuwenden, so ist er nicht ganz gehindert, allein er ist genotigt, an 
einer andern sogleich etwas fehlen zu lassen; und so kann die Natur sich niemals 
verschulden oder wohl gar bankrott werden.''^- Both of these fundamental prin- 



11 "With this model and with the key thereto one can discover an endless number of plants 
which, as a consequence, must be. That is to say, these plants, even if they do not exist, could 
nevertheless exist. They are not some pictorial or poetic images or appearances; rather they have 
an inner truth and necessity. The same law can be applied to all other living beings." 

12 "If, after that, we consider first the most perfect in the most general established types of 
different animals, which we call mammals, we find that the organizational system of nature is 
ind-ed limited. Nevertheless, because of the multitude of parts and because of the diversity of 
capacities for modification, the alteration of forms ad infinitum becomes possible. ... If we 
know and consider the parts exactly, we will find that the multiplicity of form springs from the 
fact that this or that part is granted the upper hand over the other. ... So, for example, in the 
giraffe the neck and extremities are favored at the expense of the body. The reverse situation is 
found in the mole. ... By means of this consideration we advance at the same time toward the 
law: that no part will be able to increase unless something is, on the other hand, taken from 
another part, and vice versa. . . . The organizational impulse here is put forth as the governor 
of a Kingdom, indeed limited but nonetheless well directed. The rubrics of its budget in which 



248 The University of Texas Publication 

ciples of the typological morphology from Goethe and Humboldt are based upon 
one and the same still more fundamental biological principle, that same one 
which overran and exceeded Humboldt's own epoch, helping us today to over- 
come the mechanistic philosophy of nature of the second half of the last century. 
This principle we call today the principle of holism, after Smuts, and which, of 
course, is antagonistic to any mechanism. Speaking here of mechanism and 
holism we do not mean the mechanistic or holistic general philosophy of nature, 
but both principles as causal ones which are helping us to form exactly demon- 
strated scientific theories. So, for example, when we are speaking of the mech- 
anism of atomic activity or of respiration, then we may say that we are able to 
explain these complicated phenomena in the causal way of exact physical or 
physiological research. In this sense mechanism and holism as causal principles 
are antagonistic and opposed principles but they are not contradictory in the 
sense of logic. There are many phenomena in nature that we are able to explain 
as pure mechanisms, but there are also others that we may understand only as 
holisms. So the causal principles of mechanism and holism are not contradictory 
ones, as the corresponding metaphysical systems of mechanism and holism are, 
because they — as all philosophical systems are — are concerned always and prin- 
cipally with totalities of reality, whereas scientific theories are just as funda- 
mentally concentrated in particular realities. 

The holistic causality we cannot define better than was done by Christian 
von Ehrenfels by his statement that "the whole represents more than just the 
sum of its parts." If we are not able to explain a complicated phenomenon of 
physical or biological nature by mechanistic causality, which means to analyze 
it into its most simple elements and then to summarize it out of these simple 
elements, then we are dealing with a phenomenon which is to be understood 
only in the way that holistic causality points out. That means that we have to try 
to explain the activities of the parts of any whole or holism only by starting with 
the whole as such, which of course then has to be known to us. This is precisely 
the case in Goethe's and Humboldt's morphological types. The possible compen- 
sations in and by them we are able to consider and construct only if first we have 
a clear idea of our type as a whole or as an holism in the sense of Smuts. Whereas 
the strongly mechanistical epoch, which followed the Humboldt-Goethe-Time 
and preceded our present time, has been absolutely convinced that in some future 
time we would be able to explain all phenomena of nature, including the most 
complicated biological and psychological ones, by the way of mechanistic caus- 
ality, today we recognize more and more phenomena as absolutely not being able 
to be explained sufficiently as pure mechanisms. And this is true not only in psy- 
chology and biology but in microphysics too. From Planck himself, we can 
understand a microphysical material-wave only if we consider it as a whole, the 
behavior of its microelements, quanta, being determined only by the wave as a 
holism, and not, reciprocally, the behavior of the wave as a whole summarized 
by the singular behavior of its microconstituents. And the so-called Pauli-prohibi- 

its expenditures are to be divided, are preordained. What it wishes to spend on each is, to a certain 
degree, uncontrolled. If it wishes to spend more on one it is not completely prevented from 
doing so. It is merely required, at the same time, to leave something lacking in another. And so 
nature can never go in debt nor become completely bankrupt." 



Meyer-Abich: Alexander von Humboldt 249 

tion, a fundamental postulate of modern atomphysics, has been recently charac- 
terized by March as a typological-morphological principle — in the sense of 
Goethe and Humboldt — and not as a principle of causal physics. It is indeed, 
biologically considered, really a morphological axiom of the same observance as 
the compensation principle of Goethe, to which group of logical principles it 
belongs. 

Our statements have made it perfectly clear that neither Humboldt nor 
Goethe could be mechanists. As for Humboldt, we will recognize that subse- 
quently also when dealing with his physiology. There we will see that his use of 
the term "Lebenskraff (vital force) does not mean that he was a vitalist. Not 
at all! It means only that he represented modern holism in the then usual 
Goethe-Schelling fashion and, of course, with an obviously anti-mechanistic 
tendency. Now we have to show how Humboldt applied the mentioned principles 
of Goethe's typological morphology in his Plant Geography. This science did not 
exist before Humboldt; it represents his own and his most original scientific 
creation which endures his particular epoch and ideas, being a new science of its 
own independent historical existence. It has been said that all Humboldt ac- 
complished has been geography, or at least more or less intimately connected 
with geography. With the exception of his physiology, that is absolutely true, 
and we may add that his geography culminated in his Plant Geography. Here 
we find together all that is true and perfect in Humboldt's work. The corre- 
sponding book is titled: '^Essai sur la geographie des plantes; accompagne d'un 
tableau physique des regions equinoctiales... Paris 1805'' In the 30-volume 
Corpus Americanum it bears the number XXVII, but it was the first book which 
appeared of the whole series. Humboldt himself translated it into the German 
language, the German edition being titled: ^'Ideen zur Geographie der Pflanzen, 
nebst einem Naturgemdlde der Tropenldnder, auf Beobachtungen und Mess- 
ungen gegriindet, welche vom. 10. Grade nordl. bis zum 10. Grade siidl. Breite in 
den Jahren 1799-1803 angestellt worden sind. Tubingen 1807'\ 4°, XII, 182 p. 
with 1 Tab. It has been dedicated to Goethe who has been said to have devoured 
it, and he was so enthusiastic about it that he immediately painted the corre- 
sponding table, which was still lacking in his first specimen of the book. If now 
we have to characterize the kind of plant geography Humboldt has realized 
here, we can do it with one word only: it is a very significant typological plant 
geography, typological in precisely the sense that Goethe used the term. 

Today plant geography, like all other biological sciences, is based absolutely 
upon the evolution theory. That could not be the case with Humboldt, whose 
book appeared more than half a century before Darwin. All his statements about 
the matter are clearly correlated with the principles of Linne and Cuvier con- 
cerning the constancy of species and types. Therefore all historical trans- 
formations and changes in the composition of the vegetation on earth are only 
due to the geological cataclysms and subsequent new immigrations from neigh- 
boring territories, as Cuvier has instructed us. Says Humboldt: "Die urtiefe 
Kraft der Organisation fesselt^ trotz einer gewissen Freiwilligkeit im abnormen 
Entfalten einzelner Teile^ alle tierische und pflanzUche Gestaltung an feste^ 



250 The University of Texas Publication 

ewig wiederkehrende Typen.''^^ Are those not almost the same words Goethe 
used to describe his "general type", which we quoted above? But we have to 
distinguish between two principally different kinds of types, the homologue- 
ones, which constitute the taxonomical "Natural System" of organisms and 
which are connected particularly with the characters of the sexual organs, the 
so-called organization-characters, and the analogue-ones, which refer especially 
to the functional similarities or so-called convergences, and which are connected 
particularly with the vegetative organs of the organisms. You remember the 
interesting historical fact that Linne gained his overwhelming success in forming 
his plant-system by using the organization-characters of the sexual systems, 
whereas Haller failed to succeed because of using the vegetative organ-systems 
for the same purpose. But to establish a "natural system" of plant-geographical 
types, as Humboldt intended to do, only the analogue characters of the vegetative 
organ-systems are important, because particularly by the adaptable faculties 
and functions of the organs for nutrition, sensation and motion the organisms 
are realizing their relations with the environment, which, particularly for the 
Plant Kingdom, are mostly identical with the corresponding geographical cli- 
matical zones and plant formations. So the "general types" in which Humboldt is 
interested in his Plant Geography, are the analogue-ones, which today we also 
call ecological types. Therefore, Plant Geography has the general task to investi- 
gate "oZ? man unter den zahllosen Gewdchsen der Erde gewisse Urformen 
entdecken oder oh man die spezifische Verse hiedenheit als Wirkung der Ausart- 
ung und als Abweichung von einem Prototypus betrachten kann'^^^ This is 
very true typology of the analogous types, which corresponds logically and 
exactly with Cuvier's famous typology of the homologous types. The same 
way as Cuvier in his famous struggle with Geoffroy St. Hilaire. and in which 
Goethe, too, was deeply involved, was interested to find out if all types of 
animals could be derived typologically from one and the same fundamental 
type — from one "Urtier'' as Geoffroy and Goethe postulated — or if we would 
have to consider different fundamental and autonomous types to cover the 
multiplicity of animal types, as Cuvier stated, that is also here the question for 
Humboldt's typology of analogous geographical or ecological plant formations. 
So Humboldt, too, here faces the problem: whether all these different plant geo- 
graphical formations could be typologically derived from only one fundamental 
plant formation, or if it would be necessary to accept different independent 
ecological types. His answer is the same as Cuvier's, namely, in favor of a 
plurality of different fundamental ecological types. That we have to deal here 
with true typology and not with what we today call phylogeny, becomes per- 
fectly clear if we consider only Humboldt's statement related to the correspond- 
ing animal geography: "Die kleine und schlanke Form unserer Eidechse dehnt 
sich im Silden zu dem kolossalen, schwerfdlligen, gepanzerten Korper furcht- 



13 "The basic force of organization holds all plant and animal forms in fixed unchanging and 
recurring types. This is true in spite of a certain degree of freedom for exceptional developments 
\iY the single part. 

1* ". . . whether one can discover definite basic forms beneath the countless sorts of life grow- 
ing from the earth, or whether one can observe the specific differentiations as a result of the 
breakdown and modification of one prototype only." 



Meyer-Abich: Alexander von Hum.boldt 251 

barer Krokodile aus. In den ungeheuern Katzen von Afrika und Amerika, im 
Tiger, im Loewen und im Jaguar ist die Gestalt eines unserer kleinsten Hans- 
tiere nach einem grosseren Masstabe wiederholt.''^'' This of course, is not his- 
torical phylogeny but only very true typology. 

The analogous fundamental types of Humboldt's plant geography are called 
"physiognomische Grundgestalten'' — physiognomic groundforms. Humboldt's 
main problem now is to make a typological "natural system" of them. But to do 
that we must first estabhsh a fundamental law or general principle upon which 
may be based our physiognomic "natural system." In Humboldt's case this 
fundamental law, of course, has to be the basic principle of all Plant Geography. 
To have discovered it, remains for all times one of the best achievements of 
Humboldt in science. This basic principle of plant geography establishes an exact 
relation between the vegetation of a zone and its climate. So Humboldt's law 
states first that equivalent medium temperatures create forms of vegetation, 
which are physiognomically analogous, and second, that with increasing altitude 
of the mountains or with increasing approach to the poles of the globe the tall- 
ness of the stem-organs decreases. Based upon this principle Humboldt distin- 
guished 17, later 19, physiognomic-typological groundforms of the vegetation. 
The most characteristic ones are the following: the bananatype, the palmtype, 
the treeferntype, the aloetype, the coniferoustype, the mimosatype, the lilytype, 
the cactustype, the grass- and reedtype, the mosstype, the lichentype, the mush- 
roomtype, and others. Using these physiognomic types we are now able to com- 
pose them into perfect natural "genera" and "families" of vegetation-formations. 
Two of these Humboldt has studied especially during his Latin American 
voyage: the Llanos of Venezuela and the Hylaea Amazonica. The Llanos repre- 
sents a holistic composition of grasses, mimosas and palms as its determining 
physiognomic groundtype, whereas the Hylaea, as a tropical rainforest, is defined 
physiognomically by trees belonging to the mimosa and laurel groups associated 
with palms, bamboos and heliconias. 

Our considerations and remarks are surely sufficient to illustrate the perfect 
and almost exclusively typological character of Humboldt's Plant Geography in 
the sense of Goethe's general morphology. And Humboldt's Plant Geography, 
as his most important and significant scientific creation, is absolutely repre- 
sentative of all his scientific work. The physiognomic groundforms represent, 
in Humboldt's typological plant geography, the "general types" of Goethe's 
morphology, whereas the different "genera" and "families" of the vegetation 
formations — such as the Llanos or the Hylaea — are completely based upon the 
"compensation principle" of Goethe. If we compare, for example, (and like all 
typological morphology Humboldt's Plant Geography is a comparative science!) 
the amazonian Hylaea with an African tropical rainforest or the Venezuelan 
Llanos with an Asiatic tropical savanna, then in both cases we have the same 
"genus" or "family" of vegetation formation, but in each case composed of very 



^5 "The small, slim form of our lizard, in the South dilates into the huge, awkward, mailed 
body of the frightful crocodile. The form of one of our smallest house pets is repeated on a 
greater scale in the enormous cats of Africa and America, such as the tiger, the lion and the 
jaguar." 



252 The University of Texas Publication \ 



different plant and tree species, which this way have to compensate each other, 
but under the hoHstic rule of always retaining the same constant physiognomic 
vegetation formation. In a letter to his friend Berghaus Humboldt describes the 
compensation in different formations of plants in the following words: ''Die 
Einheit der Natur ist dergestalt, dass sich die Formen einander nach bestehenden 
unwandelbaren, noch nicht durch die menschliche Einsicht ergriindeten Gesetzen 
ausgeschlossen haben. Kennt man auf irgend einem Punkte des Erdrundes die 
Zahl der Arten einer grossen Familie, z.B. der Glumaceen, Compositen oder 
hills enarti gen Gewdchse^ so kann man mit einiger W ahrscheinlichkeit sowohl 
die Totalmenge der phanerogamischen Gewdchse, als auch eine Anzahl der 
Arten, woraus die anderen Pflanzengruppen bestehen, schatzen.""^^ The word 
"holistic" as used above means nothing philosophical but only scientific, namely 
that all our physiognomic groundtypes and formations represent true holisms but 
never mechanisms, as we have already explained these terms above. Humboldt, 
like Goethe, is using in this connection the platonic and aristotelic concept of 
harmony, which belongs always only to holisms but never to mechanisms, 
which are only able to produce physicochemical equilibria instead of living 
harmonies. These considerations guide us, then, to our last remarks, concerning 
Humboldt's physiological ideas and research. 

IV 

Humboldt's main work concerned geography in the broadest sense of this 
science. He was not only the creator of modern scientific geography, he also 
first founded Plant Geography as a morphological-typological science in the 
sense of his master, Goethe. All these morphological sciences deal with holisms, 
whereas modem causal physiology is always concerned with the research of 
mechanisms. Therefore if we now consider Humboldt's kind of physiology, we 
have to examine first its relation to his holistic natural philosophy. 

Like Goethe, Humboldt too was always thinking physiologically. He surely 
was a typological morphologist, but he was always more interested in the 
functioning or in the analogous correlations of his types than in their pure 
typological constitution or homologies. When he was still a young student of 
only 19 years he wrote a letter to his brother Wilhelm, in which he speaks of the 
necessity to investigate more the physiological energies of the plants — its 
''Krdfte'' — than its taxonomical constitution only. He says that such studies in 
the future will be of greatest practical importance for human nutrition. To study 
Botany this way would be for him essentially: ''weil ich an einem Werke Uber 
die gesammten Krdfte der Pflanzen ... sammle, ein Werk, das wegen des vielen 
Nachsuchens und der tiefen botanischen Kenntnis bei weitem meine Krdfte 
iibersteigt, und zu dem ich mehrere Menschen mit mir zu uereinigen strebe.''^'^ 

1*^ "The unity of nature is such that the forms are imposed one upon the other as enduring and 
unchanging, not yet derived from scientific laws imposed by man. If one knows exactly the 
cipher of the species of a large family — for example the Glumaceae, Compositae, or pod-forming 
plants — he could estimate with great probability the total quantity of phanerogamic plants 
which belong to the same group as the former." 

^"^ "I collected into one work [everything] concerning all the forces of plants, a study which 
exceeded my own efforts because of the many inquiries and far reaching botanical knowledge, 
and a study in which I strove to unite more men with me in my effort." 



\ 



Meyer-Abich: Alexander von Humboldt 253 

So we see that his faculty to organize what we call teamwork today awoke 
early in Humboldt. 

Thinking this way in typological holisms we cannot but expect that Hum- 
boldt in his general philosophy of nature has also been a holist and neither a 
mechanist nor a vitalist. To document that we have an early treatise which 
Humboldt wrote for Schiller's ''Die Horen'' where it appeared in 1795, in Hum- 
boldt's 25th year of hfe. He pubhshed this essay, titled ''Der Rhodische Genius'' 
later also in all editions of his most popular and beloved book, the "Ansichten der 
Natur;' wherefore we must take it for granted that his viewpoint about the 
philosophy of living nature published here was never abandoned by him. Here 
Humboldt describes what was then called "Lebenskraff (vital force) with the 
following words: ''In der toten unorganischen Materie ist trdge Ruhe, solange die 
Bande der V erwandtschaft nicht gelost werden, solange ein dritter Stoff nicht 
eindringt, um sich den vorigen beizugesellen. Aber auch auf diese Storung folgt 
bald wieder unfruchtbare Ruhe^^^ In this way Humboldt describes what we 
defined above as physicochemical equilibria only. But then he continues: "An- 
ders ist die Mischung derselben Stoff e im Tier- und Pflanzenkorper. Hier tritt die 
Lebenskraft gebieterisch in ihre Rechte; sie kummert sich nicht um die demo- 
kritische Freundschaft und Feindschaft der Atome; sie vereinigt Stoffe, die in der 
unbelebten Natur sich ewig fliehen, und trennt, was in dieser sich unaufhaltsam 
suchtr^^ That is the kind of activity the principle of living harmony realizes. 
Using the word Lebenskraft to describe it does not mean vitaHsm. In his principal 
physiological book, "Versuche uber die gereizte Muskel- und Nervenfaser nebst 
Vermutungen Uber den chemischen Prozess des Lebens in der Tier- und Pflanz- 
enwelt;' from the year 1797 Humboldt rejects his Lebenskraft as a vitalistic 
principle, not considering it necessary "eine eigene Kraft zu nennen, was viel- 
leicht Moss durch das Zusammenwirken der im einzelnen Idngst bekannten 
materiellen Krdfte bewirkt werdeJ'-'' But such an antivitahstic statement does 
not mean that Humboldt has been transformed now into a mechanist. Not at all! 
He continues to be a holist as in his "Rhodischer Genius:' Even that "Zusammen- 
wirken der im einzelnen Idngst bekannten materiellen Krdfte" inside any hving 
substance is so characteristically of the organismic world that there exists nothmg 
comparable to it in any physiochemical system. The relation between the purely 
physical and the truly living is here thought of just the way Schelling thought of 
it when he stated: "Nicht, wo kein Mechanismus ist, ist Organismus, sondern 
umgekehrt, wo kein Organismus ist, ist Mechanismus.""^^ That is true holism, 
even in the modern form of it. We are of the opinion that there is only one total 



18 "Inert activity characterizes unorganized matter as long as the bonds of physical attraction 
remain in force, so long as no third element penetrates in order to associate itself with the other 
elements. However, unproductive inactivity again soon follows this disturbing penetration." 

19 "The composition of the same substance is different in plants and animals. Here the life- 
force enters as dominant in its sphere; it does not concern itself with the Democritic attraction 
and repulsion of atoms; it unites matter which rushes eternally in inanimate nature, and it 
separates that v/hich would seem to be inseparable." 

20 ". . . to term a special power that which perhaps would result merely from the combined 
efforts of material forces which are separately well known." 

21 "It is not correct to say: Where no mechanism exists, there is organism. Rather, the reverse: 
Where no organism is to be found, there is mechanism." 



254 The University of Texas Publication 

reality, which comprehends Hkewise the physical and the organic world. But the 
physicochemical principles and laws are not sufficient to cover also the organ- 
ismic reality in its totality. So we have to research the principles and laws of the 
organismic reality in the same autonomous way that we did also in physics. But 
having once discovered the biological laws, we are able to derive from them by 
logical simplification the corresponding physical laws. The inverse way, intended 
by the mechanistic philosophy of nature, is impossible, but also the vitalistic 
thesis of an essential dualism between the physical and the organismic. That is 
holism, which almost generally has been accepted by the great scientists of 
Goethe's time, not only by Goethe and Humboldt, but also by Carus, Joh. Muel- 
ler, K. E. von Baer and many other naturalists. And here, to say it again, is the 
historical meeting point where our own present science and natural philosophy 
encounters anew the Goethe-Humboldt-Time. 

Now we are sufficiently prepared to determine exactly Humboldt's position in 
the history of modern physiology. Since its foundation during the Renaissance 
epoch we must distinguish two main trends in the historical development of 
modern physiological thinking and research. One of them is absolutely new and 
original, created by the same spirit of modern causal-mathematical science 
which also shaped the modern dynamics and physics of Galileo and Newton. This 
is the new Physiology of Harvey, based, of course, upon the equally new ana- 
lytical and elementaristic Anatomy of Vesalius. The general characters of this 
absolutely new modern physiology are the same which are also borne by modern 
physics, namely dynamic, analytic, elementaristic and causal-mathematic. To- 
day we call this group of sciences the exact-mathematical-sciences. To all of them 
is Kant's statement valid, that any true science comprehends only so much real 
science as its mathematics content. The most evolved principle to create this kind 
of science is the "causal mechanism" mentioned above. 

For Humboldt, as a holist, this kind of physiology was not, of course, con- 
venient. Only one of its main features did also suit him — the mathematical 
treatment of its problems. Unlike Goethe, who disliked the newtonian way of 
science, that is, the most intimate correlation and collaboration between mathe- 
matics and science, Humboldt belonged to the most ardent adherents of this 
modern synthesis of mathematics with the study of nature. Particularly by over- 
coming its very deficiency, Humboldt made Goethe's science capable of meeting 
our actual scientific situation and helpful to us in establishing our own new 
ideals of knowledge. But first we must outhne the second historical trend of 
modern physiology, with which Humboldt has also been closely associated. 

This kind of physiology we have to characterize as comparative typological 
physiology. Its roots go down into aristotelian biology; therefore it is still outside 
the reach of causal thinking, and because of its comparative proceeding, also free 
of elementarism. Since the Renaissance this classical typological physiology ac- 
quired a new aspect, as often occurs with old manners when they come into 
contact and historical struggle with absolutely new ideologies. From the new 
mathematical physics our typological physiology took over the dynamic char- 
acter and behavior. So the static platonic and strictly kinetic aristotelian typology 
has become, since the Renaissance, a steadily and permanently always more 
truly dynamic science like modern physics. This becoming dynamic of an orig- 



Meyer-Abich: Alexander von Humboldt 255 

inally static structure we can recognize historically in Linne's famous distinction 
between "good," i.e. static, and "bad," i.e. dynamic, species and also in Goethe's 
concept of his "Urpflanze'' as a really existing individual. But in Goethe's 
"general type" he acquired definitely the dynamic type, which since then has 
been completely and perfectly conceived by Humboldt and Cuvier. 

This dynamic and comparative typological physiology has now become the 
second fundamental tendency, developing throughout the history of modern 
biology side-by-side with the absolutely new causal-elementaristic physiology. 
Both trends meet each other not seldom in the same important physiologists as m 
Haller and Joh. Miiller. But our typological physiology is realizing its own inde- 
pendent development; it begins its modern dynamic way in Albrecht von Haller. 
In his principal work, the "Memoires sur la nature sensible et irritable des parties 
du corps animar (4 Vols., Lausanne 1756-60), he states that sensibihty and irri- 
tabihty represent specific and autonomous faculties of the living substance, and 
for which there do not exist any corresponding properties in non-living matter. 
By these carefully exercised comparative physiological experiments, Haller de- 
feated definitely the first mechanistic theory about the living substance, namely 
the theory of Borelli, Baglivi, and others, which undertook to prove that all ani- 
mal organs are nothing but simple mechanical instruments: heart: pump; lungs: 
bellows; glands: sieves; teeth: scissors, etc. Haller's theory belongs to the true 
typological statements because it is only describing physiological functions as 
they are, by no means explaining them the causal way as the mechanistic theory 
from Baglivi really did it. In the following time other important physiologists, 
following Haller's method and way of consideration, stated, from more and more 
animal organs, that they too are exercising specific organismic facuhies which do 
not exist at all in non-living matter. The final conclusions of all this comparative 
dynamic-typological physiology made Joh. Miiller express his well known theory 
of the "specific energy" of all living substance in all its different structures from 
the simplest cell to the most comphcated organ. Based upon this absolutely typo- 
logical and by no means causal theory, any Hving structure exercises its own 
"specific energy", wherefore in the pure physicochemical reality no comparable 
processes exist. The fundamental faculty of all living specific energies is the 
first discovered by Haller: irritability. One can illustrate its function by a known 
example of Joh. Miiller's: Living skin reacts to burning differently than dead 
skin. If dead skin burns, it is transformed by pure chemical processes into other 
chemical substances; but if living skin is burned, it reacts in the specific living 
way by inflammation, which finally brings about additional new living skm. 
Only organismic "specific energies" act this way, if they are irritated physio- 
logically. This kind of typological comparative physiology is still continuing in 
our own days, its last significant presentation being that of Jakob von Uexkiill's 
so-called "Umweltlehre'' (Doctrine of the environment of the organisms). This 
part of comparative physiology represents nothing but the consequent continua- 
tion of Haller and Joh. Miiller from the field of organ-physiology into the modern 
ecological physiology. 

Humboldt also has his very definite position inside this line of physiological 
development and research, in between Haller and Joh. Miiller. He describes the 
general task of his above-quoted book on physiology with the following words: 



256 The University of Texas Publication 

'Teh habe gesucht^ in der nachstehenden Abhandlung alles zusammenzudrdngen, 
was ich bisher iiber Reiz und Reizempfdnglichkeit der sensiblen und irritablen 
Fiber beobachtete,'' and in his physiological biograph Wilhelm Wundt (Bruhns 
1872) refers to Humboldt's results with the following words: ''Mit der Unter- 
suchung der reizbaren Pflanzen beginnend, entwirft hier Humboldt ein fUr die 
damaligen Kenntnisse umfassendes Bild der Reizungscheinungen in der ganzen 
belebten Natur. Wiirmer^ Mollusken, Insekten, Fische^ zahlreiche Amphibien, 
Vogel und Sdugetiere unterwirft er der Vivisektion, dem galvanischen und 
mechanischen Reizversuch. Ueberzeugt von der inneren Uebereinstimmung oiler 
Organisation^ vermutet er^ dass die Reizbewegungen der Mimose und anderer 
Pflanzen^ wenn gleich der galvanische Reiz bei ihnen unwirksam bleibt, auf den 
ndmlichen Ursachen beruhen wie die Zusammenziehungen der tierischen Mus- 
kelfaser."- All this is indeed very true comparative-typological physiology. The 
idea of the "innere Uebereinstimmung aller Organisation'' is nothing but an- 
other word for Goethe's "general anatomical type." How near Humboldt in his 
final physiological conclusions comes to the general principle of the "specific 
energy" of Joh. Miiller is cleared up in the following remarks: 'Teh fange von der 
Erscheinung des Galvanismus an, weil ich durch die Art, wie ich die Versuche 
anstellte, unwidersprechlich erweisen zu konnen glaube, dass der Stimulus in 
diesem wunderbaren Phdnomen grossenteils von den belebten Organen selbst 
ausgeht, und das diese sich dabei keineswegs bloss leidend, etwa als elektro- 
skopische Substanzen, verhalteny"^ This remark about the own activity of the 
living organs when they meet with irritations from outside refers exactly to Joh. 
Miiller's "specific energy" whose fundamental faculty is, of course, vital activity. 
The typological comparative physiology in all its presentations from Haller 
until von Uexkiill still lacks the application of mathematical thinking and there- 
fore the characteristic modern synthesis of being a mathematicised science. To 
become a mathematicised science any doctrine has to fulfill two different but of 
course intimately correlated indispensable requisites. First of all it has to present 
its facts as exact measurements following the postulate of Galileo "to measure 
what is measurable, and to make measurable that which presently we are not yet 
able to measure." Second, we have to establish a causal principle which allows 
the application of mathematical calculi on measured facts to derive from them 
new, unknown statements. The modern causal-analytical physiology has taken 
precisely this way in its historical development. Harvey has been the first to 



22 "I have tried to bring together in the following study all that which I have observed con- 
cerning the stimulation and the susceptibility to stimulation of sensitive fibers. . . . Beginning 
with research on sensitive plants, Humbolt here sketches what was for that time a comprehensive 
picture of all living things which were capable of receiving stimuli. He subjects worms, mollusks, 
msects, fish, numerous amphibians, birds and mammals to vivisection in order to test for 
galvanic and mechanical response. Convinced of the inner agreement of all organization, he 
conjectured that the response of the mimosa and other plants depends upon the same causes as the 
coordinate functioning of muscle fiber in animals, also if the galvanic stimulus remains ineffec- 
tual for the plants." 

2'^ "I begin with the appearance of galvanism because I believe that in this manner I could 
prove without contradiction that the stimulus in this wonderful phenomenon arises in great 
measure from the living organs, and that this is in no way merely passive like electroscopic 
substance." 



Meyer- Abich: Alexander von Humboldt 257 

make measurable the then most significant basic phenomenon of all physiology, 
the circular movement of the blood, and Descartes has delivered the most im- 
portant philosophy for modern science since its beginnings, namely his mechanis- 
tic metaphysics, which postulates the indentity of physical and organismic reality, 
thus making possible the scientific treatment of all biological phenomena as 
essentially being only physicochemical structures too. As we know, this causal 
principle of the "mechanisms" has had the biggest success any philosophical idea 
in science has ever acquired by shaping not only modern physics, chemistry and 
physiology, but also the whole corresponding technology upon which all our 
actual human civihzation is based. But we know also that the profound crisis of 
our social, mental and political life has one of its reasons in the historical fact 
that this classical mechanistic philosophy is by no means any longer able to cover 
the most urgent and imperative spiritual necessities of mankind. We need a new 
philosophy, able to deliver a new ideal of knowledge and a corresponding prin- 
ciple of causality for all science to replace the theoretically exhausted causal 
principle of mechanism without, at the same time, abolishing its really great and 
stupendous acquisitions. 

Humboldt, being a hoHst, could not of course follow modern physiology in its 
mechanistic way. But its first requisite, to establish all its facts in form of exact 
measurement, of the so-called "constants of nature," has been a way to exercise 
science which corresponds intimately with Humboldt's own ideal of knowledge 
and in which he surpassed also his friend Goethe, as we have noted at different 
times. And not only in his particular physiological research concerning irrita- 
bility problems did he apply measuring methods, whenever possible, but also in 
his beloved plant geography he introduced statistical mathematics. Perhaps the 
most famous botanist of Humboldt's time was Alexander Brown, who first inves- 
tigated the geographical distribution of plant species and families in a given 
formation of vegetation, counting the species and obtaining in this way an 
exact correlation between the natural system of plants and its given geograph- 
ical distribution in the different floral provinces and formations. This method 
suited Humboldt best and, in elaborating the results of his botanical research 
in tropical America he applied it in the description of some plant formations 
visited by him and Bonpland. He wrote also a particular treatise about this 
kind of "botanical arithmetic" as he called it, which appeared in 1817 under the 
title: ''De distributione geographica plantarum, secundum coeli tem.periem et 
altitudinem montium, Prolegomena.'' That time this statistical treatment of the 
problems of plant geography did not provide us with important results, but that 
is only due to the small number of taxonomic species known during that time. 
Probably in the future the same statistical processes in biogeography will bring 
us very important knowledge intimately related to the results of modern geo- 
botany and geochemistry (Vernadsky, et al.). In any case Humboldt has proved 
that combining typological physiology and morphology with exact mathematical 
treatment by measuring and statistics is absolutely not only possible but an 
urgent necessity. 

To transform that way the typological comparative physiology into a new 
causal-mathematical physiology we need also the second mentioned requisite of 
modern mathematicised science, namely the invention of a new ideal of knowl- 



258 The University of Texas Publication 

edge with its corresponding causal principle. If the holistic philosophy of nature 
and its corresponding "causal principle" (Smuts) of "holisms" are able to guide 
us in that direction, the next coming future has to prove it. Essentially hereby 
would be, if the holistic ideal of knowledge will have the faculty to produce new 
holistic mathematical calculi able to synthesize holisms into mathematical the- 
ories. The mechanistic idea doubtless has fulfilled such a great task, having 
synthesized the mechanisms of nature in its grandiose new mathematical calculi 
of the infinite-simalanalysis and analytic geometry, autonomously shaped by its 
own power of knowledge. But actually they are no more sufficient to cover also 
the requisites of future science. 

Historians are not allowed to make prophecies. They normally are too much 
attached to distinct epochs and tendencies of the past to have still a free and 
unbiased look into the future. All they have to do, and only they can do it, is to 
provide the men who are actively shaping the future with a better understanding 
of the past; for it is the past which determines the future much more than the 
immediate present time. In such a sense I am of the opinion that Humboldt's 
science will have a certain importance to guide us in shaping our own future in 
science and knowledge. 

LITERATURE CITED 

Alexander von Humboldt: Voyage aux regions equinoctiales du Nouveau Continent, fait en 
1799, 1800, 1801, 1802, 1803 et 1804 par Alexandre de Humboldt et Aime Bonpland, redige 
par Alexandre de Humboldt. Grande edition. Vols I-XXX, Paris 1807-1834. 

: Ideen zu einer Geographie der Pflanzen, nebst einem Gemalde der Tropenlander, 

auf Beobachtungen und Messungen gegriindet, Tubingen 1807. 

: De distributione geographica plantarum, Prolegomena, Lutetiae Parisiorum et 



Lubeck 1817. 

: Versuche iiber die gereizte Muskel- und Nervenfaser Bd. 1, Posen, Berlin 1797. 

: Ansichten der Natur, mit wissenschaftlichen Erlauterungen, Bde. 1, 2, Stuttgart u. 



Tiibingen, 1. Aufl. 1808, 3. Aufl. 1849. 

: Kosmos, Entwurf einer physischen Weltbeschreibung, 5 Bde., Stuttgart 1845-62. 

Johann Wolfgang Goethe: Gedenkausgabe der Werke, Briefe und Gesprache 28. August 1949: 
Bde. 16, 17; Naturwissenschaftliche Schriften T. 1. u. 2; Bd. 24: Goethes Gesprache mit 
Eckermann, Ziirich 1949-52. 

Heinrich Berghaus: Brief wechsel Alexander von Humboldts mit Heinrich Berghaus, Bd. 1, 
Leipzig 1863. 

Karl Bruhns: Alexander von Humboldt. Eine wissenschaftliche Biographie im Verein mit R. 
Ave-Lallement, J. V. Carus u.a. bearb. u. hrsg., Bde. 1-3, Leipzig 1872. 

Christian von Ehrenfels: Ueber Gestaltqualitaten, Vierteljahrsschr. f. wiss. Philosophie, Jg. 14, 

1890. 

Arthur March: Die physikalische Erkenntnis und ihre Grenzen, Braunschweig 1955. 
Adolf Meyer- Abich: Naturphilosophie auf neuen Wegen, Stuttgart 1949. 

: Biologie der Goethezeit, Stuttgart 1949. 

Johannes Miiller: Ueber die phantastischen Gesichtserscheinungen, Bonn 1826. 
Adolf Naef: Idealistische Morphologie und Phylogenetik, Jena 1919. 



Meyer -Abich: Alexander von Humboldt 259 

Richard Owen: On the archetype and homologios of the vertebrate skebton, London 1848. 

F. W. J. von Schelling: Von der Weltseele, eine Hypothese der hoheren Physik zur Erklarung 

des allgemeinen Organismus, 1798. 
Jan Christian Smuts: Holism and Evolution, 2. ed. London 1927. 
W. J. Vernadsky: Geochemie, iibers. von E. Kordes, Leipzig 1930. 



Molecular Configuration, Synthesis and Gene Action 

R. p. WAGNER 

Genetics Laboratory, Department of Zoology 
The University of Texas 

The thoughts and observations expressed here are a logical result of Professor 
Patterson's having urged me to study the natural nutrition of the fruit fly in 1940. 
I will not elaborate on the sequence of events that transformed a fruit fly ecolo- 
gist into a macromolecule ecologist, but I do want to make clear my indebtedness 
to him. Once he got me started, he washed his hands, and only hoped that some- 
thing good might come of it. Although he was not always certain as to what I was 
trying to do (in which feeling I frequently had to join with him), he never 
faltered with his support and encouragement. 

The search for organismal units of structure and function in biology has led 
us to the recognition of organs, cells, chromosomes, mitochondria, plastids, macro- 
molecules, etc. All of these have the quality either of being directly observable 
with eye or instrument, or at least of being made known to us as physical entities 
by physical or chemical techniques which are known to be reasonably depend- 
able. On the other hand, we have also encountered some difficult problems in this 
quest for units. The foremost of these is the search for the hereditary units which 
segregate in definite ratios. In times past these have been called factors (Bate- 
son, 1909), differentiating characters (Mendel, 1866), genes (Johannsen, 1911). 
cistrons or recons (Benzer, 1957), etc. But the same general difficulty has been 
evident from the very beginning of genetics, when we attempt to apply precise 
definitions to these entities or concepts. It is nearly always found impossible to 
define satisfactorily what segregates by what is observed. Several significant 
contributions in this direction have been made. First we may cite the formulation 
of genotype by Johannsen (1911) whereby he essentially separated what is 
transmitted through the gametes from the determined characters. These are 
necessary terms for development of causal genetics as well as formal genetics. 
The term, gene, has proved very valuable for several reasons. It served to replace 
the older term, factor, which was used ambiguously. Gradually gene came to 
mean determiner that is transmitted from generation to generation, as opposed 
to what is determined. The gene theory gave it a locus on the chromosome and 
defined its boundaries by crossingover. By implication it became a unit of func- 
tion because when it mutated it caused a specific change in the phenotype in- 
herited as a unit change. For many years it has served as a satisfactory genetic 
unit — a unit of physical structure on the chromosome and a unit of function and 
mutation. 

Various geneticists, particularly Goldschmidt (1951), have attacked the gene 
concept, usually on the grounds that position effect, in particular the variegated 
type, as found in Drosophila, would indicate that an order of organization exists 
on the chromosomes that makes the gene unit an unnecessary assumption. More 
recently the concept of the gene has come under even stronger criticism because 



262 The University of Texas Publication 

of the recognition of the undoubted widespread distribution of pseudoalleles. The 
phenomena of cis-trans position effect (Lewis, 1955), complementation (Wood- 
ward, Partridge and Giles, 1958), etc. taken together with the fact that closely 
linked pseudoalleles have similar or related functions have necessitated a com- 
plete reexamination of what we mean by gene (Pontecorvo, 1958). In fact it is 
now quite evident that the gene as a unit as defined by previous criteria particu- 
larly as a crossover unit probably does not exist. It is necessary to redefine the 
term or abandon it entirely. 

It is my purpose here to point out some facts and theories and to attempt to 
indicate a possible new approach to the problem. Instead of taking the usual 
approach of starting in the nucleus and proceeding outward I believe it might be 
more rewarding to start in the cytoplasm, and work back into the nucleus. 

PROTEINS 

The question we ask ourselves is this: Can we use our present theories and 
understanding of protein structure and biosynthesis to help us understand the 
structure of genetic material and the consequences of its functioning? Implicit 
in this approach is the important concept that these macromolecules constitute 
units of function in the cytoplasm and that their specificity is in some way 
directly or indirectly conferred upon them by the genetic material. 

First, what are the relevant facts about proteins and their structure, function 
and biosynthesis? In summary this seems to be the picture. Proteins are macro- 
molecules of molecular weights ranging from several thousand to about a mil- 
lion. They are polypeptide chains of amino acids with secondary foldings. They 
function as "work horses," as enzymes, transporting agents (for example, oxy- 
gen or electron carriers), regulatory agents, (for example, hormones) or an- 
tibodies. In these functions they show a remarkably high degree of specificity 
of function. It now appears probable that part or all of this specificity basically 
resides in the sequential order of the amino acid residues that make up the chain. 
These sequences in turn probably determine the types of folds held in place by 
disulfide and hydrogen bonds which give the protein molecule its active con- 
figuration. Hence, the different orders of the 20 odd amino acids known to be 
involved in proteins can theoretically account for the many different kinds of 
proteins which exist or have existed. One important detail in this connection is 
that those proteins from a single source, such as insulin, ribonuclease, etc., in 
which amino acid sequences have been determined exist as chemical entities of a 
single species represented by a single type of order, and not as populations of 
different but related species of molecules. This does not mean that mixtures of 
protein species may not exist, such as in heterozygotes, but simply that within 
each species homogeneity is practically absolute. Nor does it mean that famihes 
of related proteins do not exist. It is clear that they do. Insulin from cow, sheep, 
horse or pig all have shght differences in amino acid order, but yet they are all 
obviously related insofar as they all have the physiological effect of insulin and 
all have identical amino acid sequences in the rest of their chains (Harris, 
Sanger, and Naughton, 1956). 

How is the function of a protein molecule related to its structure? This is a 



Wagner: Gene Action 263 

problem still under extensive investigation. It is clear that many proteins, par- 
ticularly those that act as enzymes or carriers, are associated with prosthetic 
groups or cofactors such as organic coenzymes, metal ions, or porphyrins such 
as heme. These non-protein groups are of extreme importance in determining the 
activity of the total protein-cofactor complex. But in addition to the cofactors, 
other aspects of the functional molecule complex must be important. For one 
thing, the same cofactor may be involved with a number of different proteins to 
produce as many different enzymes or carriers. The protein portion is basically 
an important part of the complex, therefore, since it endows the molecule with its 
specificity. It is generally assumed that this portion has a region or center called 
the "active center" which is most closely related to the relevant function of the 
molecule. This may be the site at which the coenzyme attaches, plus contiguous 
areas which dictate specificity of function. The active center may be a relatively 
restricted portion of the molecule surface or chain. It appears that in many cases 
parts of the chain of a biologically active protein are expendable when tested 
in vitro. For example, only one-third of the papain molecule is necessary for its 
activity. Up to 120 of its 180 residues may be removed without loss of activity 
(Hill and Smith, 1956) . Also, as stated above, certain substitutions may be made 
in the insulin chain without altering the general function. The differences be- 
tween hemoglobin A, S, and C discussed below are another example. 

What is the function of the part of the protein molecule not included in the 
active center? It has been argued that the residual part of a protein is (1) a 
handle or holdfast which attaches a molecule in its proper position within the 
cell, (2) a necessary addendum as an energy transfer device, (3) a means to 
maintain stability and specificity of the active center (Steinberg and Mihalyi. 
1957), or (4) a non-essential residuum formerly functional but now merely an 
appendix which may provide clues to the molecule's evolution but otherwise un- 
important (Anfinsen and Redfield, 1956). There may be an element of truth in 
each of these, but I would add a more general statement already anticipated by 
others. The residual part of the polypeptide chain acts as a buffer between the 
active center and the environment of the cell. This includes the external and in- 
ternal environment of the cell, and, within the cell, includes molecules ranging 
from inorganic ions to proteins and other macromolecular constituents. Merely 
having an active center is not enough. The active center must be protected from 
inhibitors which are otherwise natural and necessary constituents of the cell. The 
residual chain may do this by the way it is folded adjacent to the active center 
and the way in which it is folded would be directly related to the amino acid 
sequence in the chain. The substitution of a single amino acid in a chain of one 
hundred could conceivably alter the configuration of the entire molecule. 

GENETIC IMPLICATIONS 

Several conclusions may be drawn from the above observations relative to gene 
action and phenotypic expression provided one is willing to make a number of 
reasonable assumptions. If we assume that the total protein part of an enzyme or 
carrier is necessary under actual cellular conditions, and that a large part of it is 
really an environmentally selected-for buffer, we have at hand an explanation 



-64 The University of Texas Publication 

for allelic gene differences, and the action of suppressors, enhancers, inhibitors, 
dilutors, and various unnamed modifiers in general. Present research strongly 
indicates that a gene mutation can modify the structure of an active protein by 
alteration of its amino acid sequence. This may involve only the substitution of a 
single amino acid. Such a change can conceivably occur within the active center, 
or in the residual part and may result in a protein which is worthless under any 
conditions. On the other hand, the new protein may be active, but only under 
certain conditions of the environment. 

The hemoglobin studies of Ingram (1957) (Hunt and Ingram, 1958) indicate 
that the mutant hemoglobins C and S each differ from the normal A by the sub- 
stitution of a single amino acid residue. In that position in the globin chain at 
which is found a glutamic acid residue in A there is substituted a lysine residue 
m C and a valine in S. Both C and S function poorly as oxygen carriers in the red 
blood cells, but for S, at least, the oxygen dissociation curve determined at 20° C 
and pH 7 is the same for the purified hemoglobins from both S and A individuals 
(Jeffries and Allen, 1951). This indicates that the active center has not been af- 
fected drastically, but that under certain environmental conditions, i.e., within 
the intact red corpuscles, the active center of hemoglobin S is not as efficient as 
the active center in A, but under other conditions it may be just as active. This 
may be due to a change either in the active center or in the residual part. 

Turning to proteins with enzyme activities we find a number of examples of 
apparent enzyme alterations accompanying a single gene change. Yura (1959) 
reports that the enzyme, pyrroline-5-carboxylate reductase, (which catalyses the 
conversion of pyrroline carboxylic acid to proline), is present in a mutant of 
Neurospora blocked at this step in the biosynthesis of proline. The activity of 
this enzyme in the mutant, however, is about 30-fold less than the wild type. No 
significant difference was found between the enzyme from the mutant and the 
wild type with respect to relative affinity for the substrate, effective pH, or frac- 
tionation behavior, but they were strikingly different with respect to activation 
energy and thermostability. Mutant reductase has a temperature coefficient, Q,o, 
of 4.1, and a half-life at 48° C of 1.3 minutes. These values differ significantly 
from a Q^o of 1.6 and a half -life of 21 minutes found for wild type. The three en- 
zymatic differences, low relative specific activity, high Q,o and low temperature 
stability, segregate as single gene differences with the phenotype prolineless. 

Horowitz and Fling (1956) and Gest and Horowitz (1958) have demonstrated 
a difference between two tyrosinases in Neurospora apparently inherited through 
a pair of allelic genes T^ and T^. T^ determines the presence of a stable tyrosi- 
nase, and T^ a thermolabile tyrosinase. In addition, the thermostable and thermo- 
labile enzymes appear to be quantitatively different with respect to stabilization 
by Na+ and K+. Both the thermostable and the thermolabile enzymes are 
formed in T^ + T^ heterocaryons. 

Maas and Davis (1952) and Fincham (1957) have found significant differ- 
ences in enzymes formed by strains which reverted from the mutant to a strain 
similar but not identical to wild type. Their results cannot be as clearly inter- 
preted as those given for the reductase and tyrosinase, but they, along with 
numerous observations such as those made by Stadler and Yanofsky (1959) on 
tryptophan revertants in Escherichia coli^ make it evident that allelic genes prob- 



Wagner: Gene Action 265 

ably produce qualitatively different enzymes or proteins in general. Whether 
these are active or not depends upon environmental factors. For example, a 
thermolabile enzyme would be ineffective at a high growth temperature but 
effective at a low temperature (Maas and Davis, 1952); an inhibited enzyme 
would be active in the absence or in the presence of reduced amounts of the 
inhibitor, etc. (Wagner and Haddox, 1951 ) . 

Some of the work on the trytophanless mutants of Neurospora supports this 
latter conclusion directly. Tryptophan synthetase, the enzyme which forms 
tryptophan from indole and serine, is inactive or absent in those mutants of 
Neurospora which require tryptophan for growth, and cannot use indole + 
serine to replace the tryptophan requirement. However, in many of the allelic 
mutants at the td locus, which apparently control the production of this enzyme, 
a protein is present which has the same immunological specificity as tryptophan 
synthetase isolated from wild type but none of its enzymatic activity. This indi- 
cates strongly that the mutant alleles are forming altered proteins which are non- 
functional in the Neurospora cellular environment in which they initially occur. 
These immunologically active but enzymatically inactive proteins are called 
CRM. 

Suskind and Kurek (1958, 1959) extracted and purified the CRM substance, or 
a protein which fractionated with it, from a strain {tdo^) which required trypto- 
phan when grown at 25° C. At this temperature there is no detectable tryptophan 
synthetase in the crude extracts. However, after fractionation, a highly active 
tryptophan synthetase is obtained. This enzyme from the mutant is inhibited by 
an inhibitor present in both the wild type and other td mutants. Significantly, 
the mutant enzyme is ten times more sensitive to the inhibitor than is the wild 
type. The inhibitory effect can be simulated by Zn+ + . 

It is clear from this example that mutations of a gene may result in a variety 
of different molecules, all functionally related — if they have a function at all — 
but able to act only under certain specific conditions of the environment. 

Observations such as these give us a clearer understanding of the possible dif- 
ferences between wild type and what Muller (1932) called hypomorphic and 
amorphic alleles. Amorphs produce no enzyme, or one which is completely in- 
activated by an inhibitor; hypomorphs produce enzymes which are partially 
inhibited, and produce less than the wild type phenotype. The dominant wild 
type produces an enzyme which is in approximate harmony with its environ- 
ment and its potential activity is probably in excess of available substrate. 

Turning now to the problems of gene interaction we can see immediately that 
the modification of the cellular environment by the mutation of one gene may 
have a drastic effect on the action of the product of another. Suppressor genes 
may modify the environment in such a way as to relieve the inhibition of an 
enzyme as in the mutant tryptophan synthetase discussed above (Suskind and 
Kurek, 1959). (See also Wagner and Haddox, 1951.) Such an interpretation is 
in agreement with the studies on the suppressed td mutants of Neurospora. Active 
tryptophan synthetase has been extracted and purified for a number of the sup- 
pressed mutants of Neurospora and found to have the same dissociation constants 
as the tryptophan synthetase produced by wild type. This might be expected if 



^^^ The University of Texas Publication 

the mutant protein was altered in an area of the polypeptide chain outside of the 
active center in the residual part. 

Furthermore this interpretation fits the seemingly paradoxical fact that sup- 
pressors can be both exceedingly specific and non-specific. A suppressor may 
suppress the mutant action of a single allele or a number of the alleles in the 
series, but not all (Yanofsky and Bonner, 1955). On the other hand it may be 
quite non-specific and suppress the mutant action of a number of non-allelic and 
seemingly functionally unrelated genes (E. B, Lewis, cited in Wagner and 
Mitchell, 1955). These phenomena can be expressed by assuming that genes, 
allelic or not, produce different kinds of protein subject to inhibition by the same 
or different inhibitors. Mutation of another gene controlling one of the inhibitors 
can therefore effect one or more of the proteins produced by allelic or non-allelic 
genes. Similar lines of reasoning can explain the action of inhibitor, enhancer, 
etc. genes. There remains to do the actual work to test this hypothesis rigorously, 
but Suskind and Kurek have made a good start. 

What does all of this have to do with the problem of the gene as an hereditary 
unit? If we assume that the total polypeptide chain making up a functional unit 
as discussed above is necessary for that function, then, we must further assume 
that whatever template pattern the protein sequence is derived from contains the 
necessary information to organize the synthesis of a long chain of amino acids in 
a precise sequence. The primary pattern is now generally assumed to be DNA in 
those organisms with both DNA and RNA. According to Crick and co-workers 
r Crick, 1958; Crick, Griffin and Orgel, 1957) probably a minimum of three ad- 
joining nucleotides is needed to determine one amino acid, and there are twenty 
arrangements of three out of sixty-four possible triplets from four possible nucleo- 
tides which do not give "nonsense" when arranged linearly side by side. (That 
the number 20 is also the number of amino acids which make up the bulk of pro- 
tein may be no mere coincidence. ) If a triplet is required to determine a single 
amino acid, then a protein containing 250 amino acid residues would require a 
nucleic acid template of 750 nucleotides per strand. Since it is possible that the 
cytoplasmic protein is formed from a RNA template obtained from the DNA 
pattern, it would be required that each protein have both DNA and RNA 
nucleotide chains with three times more nucleotides than the number of amino 
acids in the protein chain. 

If we assume that all of the protein chain is important to the functioning of 
the protein in vivo, then all the polynucleotide determining it is necessary, and 
its limits are dictated by the limits of the functional molecule in the cytoplasm. I 
would call this length of polynucleotide chain the gene. 

At the time of pairing of homologous elements, breaks and interchanges may 
occur within this gene as well as between it and adjoining genes, but interchanges 
within the gene will not be recognized, unless there are at least two different 
triplets involved in the two synapsing chains. Breaks may conceivably occur be- 
tween the nucleotides of a triplet as well as between the triplets; however, if the 
DNA or RNA is bound to protein as nucleoprotein most of the time, permanent 
breaks should be rare. When not bound to protein the triplets might be stabilized 
by the amino acids attracted to them which would allow breaks and interchanges 
between unlike but adjacent synapsing triplets only during the rare intervals 



Wagner: Gene Action 



267 



that amino acids are not present, and homologous breaks or very close breaks 
occur simultaneously. 

If instead of using the classical interpretation of the origin of interchanges by 
breaks and refusion of strands of synapsing chains, we use the "copy choice" 
explanation (Stent, 1958) as applied to virus recombinations, we may come to a 
similar but not identical conclusion. 

Stent (1958) has proposed that the Watson and Crick (1953) explanation of 
DNA replication must be modified. He proposes in its stead the hypothesis sup- 
ported by data from virus and bacteria that a DNA double helix acts as a tem- 
plate for the synthesis of a single RNA polynucleotide which then peels off and 
carries all the information present in the DNA chain. This chain may be stabi- 
Hzed by an adhering protein. Another DNA helix is formed from this RNA 
rather than directly from the mother DNA. First, however, Stent proposes that 
identical RNA from the same mother DNA. pair in what he calls identical du- 
plexes, presumably for purposes of stability. Another DNA helix is formed from 
one of the RNA strands of the duplex as the two strands peel apart. If two homo- 
logous but somewhat similar RNA's pair (as from two allelic DNA's) the DNA 
formed on the duplex has the choice (copy choice) of copying part from one and 
part from the other. This could result in recombinations which would be recog- 
nized, if at least two differences were present in the two RNA's in the duplex. 
Thus interchanges occur not directly between paired DNA strands, according to 
Stent, but indirectly through RNA. 

This hypothesis has the merit of fitting viral and bacterial data well, because 
complementary classes are not recovered from a single synaptic event. It does 
not, however, in its present form fit the data from higher organisms such as 
fungi, where complementary crossovers are regularly observed resultmg from a 
single zygote nucleus. 

Although this hypothesis may not explain interchanges in the basic genetic 
material in organisms above virus and bacteria as well as crossingover, it may be 
of some heuristic value to consider it as a possibility in the cytoplasmic synthesis 
of protein. Classically, recombinations have only been thought to occur in the 
genetic material, but why should they not also occur in the cytoplasm? The 
reason for advancing this hypothesis, which has already been anticipated by 
Woodward, Partridge and Giles (1958) in a somewhat different form, are sev- 
eral. There are a number of phenomena currently known, but not understood, 
occurring in heterozygotes or heterocaryons which may be explained on the as- 
sumption that recombinations occur within functional units in the cytoplasm at 
the time of replication. 

If we extend Stent's hypothesis for DNA replication from a RNA duplex to 
protein replication from similar duplexes in the cytoplasm then we have the 
basis for recombinations occurring in protein. We may imagine the following 
sequence of events. In heterozygotes there must be two kinds of DNA repre- 
senting a particular locus, each corresponding to a different allele. Therefore, 
there must be two kinds of RNA formed which get into the cytoplasm. Three 
kinds of duplexes may be formed from these; two different identical duplexes 
each representing one allele, and a duplex with non-identical but homologous 
RNA's. Protein polypeptide chains may be visualized as forming on this "hetero- 



^^8 The University of Texas Publication 

zygous" duplex following one strand of a duplex at a time, but at times switching 
from one to the other in copy choice fashion. If between the paired RNA strands 
of the duplex, there exist at least two differences, then there should be at least one 
protein type produced by copy choice interchange which is not formed by the 
identical duplexes. Hence, from certain heterozygotes we should expect more 
than the two expected types of a particular protein species. Indeed we do find 
such "hybrid substances" in dove hybrids (Miller, 1954), blood serum hetero- 
zygotes in man (Beam and Franklin, 1958), and rabbit (Cohn, 1958). Hybrid 
substances produced in the cytoplasm by this means could explain the type of 
allelic gene interaction such as exhibited by antimorphs (Muller, 1932) and 
mixomorphs (Wright, 1941). Also the phenomenon of over dominance (Hull, 
1946; Crow, 1948) may find a possible explanation here. It is only necessary to 
assume that in the case of antimorphic interaction in heteroz3^gotes the hybrid 
substance or substances formed are recombination proteins incapable of being 
active agents in the cellular environment, and furthermore that they inhibit the 
activity of the active unrecombined proteins by combining irreversibly with the 
substrate. This would explain why two different alleles may be active when each 
is homozygous, but less active or inactive in heterozygotes. Alternatively, over- 
dominance could be explained by assuming that the hybrid substance is superior 
in activity to either of the unrecombined proteins produced in the pure form in 
the homozygous strains. Furthermore, the phenomenon of complementation in 
which two mutant "allelic" genes produce a wild type effect in heterozygotes 
would fit the explanation for overdominance. This could applly to complementa- 
tion in both heterozygotes of higher forms and heterocaryons of organisms such 
as Neurospora. 

RECONS AND CISTRONS 

The scheme presented in the preceding paragraphs may do little more than 
add confusion to a now already confused situation. Only the accumulation of 
more data can be expected to partly clarify the situation. Meanwhile, it may be 
helpful to consider certain other phenomena and confuse things even more. 

Coiners of terms and phrases have been extremely active since genetics in- 
vaded the domain of virus and bacterium, and physicists and biochemists arrived 
on the scene to enlighten geneticists. Some of the new terms undoubtedly fill a 
definite need, and will become a permanent part of the biological vocabulary. 
They have been necessitated primarily by the fact that recombinations can occur 
within the limits of the genetic unit called the gene. Benzer (1957) proposed the 
term recon which he defined "as the element in the one dimensional array that 
is interchangeable (but not divisible) by genetic recombination." Demerec 
(1956) uses the term site with more or less the same meaning as recon. I would 
consider site and recon synonymous, and to fit them into the framework given 
above I would postulate either that a site is a single nucleotide (or pair) of a nu- 
cleic acid chain, or at most a triplet. With respect to protein a site determines an 
amino acid in the chain. A gene then is made up of the necessary series of sites to 
produce a functional protein. Alleles at a particular locus are the result of differ- 
ences in sites. Hence there may be "identical" or "non-identical alleles" (De- 
merec, 1956). Non-identical alleles may be expected to complement one another 



Wagner: Gene Action 269 

if a functional molecule can be derived by interchange as discussed above. 
Identical alleles obviously would be, under this hypothesis, unable to comple- 
ment one another. The term cistron (also coined by Benzer, 1957) was intro- 
duced, again with very good reason, to describe those situations in which closely 
linked genes with apparently related functions show the cis-trans position effect. 
A cistron is a group of pseudoalleles — genes which give a mutant phenotype 
when two are present in the trans configuration and heterozygous with their 
wild type alleles, i.e. when each mutant allele is on a different homologue of the 
chromosome pair. When both mutant genes are on the same chromosome, their 
respective wild type alleles give a wild phenotype. 

A variety of explanations can be given to explain this phenomenon (Lewis, 
1954; Wagner and Mitchell, 1955) which all boil down to the assumption that 
the adjacent genes are related in a sequentially polarized function and in order 
to carry out this function the wild type or effective alleles must be together. The 
cistron then may be looked upon as a supragenic unit. Conceptionally, no diffi- 
culties are encountered by adding this supra unit to the functionally defined 
gene unit described above. However we are then faced with the paradox that 
complementation may occur within a gene so defined but not between genes 
which are elements of the same cistron. Obviously more facts are needed here to 
clarify the relation between genes and cistrons. However, if we use the idea that 
interchanges may occur by copy choice in the cytoplasm to produce "recom- 
bined" proteins, we may find a way out. These interchanges are possible only 
between homologous gene products but not ordinarily between non-allelic genes 
or their products. Interchanges between genes will only be maintained in the 
"germ line." As such they will be recognized as crossovers, products of gene 
conversion, or just mutations. 

CONCLUSIONS 

The thesis presented in this paper, that we can only define the gene function- 
ally by its product, makes no pretense at including the solution to all our prob- 
lems. It is, I believe, one possible working hypothesis to be considered among 
others. It does have the merit of being testable. This is also bad for its longevity, 
for hypotheses which cannot be tested often last the longest and attract the most 
attention at symposia. 

The ancillary thesis that recombinations may occur at the cytoplasmic level by 
copy choice at the time of formation of active proteins has the merit of providing 
for a more flexible system in the cytoplasm. A number of protein types are capa- 
ble of being produced — more than the previously assumed maximum of two in 
heterozygotes. These may be selected for under changing conditions of environ- 
ment. Indeed their occurrence may be a possible explanation for the apparent 
selective advantage held by heterozygotes in populations. (See Patterson and 
Stone, 1952). 

In conclusion I must point out that Haldane (1958) raises an extremely inter- 
esting point related to the above discussion. He suggests first that "the word fac- 
tor be used for the cause of an observable difference which shows Mendelian 
segregation." He then points out that though we must consider the gene a ma- 



270 The University of Texas Publication 

terial structure, we cannot localize it precisely. Its limits are certainly not exactly 
definable. A factor may be defined as a change in a few nucleotides by substitu- 
tion or by loss or duplication of nucleotides. He then goes on to say "while then 
factors are units, but not necessarily or even usually real units, genes are not 
necessarily units. I do not go as far as Goldschmidt, and say that a gene can only 
be detected because it has mutated." 

I do not agree with all that Haldane states here. We must focus our attention 
not on differences, which Haldane essentially identifies as factors, but on the real 
working units which give us the phenotype. If these are proteins, as discussed 
above, then their specificity must come from some source of information which 
we can define as that part of the genetic material which carries that sufficient 
information. This is the gene, and I think it is definable. Sturtevant's (1951) 
statement still has the ring of truth. "There is no escape from the conclusions 
that chromosomes are regionally differentiated, physiologically as well as visibly 
under the microscope; that particular and identifiable regions are necessary for 
particular reactions in the organism; and finally that these particular regions 
behave as units in heredity — specifically in crossingover." With the possible 
exception of the last phrase, "specifically in crossingover," it is my contention 
that this statement still stands as essentially true. 

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Beam, A. G. and E. C. Franklin. 1958. Science 128: 596. 

Benzer, S. 1957. In A Symposium on the Chemical Basis of Heredity, ed. by W. D. McElroy and 

B. Glass. The Johns Hopkin's Press, Baltimore. 
Cohn, C. 1958. /. Immunology 80: 73. 

Crick, F. H. C. 1 958. Symposia of the Society for Experimental Biology 12: 1 38. 
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Maas, W. K. and B. D. Davis. 1952. Proc. Natl. Acad. Sci. 38: 785. 

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Wagner, R. P. and C. H. Haddox. 1951. Amer. Nat. 85: 319. 

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