Book - Sex and internal secretions (1961) 1
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SECTION A Biologic Basis of Sex
Genetic and Cytologic Foundations for Sex
John W. Gowen, Ph.D.
Department of Genetics, Iowa State University, Ames, Iowa
I. Basic Literature
In the first edition of Sex and Internal Secretions published in 1932, Bridges discussed the closely prescribed problem of the genetics of sex, particularly as it was related to one species, Drosophila melanogaster. The treatment was sharply focused on the advances made, chiefly through his own researches, in understanding the functions of the genetic and cytologic factors operating during embryologic development which ultimately establish the sex types. The emphasis was on gene action through interchromosomal balances as they may aff"cct sex expression. The second edition of 1939 brought this material up-to-date and, at the same time, offered a much broader treatment by the inclusion of accumulated evidence on how differentiation for sex comes about in other forms. There is no substitute for a careful reading of these two presentations as background material for a present day understanding of sex determination. The current presentation makes no attempt, except in the barest outline, to repeat this early material other than in those aspects which bear on the advances that have been made since the printing of the second edition.
Immediately following the publication of the first edition of Sex and Internal Secretions there was a resurgence of interest in the problems considered in that book. The resulting research led to notable advances in available knowledge. Seven years later the second edition was published. A second wave of accomplished research appeared. In the interim of the past 21 years extensive advances have been recorded, particularly in understanding the mechanisms of sex differentiation in plants as well as in many animals. Among others, the data on Melandrium, Asparagus, Rumex, and Spinacia have been of first importance. Further information on Drosophila, Habrobracon, and Xiphophorus has notably broadened our viewpoints. Beginnings have been made to a better understanding of the conditions for sex separation in bacteria, protozoa, bees, birds, goats, mice, and man. To these specific contributions may be added the basic advances in understanding the methods by which genes are transmitted from one generation to the next, accomplish their actions in development, and by which chromosomes reorganize and reconstruct gene groups.
The period has also been one of excellent monographic treatments on different phases of the subject. Wilson's The Cell in Development and Inheritance (1928) and earlier, Morgan's Heredity in Sex (1914), Schrader's The Sex Chromosomes (1928) and Goldschmidt's Lymantria (1934) retain their pre-eminence. To this list have now been added the publication of extensive tabulations of chromosomes of cultivated plants by Darlington and Janaki Ammal (1945), of animal chromosomes by Makino (1951 ), and the yearly index to plant chromosomes beginning with 1956, compiled by a world-wide editorial group and published by the University of North Carolina Press, Chapel Hill, North Carolina. Those volumes give access to the basic chromosomal constitutions and their sex relations for far more species than were heretofore available. Inheritance information has been made more accessible and at the same time in more detail. The tabulations of mutants observed in particular species as in D. melanogaster (Morgan, Bridges, and Sturtevant, 1925; Bridges and Brehme, 1944) have been followered by those of corn, mouse, domestic fowl, rat, rabbit, guinea pig, Habrobracon, and many other animal forms, together with similar tabulations for cereals and a number of other plant species. Books of particular interest include those of Hartmann. Die Sexualitdt (1956), White, Animal Cytology and Evolution (1954), Goldschmidt, Theoretical Genetics (1955,1, Hartmann and Bauer, Allgemeine Biologic { 1953) , and Tanaka, Genetics of Silkworms (1952), and special reviews in various volumes of Fortschritte der Zoologie (1 to 12) (Wiese, 1960). Basic normal development of Drosophila has been presented in convenient book form in papers by Cooper, Sonnenblick, Poulson, Bodenstein, Ferris, Miller and Spencer, Biology of Drosophila (Demerec Ed., 1950) . Special papers having a direct bearing on the subject matter include particularly those of Pipkin (19401960) in analyzing the various chromosomes of Drosophila for sex loci, of Tanaka (1953) and Yokoyama (1959) for their studies and reviews of the genetics of silkworms, and of Westergaard (1958) on sex determination in dioecious flowering plants.
II. Mechanistic Interpretations of Sex
The records of search for basic mechanisms involved in the determination of sex have been foremost in the writings of man from the beginning of historical record. These ideas have included all forms of mechanisms both intrinsic and extrinsic to the organism. The most constructive advance, although not realized at the time, came through the recognition of cell structure, chromosome maturation and the discrete behavior of the inheritance. Differences in clu-omosome behavior were noted but not alwaj^s related to facts of broader significance. The period was one of expanding observations and development of ideas as they applied to species in general and also to the further complications which arose with more extended study. These complications included differences not only in one chromosome pair but in several. The significance of a balance between these chromosome types as well as with the environment was grasped by Goldschmidt and particularly by Bridges where his more favorable material brought out sharper contrasts in types and in the chromosome behavior. Ideas related to chromosome balance as they may affect developmental processes were developed. Goldschmidt emphasized that this balance could include factors in the cytoplasm as well as in the chromatin material. Bridges' observations on the other hand, pointed most strongly to the chromatin elements where changes in chromosome numbers were often accompanied by sharp differentiation of new sexual types.
Concepts of sex determination broadened. They came to include all the chromosomes in the haploid set, a genome, as a whole. The important concept of single chromosome difference being all important has been replaced with that of a balance between the chromosomes. It has sometimes been emphasized that it is this balance which is the most significant element in sex determination. Present day research seems to be pointing rather to even finer structures than the chromosomes in that the main search is on for the particular genes within the inheritance complex which contribute the characteristics of sex. Bridges' work has sometimes been asserted as committed to a particular type of balance in the chromosome. However, his writings 1 1932) make it clear that this balance is, in his judgment, basically due to the genes actually contained within the chromosomes rather than to the chromosomes themselves. This view is further emphasized in the second edition of Sex and Internal Secretions. "Sex determination in numerous forms with visible distinctions between the chromosome groups of the two sexes was at first interpreted on a 'quantitative' basis, as due to graded amounts of 'sex-chromatin.' But because even more species were encountered in which no visible chromosome difference was detected, the formula
tion was changed to include also 'qualitative differences in sex-chromatin.' Now, sex is being reinterpreted in terms of genes, with investigation by breeding tests penetrating to detail far beyond the reach of cytological investigation. The chromosomal differences are now treated as rough guides, and chromosomal determination is l)cing resolved into genie determination
Hence the difference in sex must be put on the same basis as that of any other organ differentiation in plant or animal, for example, the difference between legs and wings in birds— both modifications of ancestral limbs."
In contrast to what some have interpreted. Bridges was not committed to a specific chromosome balance, as for instance the X chromosomes to the A chromosomes in Drosophila, for all species. Rather he looked on that particular balance as but one mode of gene association in chromosomes that could bring about differences in gene action on development which would lead to sexual differentiation in its various forms. In that sense his ideas prepared him for, and were in conformity with, the types of sex differentiation which later became established in such forms as Melandrium and in the silkworm. In his concepts of sex determination and in his active interest in making genetics more specific, there can be little doubt that his theory would include those of the present and would also be welcome as refining and more sharply defining the significance of cytologic and genetic elements important to sex differentiation.
A. Concept of Sex Determination
As frequently happens, observations are made often before their significance is more than dimly understood. Background information may be insufficient for the facts to become clear. This was true when a lone chromosome was discovered as a part of the maturation complex in sperm formation of Pyrrhocoris. The association of the presence of this element with the idea of its being the arbitrator between male and female development in certain species was not made until 10 years later. Henking (1891) observed in Pyrrhocoris 11 paired chromosome elements and one unpaired element which after spermatocyte divisions led to sperm of two numerically different classes, one carrying 11 chromosomes plus the accessory chromosome and the other carrying only the 11 chromosomes. His observations were confirmed by several other investigators and in principle for other insect species over a decade following. The first suggestion that this chromosome behavior was related to sex diiTerentiation came from McClung's (1902) observations on the "accessory chromosome" of Xiphidiiim fasciatum. It is of some interest to examine the steps l)y which these conclusions were reached.
"The function exercised by the accessory chromosome is that it is the bearer of those qualities which pertain to the male organisms, primary among which is the faculty of producing sex cells that have the form of spermatozoa. I have been led to this belief by the favorable response which the element makes to the theoretical requirements conceivably inherent in any structure which might function as a sex determinant.
"These requirements, I should consider, are that: (a) The element should be chromosomic in character and subject to the laws governing the action of such structures, (b) Since it is to determine whether the germ cells are to grow into the passive, yolk-laden ova or into the minute motile spermatozoa, it should be present in all the forming cells until they are definitely established in the cycle of their development, (c) As the sexes exist normally in about (■(lual projoortions, it should be present in half the mature germ cells of the sex that bears it. (d» Such disposition of the element in the two forms of germ cells, paternal and maternal, should be made as to admit of the readiest response to the demands of enA'ironment i-egarding the ])i'oportion of the sexes, (el It should show variations in structure in accordance with the vai-iations of sex potentiality observable in different species, (f) In parthenogenesis its function would be assumed by the elements of a certain polar body. It is conceivable, in this regard, that another form of polar body miglit function as the non-determinant bearing germ cell." The important fact established by this reasoning was that a chromosome could be the visual differentiator between the fertilized eggs developing specific adult sexual differences. It w^as of little consequence perhaps that for sex itself, in this species, the chromosome arrangement was misinterpreted. The validity of the rules was probed through examinations of the cells of many species by many different observers.
B. Sex as Associated with Visible Chromosomal Differences
Variations in the chromosome complexes contributing to sexual differentiation were soon found. Probably the most frequent type observed in animals and plants was that in which a single accessory chromosome of the male was accompanied by, and paired with, another chromosome either of the same or of a different morphologic type. This other chromosome, as distinguished from the accessory chromosome now generally called the X, was designated the Y chromosome. For different species the Y ranged in size from complete absence, to much smaller, to equal to, to larger than the X. Besides the X and Y chromosomes, the autosomal or A chromosomes, the homomorpluc pairs, complete the species chromosome complement. In this terminology
Sperm (Y + A) + egg (X + A) = XY + 2A cells
of
lie determinino; tvpe
Sperm (X + A) + egg (X + A ) = 2X + 2A cells
of female detei'mining type
Variations in numbers and sizes of chromosomal pairs making up the autosomal sets of different species are familiar cytogenetic facts. These variations may extend from one pair to many chromosome pairs depending on the species. Similar variations may occur in chromosomes of eitluM- the X or Y types. The X is commonly a single chromosome but may be a compound of as many as 8 chromosomes in Aiicaris inciirva (doodricli. 1916). The Y mav be lacking entirely in some species or may be reduplicated in others. Males which form two types of sperm are often called heterozygous for sex, although the term digametic may be better, whereas the females are homogametic.
In other species the females show the chromosomal differences whereas the males are uniformly homogametic. These types are principally known in the Lepidoptera, more primitive Trichoptera, Amphibia, some species of Pisces, and Aves. The single accessory chromosome is present in the females whereas the males have two. It may l)e unpaired or be with another chromosome of different morphology and size. The sex types may be symbolized by designating the accessory chromosome by Z and its mate W as a means of separating possible differences between them and the X and Y chromosomes. The significance of these differences is not clearly established with the consequence that some investigators prefer to substitute the XY designation for the females and the XX condition for the males of these species. The zygotic formulae are:
Sperm ZA + egg ZA = 2Z + 2A male Sperm ZA + egg WA = ZW + 2A female
A third major chromosomal arrangement accompanying sex differentiation came to light as a result of Dzierzon's 1845 discovery of parthenogenesis in bees. Chromosomal and genetic studies have shown both Apis and Habrobracon of the Hymenoptera to be haploid, N, for each germinal cell in the male complex and diploid, 2N, in the female. N may stand for any number of chromosomes, as 16 for Apis or 10 for Habrobracon. The same chromosomal pattern characterizes, so far as known, the other genera of Hymenoptera. Haploidy vs. diploidy is viewed as the most obvious feature predicting differentiation toward the given sex type even though, as in Habrobracon, there is evidence for a particular chromosome of the N set carrying a locus for sex differentiating genes. The zygotic sex formulations are:
No sperm + egg N = N male Sperm N + egg N = 2X female
In development, as in some other species of widely diverse origins, chromosome polyploidy may take place causing the soma cells to differ from the germ cells in their chromosome coniponents.
The common phenotype for plants and lower animals has differentiated sex organs which are combined in the same individual. Plant species seldom depend on any but hermaphroditic types for their reproduction. Lewis' (1942) tabulation for British flora had but 8 per cent of all species depend on other than this form of reproduction. Those that had perfected dioecious systems were not all alike in the system adopted, although species with XY + 2A males and XX + 2A females were in high frequency. Dioecious reproduction was rarely the system common to a whole genus. Recent and multiple origins of dioecious types are indicated by the irregular distribution of species with bisexual reproduction within the different genera and families. Methods for preventing inbreeding have taken other channels as selfsterility genes reminiscent of fertility or sex alleles found in the older bacteria and protozoa. Animals of the lower phyla are those which are most frequently hermaphroditic. Within hermaphroditic species chromosomal distinctions are ordinarily absent. The higher forms with sex and chromosome differences, on the other hand, may show reversion to the hermaphroditic condition from the dioecious or bisexual states.
Other less common cytogenetic controls of sex development have become recognized and better understood. Discussion of their gene and chromosomal arrangements will be considered when these cases arise. The al)ove types will be sufficient to furnish a basis for interpreting the newer data.
C. Changing Methods Of Cytogenetics
The earlier studies of sex determination depended on the natural arrangements of chromosomes found in different species and on occasional chromosomal rearrangements occurring as relatively rare aberrant types. Further development of genetics has increased the tools for these studies. Mutant genes have been shown to control chromosome pairing in segregation (Gowen and Gowen, 1922; Gowen, 1928; Beadle, 1930).
Different drugs such as chloralhydrate and particularly colchicine and various forms of radiant ^energy have made it possible to create new types by doubling the chromosomes, by chromosome rearrangement and by changing the chromosome number. Better genetic tester stocks of known composition have been organized, which together with a more exact understanding of the inheritance structure of the species have made for more critical studies. Techniques, which have improved chromosome differentiation and structural analysis through better methods, better dyes, tracers to mark chromosome behavior (Taylor, 1957), and the use of hypotonic solutions in the study of cells (Hsu, 1952), have removed doubts that were created by the earlier technical difficulties. Instrumentation has improved for the measurement, physically and chemically, of cell components.
D. Chromosomal Association with Sex
The first genetic linkage groups were associated with the sex chromosomes and were soon shown to follow the patterns of the different chromosome sets observed within the different species. In man, hemophilia inheritance was observed to follow that which was presumed for the X chromosome. Barring in birds and wing-color pattern in Lepidoptera on the other hand were found to follow the chromosomal patterns of their species where the male was ZZ and the female Z or ZW for the sex chromosomes. The utility of these methods was further probed by Bridges' observations that when chromosome behavior in Drosophila resulted in sperm or eggs carrying uncxjx'cted chroiiiosomc combinations there followed ('(nmlly unexpected phcnotypes in the progeny. The cliaractei'istics of these unexpected progeny, in turn, I'ollowt'd those expected if genes for them were carried m the sex chromosomes. The use of these linkage groups as tracers, both as they naturally occur and as they may be reorganized through the treatment effects of such agents as radiant energy, open the possibility ol assigning sex effects to, not only chromosomes, but also to particular i-laces within the chromosomes.
Both polyploids and aneuploids, as occurring naturally and as marked by tracer genes, give insight into sex differentiation and its dependence on chromosome inheritance behavior. True polyploids in a species are formed as a consequence of multiplying the entire genome. The possible types may have a single set of chromosomes and genes in their nuclei (haploid ) , 2 sets (diploid) , 3 (triploid), 4 (tetraploid), and so on, representing the genomes 1. 2. 3. 4. to whatever level is compatible with life. Multiplying the genomes within the nucleus often increases cell size but seldom gives the organism overtly different sex characteristics.i Aneuploids, on the other hand, give quite different results, the result being dependent upon the particular chromosome that may be multiplied. Particular trisomies in maize, wheat, and spinach, for example, are distinguishable by marked differences in phcnotypic appearance that is not attributable to cell size but rather to abrupt deviations in particular characteristics. The genes in the particular trisomic set are unbalanced against those of the rest of the diploid sets within the organism. Their phenotypes express these differences.
E. Balance of Male and Female Determining Elements in Sex Determination
The foundations for the basic theory that sex determination rests on quantitative relationships of two genes or sets of genes localized in separate chromosomes rests largely on the work of Morgan, Bridges and Sturtevant, 1910, with Drosophila and the breeding work of Goldschmidt, 1911, with Lymantria. The work of Goldschmidt soon gave extensive descriptions of diploid intersexuality in L]imantria dispar. The details of this work scarcely net'd review because they have \)vvn repealed several times and have been smnmarized recently in Goldschnu(h's Theoretical Genetics (1955). Fioui (hildschmidt's viewpoint, the basic point was that definite conditions between the sexes, that is, interscxuality, could be pro.lnced at will l)y proper genetic combina, tions (crosses of subspecies of L. dispar)
• The notable exception of the Hynienoptera will I )c discussed later.
without any change in the mechanism of the sex chromosomes, and this in a typical quantitative series from the female through all intergrades to the male, and from the male through all intergrades to the female, with sex reversal in both directions at the end point. The consequence was: (1) the old assumption that each sex contains the potentiality of the other sex was proved to be the result of the presence of both kinds of genetic sex determiners in either sex; (2j the existence of a quantitative relation, later termed 'balance' (though it is actually an imbalance), between the two types of sex determiners decides sexuality, that is femaleness, maleness, or any grade of intersexuality; (3) one of the two types of sex determiners (male ones in female heterogamety, female ones in male heterogamety) is located within the X-chromosomes, the other one, outside of them; (4) as a consequence of this, the same determiners of one sex are faced by either one or two portions of those of the other sex in the X-chromosomes; (5) the balance system works so that two doses in the X-chromosomes are epistatic to the determiners outside the X, but one dose is hypostatic; (6j intermediate dosage (or potency) conditions in favor of one or the other of the two sets of determiners result, according to their amount, in females, males, intersexes, or sex-reversal individuals in either direction; (7) the action of these determiners in the two sexes can be understood in terms of the kinetics of the reactions controlled by the sex determiners, namely, by the attainment of a threshold of final determination by one or the other chain of reaction in early development; while in intersexuality the primary determination, owing to the 1X-2X mechanism, is overtaken sooner or later — meaning in higher or lower intersexuality — by the opposite one, so that sexual determination finishes with the other sex after this turning point. The last point is, of course, a problem of genie action."
Bridges developed his idea of "genie balance" as a consequence of his observations on chromosomal nondisjunction, particularly as it illustrated the loss or gain of a fourth chromosome in modifving nonsexual characters. The similarities and contrasts of this view from that of Goldschmidt are indicated by the following quotation (Bridges, 1932) : "From the cytological relations seen in the normal sexes, in the intersexes, and in the supersexes, it is plain that these forms are based upon a quantitative relation between qualitatively different agents — the chromosomes. However, the chromosomes presumably act only by virtue of the fact that each is a definite collection of genes which are themselves specifically and qualitatively different from one another. There are two slightly different ways of formulating this relation, one of which, followed by Goldschmidt, places primary emphasis upon the quantitative aspect of individual genes. The other view, followed by the Drosophila workers, emphasizes the cooperation of all genes which are themselves qualitatively different from one another and which act together in a quantitative relation or ratio. Goldschmidt developed his idea through work with the sex relations in Lymantria and has sought to extend it to ordinary characters. The other formulation, known as 'genie balance,' was developed from the ordinary genetic relations found in characters. Both are crystallizations of fundamental ideas with which the earlier literature was fairly saturated and no great claim to distinctive originality should be ventured for either or denied for one only. Both are physiological as well as genetic — that is, they are formulations of the action of genes, not merely statements of the genie constitutions of individuals nor merely studies of the way genes act. The physiological side has been emphasized by Goldschmidt and the genetic side by Drosophila workers. But Goldschmidt's tendency to represent the view of genie balance as without, or even as in opposition to, such physiological formulation is groundless — as groundless as would be the reciprocal contention that Goldschmidt's theory is only one of 'phenogenetics.'
"A common element in the foundation of both formulations is that if a gene is represented more than once in a genotype the phenotypic effect is expected to be different, though roughly in the same direction as before and roughly proportional to the quantitative change in the genie constitution."
Since the sexually different types observed by Bridges were accompanied by whole chromosomal differences, he could point to losses or gains of autosomes, with an internal preponderance of genes tending to develop male organs, as balanced by genes of the X chromosomes tending in the direction of producing female organs or of suppressing alternative male organs. The net effect of the X chromosome favoring femaleness, and of the set of autosomes favoring maleness, terminates in the development of male or female, according to the ratio of these determiners in the whole genotype. The effect of the X chromosome goes on the basis of whole numbers 1, 2, 3, 4, as does the similar variation in the sets of autosomes.
TABLE LI
Chromosomal numbers and kinds for the different
recognized sex types of Drosophila
Type
Superfemale . . . Triploid meta female
Female*
Female
Female
Female
Female
Female
Female
P'emale
Female
Female
Intersex*
Intersex
Intersex
Male
Male
Male
Male
Male*
Supermale
Chromosomes
X
Y
A
3
2
4
3
4
4
3
3
3
1
3
3
2
3
2
2
2
1
2
2
2 + vS
2
2
2 + yL
2
2
2
2
1
1
3
4
2
3
2
1
3
2
1
2
2
2
3
2
4
3
X/A Balance
1.5
L3
LO
LO
LO
LO
LO
1.0
LO
LO
LO
LO .75 . ()7 .()7 .50 .50 .50 .50 .50 .33
These forms are cited by Bridges from his own observations, from L. V. Morgan (1925) and from Sturtevant (unpublished). As yet but limited studies of these forms, which must be rare, have been published. Fourth chromosomes generally, but not always, equal mimber of the other individual chromosome gn)U])s.
Goldschmidt, since he was dealing with males and females of the diploid type, took a corresponding view for the Z chromosomes of his moths with this difference. Since the Drosophila chromosome pattern is XY + 2A for the males as contrasted to 2X + 2A for the females and the pattern for Lymantria is ZZ -|- 2A for the males and ZW + 2A for the females, it was necessary to use a relation which was reciprocal to that of Drosophila; the male determining element or elements were assigned to the Z chromosomes. Lymantria has a rather large number of different races found in different geographic locations. Within any one of these races this formulation apparently sufficed. However, from crosses between races it was soon observed that the progeny showed ranges in sexuality all the way from phenotypic males to phenotypic females although these females were actually genetic males. To Goldschmidt, this variation indicated different potencies of the male-determining element. Similar differences were attributed to the female element which he had first assigned to the cytoplasm but for which he later favored a W chromosome location.
In applying this postulate of discrete chromosome contributions to sex according to their number, Bridges made the further assumption that female-producing genes l)redominate in the X and are scattered through it in more or less random fashion as are the genes affecting so-called somatic characteristics as wing shape or bristle pattern. The quantitative relations for the different chromosomal types, together with their descriptions, are indicated in Table 1.1.
In the formulation of Table 1.1, the X and Y chromosomes are counted separately, whereas a set of A chromosomes (autosomes), is allowed a value of but one even though comjiosed of a 2nd, a 3rd, and a 4th chromosome for the haploid genome. The dii)loid set of autosomes is given a weight of two, and so on. From these data Bridges observed that the presence or absence of a Y chromosome did not affect the sex types. The X chromosomes and autosomes were, however, important. He held that their importance stood as the ratio of thcii' prcsuiiK'd iM'oducts to each other. The ratio is 1 for the perfect female and 0.5 for the perfect male. Between these values intersexual conditions develop. Beyond the value 1, development is overbalanced byexcess female genes resulting in the superfemale. Values less than 0.5 create a deficiency in the female elements or excess of male elements and a supermale results. Schrader and Sturtevant (1923) proposed another system. Instead of the ratio of X clu-omosomes to autosomes, they suggested that a straight difference between the products of the female determining elements of the sex chromosomes and the male effects of the autosomes causes the sex changes. ]3ridges criticized this system on the basis of the fact that progressive polyploidy did not change the sex type or ratio between the X and autosomes, whereas the numerical difference between them would be progressively increased. While keeping the X/A ratio as descriptive of the ultimate effects of the genes in these chromosomes, he modified their proposed weight from 1 : 1 for X:A to 1 for the X and 0.80 for the A.
All formulations for explaining sexual differences are beset with a lack of an unbiased quantitative scale by which these differences can be measured. The estimates of the changes in sexuality are left to the insight of the observer. It seems reasonable to suppose that the quantitative relations between the male and female sex determining elements should have intermediate values when the specimens under observation show a mixture of organs of either sex. This agrees with Bridges' considerations of this problem. It is not so clear, however, that the so-called supersexes^ really are what the names may connote to many readers. The superfemales, with their three X chromosomes and two sets of autosomes, are quite inviable; small in size, wings reduced and irregularly cut on the margins, ovaries developed to only the early pupal stage, and reproductive tracts much reduced in size.^ The supermales with one X
- Recently termed metafemales bj' Stern (1959b).
'^ Further development of the ovaries is able to take place in normal XX + 2A hosts. Larval XXX + 2A ovaries transplanted into fes/fes hosts IM-oduce eggs in the recipient host which on fertilization are capable of developing into adult imagoes. These imagoes show that there is a low per
chromosomc and three sets of autosomes are described as resembling males but are sterile. The wings arc somewhat spread and bristles less in size. They are late emerging and poorly viable. Neither type can be referred to as superior to normal female or normal male in anatomic develoi)ment or physiologic functioning. A new type, recently described by Frost (1960j, emphasizes this difficulty. Females, called triploid metafemales, Table 1.1 had 4X chromosomes and 3 second, 3 third, and 2 fourth autosomes. Viability was greater than superfemales but still low. Fertility was about 10 per cent. Progeny per female about 10. The flies were like triploids in bristles, eye and wing cells large, sex combs absent. They showed characteristics of superfemales in rough eyes, narrow wings without inner margins, and smaller body build. The distribution of these different Drosophila sex types (Table 1.1) showed that optimal development comes when the X/A values are 1.0 and 0.5. Any deviation away from these values tends to make the sex system less rather than more efficient.
In normal Drosophila sex differences are probably expressed in every cell making up their bodies. These differences are made visible to us only under special conditions. In adult organ differentiation, the sexual differences are manifest through such things as the body size of the males being about three-fourths that of the females, differences in coloration of the tergites, the appearance of sex combs, the development of the gonads into ovaries and testes, and the formation of a secondary reproductive system composed of several glands and ducts. The origin of these last elements is of particular interest. The ovaries can be distinguished from the testes as early as the second instar through their size and position within the fat body. They are located about two-thirds the larval length back from the mouth parts. The sex combs, on the other hand, take their origin from imaginal discs which are located in the head region, possibly one-third back from the mouth parts. The secondary reproduc centage of crossing over and high nondisjunction rate in the XXX + 2A ovaries and a high mortality rate in the offspring (Beadle and Ephrussi, 1937).
tive system for both the males and females takes its origin from a disc at the posterior extremity of the larvae. The testes or ovaries are only brought together with the secondary reproductive tracts during late development in the pupae, whereas the sex combs retain their separate development. The striking action of chromosome balance in sex determination is that all of these organs and others are jointly affected and simultaneously develop directly into either the male or female sexual types. Of these characteristics, the sex combs are particularly trustworthy indices of maleness. In the male, these combs are heavily sclerotized and pigmented. They have 9 to 13 rather blunt teeth on their margins. The normal females lack these sex combs entirely. The intersexes have well developed combs, whereas the triploid females and superfemales lack them. This all-or-none situation resembles that observed in the rest of the block of sexually differentiating characteristics found in normal males or females, save that in the intersexes development may result in the organs of either sex appearing in the primary or secondary reproductive systems. The all-or-none character of the sex combs may, however, be bridged in that the appearance of certain mutations leads to the production of these combs in all of the different sexual types. The combs differ in size, in thickness and length of the teeth, and their number. The sex combs meet the conditions of a quantitative character which may be counted or measured in unbiased units. Thus in the case of sex combs, it is possible to obtain quantitative information on the effects of clu'omosomal changes on the expression of this form of sexuality.
TABLE L2
Mean sex conih teeth for flies having (lijj'erent X and
A chromosome
numbers and heteroziiqons for
the Hr gene
Sex Type
Chromosome
Mean Sex
Genotype
Comb Teeth
Males
X + 2A
11.4
Intersex
XX + 3A
9.1
Female
XX + 2A
().9
Superfemale ....
XXX + 2A
5.0
Triploid female.
XXX + 3A
4.8
Two gene mutations in D. melanogaster have aided in this search. The first is the dominant gene, Hr, which causes the male reproductive tract to be added to that of the female when this gene is heterozygous (Gowen, 1942 j . The extensive effects of this gene on the whole sex determining system have been described by Gowen (1942, 1947) and Fung and Gowen (1957a). The second gene is that of Sturtevant's (1945) transformer, tra, which operates on the female phenotype to convert it to that which corresponds to the male in having sex combs, full male reproductive system including testes, while still leaving the female characteristic body size. These genes are located in the third chromosome of D. melanogaster. Crosses between them show allelomorphic effects. Combinations of these genes in conjunction with different chromosomal arrangements make possible a series of different sex types which are distinguishable from ordinary males and females (Gowen and Fung, 1957). As some of these types bear sex combs, a quantitative character is furnished in the variation of the sex comb teeth which may be used as an impersonal measure of departure of these types from ordinary males or females. The groups having particular interest are those carrying the Hr gene in heterozygous condition with the X and A chromosomes having various numbers. The mean numbers of sex comb teeth for these various groups are shown in Table 1.2.
Analysis of these data for the contributions made by the sex chromosomes and autosomes to the numbers of teeth found on the sex combs of these different genotypes shows the following relation.
Sex combs == 12.82 - 3.42 X + 0.89 A
From this equation it is seen that the X chromosome has four times as much effect on lowering the number of teeth on the sex combs as a set of autosomes. The direction of effect is, as would be expected, increasing the number of X chromosomes tends toward making the individual more female-like in that the sex combs become smaller and less l)ronounced. Increasing the number of autosome sets on the other hand tends to push the indiviihial toward the male type with larger sex combs. Some 95 per cent of the variation in sex comb teeth has been accounted for by this equation.
The above equation results when the effect of the sex chromosomes and autosomes is considered as operating on a simple additive basis. It is interesting to consider these effects on the basis of the ratio of sex chromosomes to autosomes as utilized by Bridges. As is customary, the male genotype is given a weight of 0.50, the intersex 0.67, the female 1.00, the superfemale 1.50, and the triploid female 1.00. With these values the data on the sex combs are fitted by the equation
Sex combs = 13.40 - 6.38 X/A
The fit of this equation to these data shows control of less of the variation in the sex comb teeth. Only 76 per cent of the variation is accounted for by these methods whereas 95 per cent is accounted for when the effects of the X and A chromosomes are considered as additive.
If it is agreed that the condition of the sex combs is a good unbiased measure of the degree of sexuality of the Drosophila, it follows that it would be more probable that the genes in the X chromosomes operate additively with those of the autosomes.
III. Sex Genes in Drosophila
A. MUTANT TYPES
Bridges' concept of sex determination turned on the action of sex genes located more or less fortuitously throughout the inheritance complex of the species. In Drosophila it happens that the major female determining genes seem to be located in the X chromosome and the male determining genes in the autosomes, whereas the Y chromosome seems essentially empty of sex genes. In support of this concept limited data are cited on specific genes affecting the reproductive system or its secondarily differentiated elements and two cases where genes affected the primary reproductive system as a whole. During the interim between 1938 and the present, the numbers of these genes and the breadth of their known effects have been notably increased. Again the genes as a whole affect every phase of sexuality, morphology, fertility,
and physiology. Single genes may occasionally alter both sexes or may frequently affect only male or only female phenotypes. Single genes may appear to influence two or more distinct characteristics observable in the developing flies, although this multiple phenotypic expression may go back to a gene action which is controlling a single event in development. Genes affecting the structural development of either male or female organs frequently are accompanied by sterility of various degrees. A very large category of genes is known only through its effects on sterility of either or both sexes. Experience has shown that when properly analyzed anatomic changes are probably basic to the sterility. In this sense genes for sterility should be considered genes for sex characters. Berg (1937) furnished data on the relative frequency of sterility mutations in the X chromosome as against those in the autosomes. 12.3 per cent mutations in which the males were sterile were found in the X chromosome against 4.5 per cent found in the second chromosome. These results show that the X chromosome has many gene loci occupied by genes capable of mutating to sterility genes which affect males. Sterility is also common for the females but requires more testing. The loci for these genes are widely distributed both within and among the chromosomes.
Genes affecting sex morphology are found in all Drosophila chromosomes. Of 17 which have been recently studied; 6 were in the 1st chromosome, 5 in the 2nd, 5 in the 3rd, and 1 in the 4th. Insofar as can be determined these genes are no different than those affecting other morphologic traits. They may be dominant, they may be recessive, and a limited number of them may show partial dominance. They affect a variety of sex characteristics and do not always involve sterility of one or the other sex. The loci occupied by these sex genes may have several alleles, some of which may lead to sterility, others not. Most affect characters like size and development of the ovaries, the characteristics of the eggs, duct development, spermathecae, ventral receptacles, parovaria, paragonia, sex combs, position of genitalia, and so on.
Altlioiigh Drosophila is the leader in furnishing types for analyses of the inheritance basis for sexuality, it is well known that similar conditions exist in other forms of life.
B. MAJOR SEX GENES
Major sex genes affecting the dichotomy of the sexes have appeared among the observed mutational types. Sturtevant (1920a, 1921 j in Drosophila simulans isolated a gene in its second chromosome which when homozygous could convert diploid females into intersexuals and render XY males sterile. Phenotypically these intersexuals were female-like in that they lacked sex combs and had 7 dorsal abdominal tergites, ovipositor of abnormal form, 2 spermathecae, and lacked the penis. They were male-like in having first genital tergite although abnormal in form, lateral anal plates, claspers, black pigmented tip to the abdomen. The gonads were rudimentary. The gene was recessive and, as expected, showed no effect on D. simulans X D. melanogaster hybrids.
In 1934 a new intersexual type was observed by Lebedeff in D. virilis. Intensive study of this type showed that it depended on a fairly complex inheritance. A 3rd chromosome recessive gene ix™ at 101.5 converted the XX females into sterile males, showing only one noticeable female characteristic, the presence of a rudimentary 5th stcrnite. Sterility could be accounted for, even though the internal genitalia were completely male, by the small size of the testes and the degeneration of the germ cells largely at the spermatocyte stage. Observable effects of this gene appeared only in females, XX + 2A. Genetic modifiers were isolated which acted on the ix"yix'" complex. Other gcnotyi)es were derived through recombinations of these genes. The resulting phenotypes showed a greater variation of the male-female mosaics. One of these types was sterile but fully hermaphroditic having a complete set of external and internal genitalia of both male and female. Genetically, this type was homozygous for the ix'" gene but in addition luid both a previously found dominant scmisu})pressor and also a second semisuppressor. Another line having the ix'" gene homozy
gous had a full set of male organs but there
were rudimentary ovaries attached to the
testes. This line showed the dominant semisuppressor as the restraining element on the
developmental pattern of the ix'" gene. Another type was separable in that it was still
more female-like, yet had male external
genitalia and rudimentary testes. This type
was the result of the homozygous ix'" genes
operating in conjunction with 3 different
semisuppressors. Extensive embryologic
studies were interpreted as indicating that
gonads, ducts, and genitalia had started as
in females with the XX constitution.
Shortly thereafter male organs appeared as
new outgrowths from the same imaginal
disc. The development of the two sets of
organs was then simultaneous but still depended on the gene pools present in the
particular strain.
Another intersexual type appeared in D. pseudoobscura as a mutation, presumed due to a single dominant gene (Dobzhansky and Spassky, 1941). In a series of cultures having "sex ratio," two cultures gave 234 females, 7 males, and 266 intersexes. The females' progeny transmitted only the normal condition to their Fs progeny. The males were sterile presumably because they lacked a Y chromosome. The intersexes were also sterile so that further study of the genetic condition became impossible. The evidence, however, is interpreted as showing that the intersexes were transformed females which had inherited a dominant gene governing this condition. The intersexes were characterized by two sets of more or less complete genital ducts and external genitalia but only one pair of gonads. One set of ducts and genitalia was almost always more female-like and the other more male-like. Sex combs were present, the distal comb had 2 to 4, and the proximal 4 to 6 teeth, compared with the normal male number of 4 to 7 on the distal and 6 to 9 on the proximal comb. Body develo{)ment was more like that of the female. Cytologic examination showed two of the intersexes had (wo ()\-aries each. One of these two had a rudimentary testis. The chroinosoiiu' complement was 2X + 2A.
A dominant gene which causes intersexiiality in diploid females of D. virilis was estal")lished by Briles. Stone (1942) located this gene in the second chromosome. Price ( 1949j placed the gene within the second chromosome near the locus of "brick" by inducing crossing over in males by exposing them to x-rays. Newby (1942) extensively studied the embryologic sequence in development of the organs of these intersexual types. The Ix^ gene did not affect the males, XY 4- 2A, but did change the females XX + 2A into intersexes when it was in the heterozygous condition. The intersexuals had 9 tergites; the first 6 were like those in the normal female and the last 3 were small and irregularly formed. There were 6 sternites, the first 5 being normal, but the 6th malformed. Anal valves were lateral as in the male but a third small valve was also present at the ventral side of the anus. The plates forming the claspers were of irregular pattern and found ventral to the anus. The vaginal plates were often extruded into a genital knob and were below the claspers. The knob occasionally became heavily pigmented. The internal organs ranged from nearly female through those which were of hermaphroditic type containing representative organs of both sexes to individuals almost wholly male. Newby concluded that intersexuality expresses itself as a response to the developmental pressures of both sexes, not as development in the one direction followed by a change.
Gowen in 1940 established a stock carrying the dominant gene Hr which had apl)eared as a mutant in one of his cultures of D. melanogaster. This gene affected diploid females of XX + 2A type but not the males of XY -I- 2A constitution. In the presence of the Hr gene the diploid phenotype of the females changed into a sterile type with male secondary reproductive system associated with the female counterpart. The first 6 segments were complete with 6 spiracles. The 7th was small with spiracle. The 8th was small but without spiracle. Sternite forming rudimentary ovipositor was usually protruded. Ninth and 10th segments resembled those of males with large tergal plates. Claspers were abnormal and had a pair of small plates flanking the anus majoi-ly in vertical position. Organs formed, although sometimes modified or missing, included: sex combs of 6.9 long slender teeth, gonads distinguishable from those of the ordinary male or female in the 3rd and possibly the 2nd instar, genital ducts male and/or female, male accessory gland, penis deformed, sperm pump, vas deferens, spermatheca, ventral receptacle often displaced, and occasionally parovaria. The primary gonads were often abnormal ovaries but in rarer instances bore a crude resemblance to testicular tissue. The yellow of the testes was frequently present as material clinging to the ovary.
Superfemales with one dose of the Hr gene had sex combs and developed parts of both the male and female external and internal reproductive systems. Sex combs had an average of 5 long and slender teeth. Abdominal segment 8 developed as in the female, and formed the vaginal plate. The latter was abnormal in shape, ordinarily becoming a sclerotic protuberance. Segments 9 and 10 developed more as in the male but were incomplete and abnormal. The genital arch did not develop but the inner lobe of tergite 9 showed irregular and abnormal growth as for the claspers. Segment 10 developed, as in the males, into longitudinal plates flanking the anus. The internal genitalia were underdeveloped but consisted of mixtures of male and female organs. Gonads were rudimentary but generally consisted of a pair of ovaries with small traces of yellow pigmentation.
The triploid fly with one dose of the Hr gene was largely female with developed ducts, ovaries and eggs, but was sterile. The male characteristics were small sex combs and dark abdominal plates. In superfemales, sex combs were present and teeth were intermediate between those of the diploid female and triploid female. The gene showed a dosage effect in triploids which was less than that observed in diploids and was in relation to the relative balance of the gene with its normal alleles, 1:2 for the triploid and 1 : 1 for the diploid. The developmental effects of Hr as well as the pigment producing potentialities of testes, ovaries and hermaphroditic gonads have been discussed by Fung and Gowen (1957a, b).
Hr has been shown to be allelomorphic to a recessive gene, tra, described by Sturtevant (1945) and known to be located in the 3rd chromosome. The location of this allele, tra, is at 44 to 45 or between the genes scarlet and clipped. When homozygous the gene transformed diploid females into sterile males. Heterozygotes showed no detectable differences from normal females of XX constitution. Males XY homozygous for tra or heterozygous for it were indistinguishable from normal males. The homozygotes XX, tra/tra were female in body size, but otherwise were nearly male in appearance. They had fully developed sex combs, male colored abdomens, normal male abdominal tergites, anal plates, external genitalia, genital ducts, sperm pumps, paragonia, and showed the usual rotation of the genital and anal segments through 360 degrees. They mated with females readily and normally. The testes, however, although normal in color, elongated, curved, and attached to the ducts were of small size. Testis size was never that found in normal brothers. The addition of a single Y or two Y's did not alter fertility.
The triploid females 3X + 3A homozygous for tra, had large bodies, ommatidia and wing cells. They resembled the diploid homozygous tra individuals in having male external genitalia, well developed ejaculatory ducts, sperm pumps, and accessory glands, testes elongated but narrower than those of normal males, and sex combs averaging about 9.6 teeth. They mated with females but were completely sterile.
Triploids with one or two doses of tra were like wild type triploids in having no sex combs and being female throughout. Intersexes having one or two doses of tra were similar to intersexes having only wild type genes in the locus.
Sturtevant obtained one superfemale which was homozygous for tra. It had male genitalia and sex combs with only about half the normal number of teeth. This individual argues for a greater balance toward the female side of sexual development than either the diploid or triploid females previously discussed. The evidence is, however, contradictory to that furnished by the Hr gene as indicated earlier.
A combination of two or more genes, Beaded and various Minutes, having well known phcnotypic effects, has sometimes
produced phenotypes which have been interpreted as peculiar, low grade types of intersexuality in males (Goldschmidt, 1948, 1949 and 1951). The data showed that the Beaded cytoplasm favors the low grade intersexual male whereas the Minute cytoplasm favors the reduced male with the hetcrozygote being intermediate. Just how far these types may be related to the other types strongly affected by specific genes is a matter of question, having at least other interpretations (Sturtevant, 1949).
In 1950, Milani trapped an inseminated female of D. subobscura w^iich segregated intersexual progenies. Spurway and Haldane (1954) studied these intersexual types. A recessive guiding development toward these intersexes was located on the 5th chromosome of subobscura. When present it caused the XX homozygous females, ix/ix + 2A, to have sex combs on both the first and second tarsal joints. The numbers of teeth making up the sex combs w^ere reduced as also were the sizes of the teeth. The illustration in Spurway and Haldane's (1954) paper indicates that the number of teeth was 7 on the first tarsal joint and 5 on the second joint, whereas the sex combs of the males had 11 teeth on the first joint and 9 on the second. A series of changes were observed in the genital plates which graded from those resembling true females to those approaching the male type.
C. OTHER CHROMOSOME GROUP ASSOCIATIONS IN DROSOPHILA AMERICANA
I), aniericana has 4 chromosomes in the female genome and 5 chromosomes in the male genome. As compared with D. virilis the X chromosome is fused with the 4th chromosome and the 2nd chromosome is fused with the 3rd, the 5th and 6th chromosomes are free in the female, whereas in the male genome the Y chromosome, 4th, 5th and 6th chromosomes are free and the 2nd and 3rd fused. Stalker (1942) has shown that the three female genomes are balanced and lead to triploid females as they do in D. melanogaster. D. aniericana triploids differ from their diploid sisters in having bigger ommatidia, larger wing cells and somewhat larger bodies. When these triploids are bred to diploid males they give rise to 6 chromosomally different types of offspring: diploid males, diploid females, triploid females, intersexes, females carrying a Y and a male limited 4th chromosome hut otherwise diploid, and tetraploid females. All intersexes cytologically show a Y and a 4th chromosome present. Intersexes without the Y are presumed also w^ithout male limited 4th chromosomes and would be expected to be inviable or very weak. No supermales or superfemales, that w^ould correspond with those found in D. melanogaster, were observed so are presumed to l)e inviable due to unbalance for the 4th chromosomes. Among 948 progeny of triploid females X diploid males there were 9 individuals that were phenotypically abnormal females. They had slightly spread, ventrally curved wings with slightly enlarged wing cells. In 8 of the 9 the first section of the costal vein was shortened so that no junction was made with the first vein at the distal costal break. Heads were large with rough eyes, thoraxes shortened, legs fre(luently malformed, and abdomens small with unusually wide 7th sternites. Genitalia were apparently normal with well developed ovaries. This type carries three doses of any genes contained in the 4th chromosome to two doses of the genes in the other chromosomes. Its phenotype represents a trisomic condition.
The sex characteristics of the flies observed in D. americana seem to follow the same patterns as those of D. melanogaster as judged by the numbers of the X chromosomes and autosomes. A Y chromosome in the intersex was not observed to affect sex expression. The intersexes could be grouped into six classes ranging from extreme male type to the most female type. The male type showed largely male organs, courted females, and had motile but nonfunctional sperm. The most female type had nearly normal ovipositor plates, well developed uterus, ventral receptacle, spermathecae and oviducts. At least one gonad showed egg strings, although a small patch of orange-red tissue was present at the tip. Two types of chromosomes were observed in the nuclei of these extreme female type intersexes; those like the chromosomes in any normal diploid cells and some which were so swollen as to be almost unrecognizable. Such swollen chromosomes were not found in the other classes of intersexes or in diploid or triploid individuals. They are suggestive of some noted by Metz (1959) in Sciara. Most of the intersexes were of the male type, 45 per cent, with decreasing numbers for each of the other five classes until those in the most female class constituted only about 4 per cent of the total.
D. LOCATION OF SEX-DETERMINING GENES
The problems of isolating and determining the modes of action of the factors normally operating in sex determination have received extensive study since they were reviewed by Bridges in 1939: Patterson, Stone and Bedichek (19371, Patterson (1938), Burdette (1940), Pipkin (19401942, 1947, 1959), Poulson (1940), Stone (1942), Dobzhansky and Holz (1943), Crow (1946), and Goldschmidt (1955). From his work on Lymantria dispar, Goldschmidt concluded that sexual differentiation was controlled by a major male factor, M in the Z chromosome and a factor F directing development toward the female and at first assumed to be in the cytoplasm but later considered to be in the W chromosome. The heterogametic female of this species would then be FM and the male be MM. In considering this problem, Goldschmidt attempted to distinguish between the sex determiners responsible for the F/M balance and modifiers affecting special developmental processes (Goldschmidt, 1955). At the other extreme Bridges' study of triploids led him to consider that sex in Drosophila was determined by the interaction of a number of female tendency genes found largely in the X chromosome and of genes having male bias located largely in the autosomes. These numerous genes were considered as being distributed throughout the whole inheritance complex. Search for the more exact locations of these genes within the different chromosomes of Drosophila has largely taken the form of determining the variation in sex types as induced by the addition or deletion of various pieces of the different chromosomes to either the normal male, normal female, or triploid complexes.
The sections of the chromosomes added
were derived from previous translocations
generally to the 4th chromosome and were
of varying lengths determined through cytologic and genetic study. Summaries of
these comparisons are found particularly in
the papers of Pipkin (1940, 1947, 1959). In
their search for a major female sex factor
in the X chromosome, Patterson, Stone and
Bedichek (1937), Patterson (1938), Pipkin
(1940), and Crow (1946) finally were unable to show that any single female sex determiner, located in the sex chromosome,
was of primary importance to sex. Evidence
for multiplicity of genes with a bias toward
female determination was found by Dobzhansky and Schultz (1934) and by Pipkin
(1940). They were able to transform diploid intersexes into weakly functioning hypotriploid females by the addition of long
fragments of the X chromosome to the
2X + 3A intersex complement. Short sections of the X chromosome in some cases
shifted the sex type in the female direction.
Pipkin found that additions of short X
chromosome sections, to the 2X -|- 3A chromosome sets, although covering in succession the entire X chromosome, were insufficient to make the flies other than of the
intersexual type. Longer and longer fragments from either the left or right end of
the X chromosome caused a qualitatively
progressive shift toward femaleness.
Weakly functional hypotriploid females resulted when either right- or left-hand sections of two translocations with t-lz (17)
and Iz-v (W13) breaks were present in the
2X + 3A chromosome complement. These
duplication intersexes possessed 1 or 2 sex
comb teeth when reared at 22 °C. and up to
5 well developed teeth when reared at 18°C.
These facts give support to the multiple
sex gene theory of Bridges or at least that
quantitative differences in sex potencies
exist within the X chromosome. This conclusion was further strengthened by the hypointersexes lacking a short portion of one
of their two X chromosomes although possessing three of each autosome, inasmuch
as these types were shifted strongly in the
male direction. These studies of the X cliromosonie show that several parts of this
chi-omosomc are concerned witli female dif
ferentiation and that the effects are irregularly additive.
Similar search of the autosome II and III for genes of male potency showed that small shifts in the male direction were found in hyperintersexes for several short regions of chromosome III but for none of chromosome II (Pipkin, 1959, 1960). Tl^ree slightly different right-hand end regions of chromosome III produced the largest shifts in the male direction in hyperintersexes, but no increase in number of sex comb teeth. These changes were comparable with those produced in the female direction by the addition of very short sections of the X-chromosome to the 2X + 3A intersex complement. On the other hand, none of the seven different hypointersexes lacking a short section of the 3rd chromosome from the 2X + 3A complement showed a shift in the female direction. This is rather surprising as hypointersexes for two short regions in the X chromosome were shown by Pipkin (1940) to shift the sex type in the male direction as was to be expected. From these results Pipkin (1959) derives the conclusion that 3rd chromosome aneuploids as well as those of the 2nd chromosome and X chromosome support the deduction that dosage changes of portions of the X chromosome are more powerful than dosage changes of portions of either of the large autosomes in affecting sex balance. This view receives further support through changes of size and number of sex comb teeth as observed in the chromosomal types carrying the gene Hr and reviewed earlier.
Influence of the Y chromosome on sexual differentiation has generally been ruled out as XO nondisjunctional flies are male although they are sterile (Bridges. 1922). Similarly Dobzhansky and Schultz (1934) ruled out the Y chromosome as an effective influence on sex types of triploid intersexes of D. melanogaster since the mean sex type of Y -(- 2X 4- 3A intersexes did not differ significantly from the sex tyyies of siblings 2X + 3A. "
The steps taken in the studies of chromosome IV are of interest. Dobzhansky and Bridges in 1928 concluded that the 4th chromosomes play no part in sex determination in D. melanogaster. The evidence was of two kinds. Triploid females were outcrossed separately to males of two diploid stocks. The triploid daughters from these crosses were again crossed to males like their fathers. This repeated outcrossing to the different stocks resulted in a shift in the grade of the intersexes in both cases, in one case to a very high proportion of extreme male-like intersexes and in the other to nearly as high a proportion of extreme female-type intersexes. These results were interpreted as showing that the grade of development of the sexual characters was dependent on genetic modifiers. The second experimental test consisted of subjecting a triploid stock to selection toward a line which produced a high proportion of extreme male type intersexes and to another line which would have a high proportion of extreme female type intersexes, each being much higher than the original stock. The l)rocedure established a line with a high proportion of extreme male type and another line which was not so extreme in its proportions but was definitely higher in female type intersexes than the original stock. Again these results w'ere interpreted as indicating the selection of modifying genes of unknown positions within the inheritance complexes.
This evidence had been preceded by Bridges' (1921) discussion in which he wrote "the fourth-chromosome seems to have a disproportionately large share of the total male-producing genes; for there are indications that triplo-fourth intersexes are predominately of the 'male-type', while the dijjlo-fourth intersexes are mainly 'femaletype'." In 1932, Bridges concluded for Drosophila intersexes that, in spite of the favorable genetic checks, in repeated and varied tests, it has been impossible to state with any assurance whether the 4th chromosome is or is not a large factor in the variability encountered.
In our own work a stock of attached X triploids has for many years consistently produced only male type intersexes. This is in contrast to what we frequently see within other lines of triploids as made up utilizing the cIIIG gene (Gowen and Gowen, 1922). Lines established from these triploids ordinarily have three intersexual types: male,
intermediate, and female. These lines, however, may be subjected to selection in both directions. In our experience, male intersex lines are established rapidly and remain relatively permanent. On the other hand, female intersex lines take many more generations and are less stable. These lines have been extensively examined for their 4th chromosome constitutions (Fung and Gowen, 1960). The male intersex lines seldom show more than two 4th chromosomes. On the other hand, the female intersex lines rarely show two 4th chromosomes but generally have more than three, the number sometimes going as high as four. More tests are needed but the evidence would seem to indicate that the fourth chromosome does have sex genes. These genes, contrary to the first notion of Bridges, are more frequently of the female determining type than of the male determining type. This would make the 4th chromosome like the X in that it carries an excess of female influencing genes and is not like the rest of the autosomes which have an excess of male determining genes. These observations are of particular interest in view of Krivshenko's (1959) paper. In this investigation on D. busckii, cytologic and genetic evidence was presented for the homology of a short euchromatic element of the X and Y chromosome with each other and also with the 4th chromosome or microchromosome of D. melanogaster. This conclusion is based on (1) observed somatic pairing of the X and Y of D. busckii by their proximal ends in ganglion cells and the conjugation of the short euchromatic elements of these chromosomes at their centromeric regions in the salivary gland cells; (2) the presence in the short Y chromosomal element of normal allelomorphs to four different mutant genes of the short X chromosomal element; (3) the presence in the short element of the D. busckii X chromosome of chromosome IV mutants: Cubitus interruptus. Cell and shaven of D. melanogaster. These considerations furnish proof for the homology of this X chromosomal element with the 4th chromosome of Drosophila.
These observations of Krivshenko support our findings that the 4th chromosome of D. melanogaster has an excess of female determining functions. It would further show that autosomes may behave differently with regard to their sex-determining properties according to the chance distribution of sex genes which happen to fall within them as they do in Rumex (Yamamoto, 1938j. The finding that the chromosome IV has a bias toward female tendencies further strengthens Bridges' multiple sex gene theory and weakens the theory of an all-or-none action of the whole X chro
IV. Sex under Special Conditions
A. Species Hybridity
Hybrid progeny coming from species crosses are apt to represent but a very few of the possible genotypes of the total number that conceivably could come from the gene pool. The hybrid phenotypes may display three kinds of characteristics. The common set is that derived from genes in either or both parents through ordinary meiotic segregations and dominance. The second set shows intermediate development of the characters found in the two parent species. The third set of characters that complete the animal is new to those observed in either parent species. These new characters may be the loss of a few dorsocentral and scutellar bristles, broken or missing cross veins, or abnormal bands in the abdomen as in D. simulans x D. melanogaster hybrids (Sturtevant, 1920b), extra antigenic substances as found in dove hybrids (Irwin and Cole, 1936), or more numerous characteristics as in the mule. Frequent among these new characteristics is sterility. The sterility may extend to either or both sexes and affect the secondary sex ratios. As Sturtevant (1920b) points out, crosses between the domestic cow and male bison give male offspring with humps derived from the bison which are so large as to prevent their being born alive. The female hybrids lack these humps and are conse(iuently born normally. The abnormal sex ratio observed at time of birth is due to causes external to the hybrid itself and attributable to the structure of the mothers.
A comparable case was found during the study of female sterility in interspecies hybrids of Drosophila pseudoobscura in which Mampell (1941) showed that in the hybrids of certain strains, the females produced no or few offspring because of interspecies lethal genes connected with a maternal effect. Comparable cases as well as those dependent on other mechanisms are known for other groups. The progeny may also be altered to give new sex types, generally intersexes. These intersexes often replace either the male or the female sex group. However, despite their apparent relation, the changes in the sex ratio and the appearance of intersexes can have different causes. D. simvlans X D. melanogaster hybrids emphasize that there may be no relation between the peculiar hybrid sex ratios and the intersexes since extreme differences in sex ratio occur but no intersexual types.
Species, however, may have natural differences in the sex potencies of their X chromosomes and/or their autosomes. In crosses between D. repleta and D. neorepleta involving a sex-linked recessive white-eyed mutant type of D. repleta Sturtevant (1946) obtained about 15 per cent fertile matings in 500 mass cultures, a total of 532 females to 635 males. All progeny as expected were wild type in character. The males, however, had long narrow testes and were totally sterile, a condition later shown to be due to a gene in the X chromosome located near the white locus. Females suggested intersexuality in having three anal plates instead of the usual two. Mating of Fi hybrid females to white D. repleta males gave 9 per cent fertility, the 179 offspring being distributed as 70 wild type females, 9 white females, 42 wild type males, and 58 white males, although the expectation for the classes was equality. Evidence indicates that some of the 9 white females were intersexes as were possibly some of the white males. The wild-type males again had the long narrow testes and sterility of the Fi male progeny. Wild type females were moderately fertile. By continued backcrossing to 1). repleta males having white or whitesinged, a female line was picked up which continued to have the unusual sex ratios but had more fertility. It was presumed that the D. neorepleta gene responsible for the unusual ratios was originally associated with MHother gene that decreased fertility in females largely of D. repleta constitution and that the foundation female for the more fertile line came as a result of a crossover between an infertility gene and that responsible for the unusual sex ratios. Continued back crosses of females of this line to white D. repleta males have been made. Out of 33 fertile cultures, 16 gave approximately equal ratios of wild-type and white females, wild-type and white males; and 17 gave 472 wild-type females, 5 white females, 63 white intersexes, 482 wild-type males, and 339 white males. The white females presumably represented crossovers between the loci of white and the critical gene in the X derived from D. neorepleta. The intersexes were of extreme type with gonads very small (rudimentary ovaries in those cases where they were found at all). External genitalia were missing or of abnormal male type. Other somatic characteristics included weakness which prevents emergence and accounts for the loss of about 88 per cent of the flies expected in that class. The intersexual condition was suggested as being caused by an autosomal dominant gene derived from D. neorepleta which so conditions the eggs before meiosis that two D. repleta X chromosomes result in the development of intersexes rather than females. The action of this gene occurs before meiosis and may in fact be absent from the intersexes themselves. This was confirmed by crosses of white brothers of the intersexes to pure D. repleta females when the offspring were normal for both sexes; but when these Fi daughters were mated to D. repleta males only intersexes and males resulted. This last cross further showed that although this gene was derived from D. neorepleta in D. neorepleta cytoplasm, the D. neorepleta cytoplasm was not necessary for the intersexes to result. The case also has an important bearing on the location of the sex-determining factors, for in this cross the characteristics were only secondarily governed by the cytoplasm through earlier determination by genes of the mothers' nuclei.
Significant parallels are found between
the autosomal gene of D. neorepleta and
the third chromosome Ne gene of D. melanogaster (Gowen and Nelson, 1942) described in the section on high male sex ratio. The D. neorepleta gene caused the
cytoplasms of the eggs laid by mothers
carrying it to become more male potent.
The female potencies of two X chromosome
D. repleta zygotes were unable to balance
these male elements. Many died late in development. Those able to emerge became
intersexes. The Ne gene also sensitizes the
cytoplasms of all eggs of mothers carrying
it causing any 2X + 2A, 3X + 3A, 3X +
2A or 2X -h 3A intersexes of female type to
die in the eggs at 10 to 15 hours whereas
males XY + 2A and male-type intersexes
live.
Other mechanisms for causing sex and sex ratio changes are known, i.e., Cole and Hollander (1950), but few are as well worked out as that of D. repleta x D. neorepleta. New mechanisms will certainly be found for the opportunities for genetic analysis of sex in hybrids are many.
B. Mosaics For Sex
Recent genetic work has emphasized the fact that individual D. melanogaster may be composed of cells of more than one genie or chromosome constitution. The main type of sex mosaic is the gynandromorph composed of cells of female constitution on one side, XX + 2A, and male, X + 2A on the other, the loss of the X chromosome coming at an early cleavage (Morgan and Bridges, 1919; L. V. Morgan, 1929; Bridges, 1939). The mosaic areas are large since the cells of each type may be in nearly equal numbers.
At the other extreme Stern (1936) has shown that phenotypic mosaics may develop as a consequence of the somatic chromosome pairs crossing over at late stages in embryologic development. Special genes, Minutes, materially increase the frequencies of these crossovers. The proportion of the body occupied by the cross-over type cells is small because crossing over takes place so late in development.
Recently another agent in the form of a
ring chromosome has been discovered which
greatly increases the production of sex mosaics. Some ring chromosomes are relatively
stable whereas others are quite unstable,
the instability depending to some extent on
aging of the eggs and environmental factors
(Hannah, 1955). The instability is manifest by frequent gynandromorphs, XO males,
and dominant lethals among the rod and
ring zygotes. It has been suggested that the
instability is due to heterochromatic elements. Hinton (1959) has observed the
chromosome behavior of these types in
Feulgen mounts of whole eggs that were in
cleavages 3 to 8. He found strikingly abnormal chromosome behavior in these
cleaving nuclei. For some cell divisions
chromosome reproduction was interpreted
as being through chromatid-type breakage
fusion bridge cycles. As a result of this behavior mosaics are formed which are intermediate between those of the half gynandromorphs and those which occur much
later because of somatic crossing over. In
terms of volume of cells included, the abnormal types may include only a few cells
of the total organism, a fair proportion of
the cells, or a full half of the whole body.
These unstable ring chromosome mosaics
may be a part of the secondary reproductive system or for that matter any other
region of the body. When the mosaic cells
are incorporated in the region of sex organ
differentiation male or female type organs
or parts of organs may develop as governed
by the cell nuclei being X, XX or some fraction thereof.
Gynandromorplis appear sporadically and
rarely in many species but in some instances genes which activate mechanisms
for their formation are known. In the presence of recessive homozygous claret in the
eggs of D. siniulans, gynandromorphs constitute a noticeable percentage of the
emerging adults. The gene nearly always
operates on the X received from the mother
causing it to be eliminated from the cell.
The resulting gynandromorphs are similar
to those of D. melanogaster. The fact that
the claret gene should affect the X and a
particular X chromosome is suggestive of
the manner in which given chromosomes arc
eliminated in Sciara. Other types of sex
mosaics will be found in the descriptions of
other species, i)articularly in the Hymenoptci'a.
C. Parthenogenesis in Drosophila
Parthenogenesis is of interest as it changes the sex ratios in families and brings to light new sex types and novel methods for their development (Stalker, 1954), A survey of 28 species of Drosophila showed a low rate of parthenogenesis in 23 species. Adult progeny were obtained for only 3 species. For D. 'parthenogenetica the original rate was 8 in 10,000 whereas that for D. polymorpha was 1 in 19,000. These rates could be increased by selection of higher rate parents: 151 and 70 per 10,000 unfertilized eggs of the first and second species respectively.
D. parthenogenetica diploid virgins produced diploid and triploid daughters as well
as rare XO sterile diploid males. Triploid
virgins produced diploid and triploid females and large numbers, 40 per cent, of
sterile XO diploid males. Diploid virgins
heterozygous for sex-linked recessive garnet
produced homozygous and heterozygous
diploid females as well as +/+/g and
+ /'g/g triploid females. No homozygous
wild-type or homozygous garnet triploid females or garnet mosaics were found. Diploid females crossed to fertile diploid males
produced few if any polyploid progeny or
jjrimary X chromosome exceptional types.
Of the unfertilized eggs from diploid virgins
which started development, 80 per cent died
in late embryonic or early larval stages.
The parthenogenesis in diploid females depended on two normal meiotic divisions followed by fusion of two of the derived haploid nuclei to form diploid progeny, or the
fusion of three such nuclei to form triploitl
progeny. In the triploid virgins similar fusions of the maturation nuclei may produce
diploid and triploid females but the large
number of diploid XO sterile males were
picsunicd to be the result of cleavage without prior nuclear fusion. Such cleavages
without fusion in eggs of dijiloid virgins
would lead to the production of haploid
embryos. They were presumed i-esponsible
for the large early larval and embryonic
<h'atlis. These observations have been confirmed by the study of S|)rackling (1960)
involving some 2200 eggs at various stages
of cleavage. Evidence from XXY diploid
virgins indicated that biiuiclcar fusion in
unfertilized eggs involved two terminal
haploid nuclei or two central nuclei. The
fact that tetraploids were not observed as
progeny of ti'iploid vii'gins was considered
indicative of relative inviability of this type. Successful parthenogenesis was under
partial control of the inheritance as 80 generations of selection increased the rate
about 20-fold. Similarly outcrossing to bisexually reproducing males and reselection
resulted in both pronounced increases in
and survival of the parthenogenetic types.
Carson, Wheeler and Heed (1957) and
Murdy and Carson (1959) have established
a strain of Drosophila mangabeirai with
only thelytokous reproduction. Males have
been captured in nature but are rare. Fecundity of the virgin females is low but the
egg hatch is 60 per cent and 80 per cent
survive to adult stage. The progeny of the
virgins are always diploid. In meiotic spindle formation D. mangabeirai differs from
other Drosophila species in that its orientation increases the probability for fusion of
two haploid nuclei into structurally heterozygous diploid females. The study of
Feulgen whole mounts of freshly laid eggs
indicated automictic behavior with two
meiotic divisions followed by a fusion of
two of the four haploid meiotic products.
The absence of adult structural homozygotes in wild populations is probabl}^ explained by death during early development
and by possible fusion of second division
meiotic products derived from different secondary oocytes. Stalker (1956a) postulated
such selective fusion in order to account for
the heterozygous condition in females of
Lonchoptera dubia, an automictic, parthenogenetic fly in which males are rare or unknown.
A
case in which jihenotypically rudimentary females, supposed homozygous for r/r
give rare and unexpected type progeny, has suggested that polar body fusion may also take place in D. melanogaster (Goldschmiclt, 1957). The rudimentary mothers producing the peculiar type are interpreted as formed by a most unusual series of events: a fertilization nucleus derived from the fusion of an r containing egg fertilized by an r containing sperm and a polar copulation nucleus derived from the fusion of a polar body containing an r genome and one having wild type. Both cell types become incorporated into the ovary. The progeny which come from these supposed rudimentary mothers are presumed to be derived from the maturation into eggs of the cells derived from the heterozygous i)olar copulation nuclei. If these progeny-producing rudimentary mothers arise in the presumed manner they give the basis for a parthenogenetic mode of reproduction and the sex types which have been described in other Drosophila species by Stalker and Carson. There are similar, as well as other forms of parthenogenesis which affect sex (Smith, 1955) or assist in maintaining trijiloid conditions (Smith-White, 1955). A number of these types have been reviewed by Suomalainen (1950, 1954). The reader may be referred to this material for other cases and chromosome behaviors.
D. Sex Influence of the Y Chromosome
The first function discovered for the Y chromosome in D. melanogaster was that it was necessary to male fertility (Bridges, 1916). Two and possibly more Y chromosome-borne, genetic factors were involved (Stern, 1929). Gamete maturation when these factors were lacking ceased just short of the sperm's becoming motile (Shen, 1932). The motility conferred on the sperm by the presence of the Y chromosome factors was fixed for the testes at an early stage of development as transplantation experiments, sterile testes to fertile larvae and fertile testes to sterile larvae, showed motility to be a property determined by early localized somatic influences on the developing gametes or predetermined in the diploid phase (Stern and Hadorn, 1938). This Y chromosome function was sex limited, because females without a Y were the normal fertile females and those with an extra Y also were fertile.
Neuhaus (1939) followed by Cooper
(1952, 1959) and Brosseau (1960) further
analyzed the Y chromosome for fertility
loci. The latter showed at least two fertility
loci on the short arm and five on the long
arm of the Y chromosome. Data are compatible with a linear order of the genes. An
additional fertility factor common to the X
and Y was suggested. The Y chromosome
fertility factors consequently fall in line
with Bridges' concept of multiple gene loci
distributed in a more or less random manner which may affect sex.
The Y chromosome has other attributes
which help to explain its significance to secondary if not primary sex characteristics.
As with many other species it has loci for
genes which are also found in the X chromosome, i.e., bobbed, as well as a limited
number of bands in the salivary gland chromosome. Nucleolus organizers and at least
two specialized pairing organelles similar
to those in the X chromosome are found
within the Y chromosome (Cooper, 1952,
1959). One of the most significant properties is the effect of extra Y chromosomes on
the variegation observed for various gene
phenotypes either in the normal chromosome pattern or in that accompanying
translocation. In variegation one Y chromosome as extra to the normal complex is
sufficient to eliminate or more rarely to
much reduce the variegated expression
(Gowen and Gay, 1933). When the Y chromosomes are 2 above the normal complement, the phenotypes gain two new features
(Cooper, 1956). Both males and females
become variegated in the expression of their
eye characteristics. The males become sterile. These effects are unexpected for they
are counter to any previous trends in the Y
effects on these characteristics. They have
reversed the direction of the effects as established by the two previous chromosome
types. The variegation of the XX2Y + 2A
females and X3Y + 2A males resembles
that of the XX + 2A females and XY +
2A males but is more extreme, whereas the
XXY + 2A and X2Y + 2A are largely
nonvariegated. The fertility relations are
equally aberrant: the X3Y + 2A males
have the same type sterility through loss of
sperm motility as that of the XO + 2A
males. Bundles of sperm are formed but
they do not become motile. Full-sized Y
chromosomes are not required to bring
about these effects, because females having
a whole Y plus a piece of a second, or males
with 2Y plus a piece of a third, will show
the effects.
The fractional Y chromosomes furnish
opportunities to test for the partial independence of the variegation and sterility effects. The two Y hyperploid males differ in
their degree of fertility according to the
fraction of the Y chromosome which may
be present whereas the effects on variegation may be constant among groups. This is
in accord with the sterility being in part independent of the factors causing the variegations. Similarly, the variegations may
be shown to be partially free of the action
of some elements that are not themselves
members of the two sets of factors influencing fertility in the normal male.
Other phenotypic irregularities appear;
eye facets may be roughened, legs shortened, and wing membranes become abnormal. On the negative side two extra Y chromosomes in females homozygous for the
transformer gene, tra, do not increase in
maleness or function.
The variegations of the so-called V-type
position effects with translocations are suppressed, as in the normal type described,
by one extra Y but are nonetheless variegated when two extra Y chromosomes are
present. That these effects are caused by
there being two supernumerary Y's is indicated by the fact that XX2Y females are
fertile and X3Y males sometimes lose a Y
chromosome in their germinal tracts and
become fertile. A number of mechanisms
have been suggested to account for these
results but most have proven unsatisfactory. A balance interpretation for the X, Y
and autosomes like that for sex, as suggested by Cooper (1956) is compatible with
the somatic cell variegations for euchromatic loci transferred to the heterochromatic regions and the sterility-fertility
relations expressed by the different chromosomal types.
The variegation effects of the Y chromosome take on further significance. Baker
and Spofford (1959) have shown that 15
different fragments of the Y chromosome
when studied for their contributions to variegation differ in their effects with the differences often not related to the size of the
fragment, thus indicating that the Y chromosome has linearly differentiated factors
capable of modifying the variegated phenotype.
Other indications of genetic activity of
the Y chromosome were given by Aronson
(1959) in her study of the segregation observed in 3rd chromosome translocations. A
deficiency for the region of the 3rd chromosome centromere is lethal when homozygous. Both males and females are fully
\'iable when this deficiency is heterozygous.
XO or haplo IV deficient males die. However, in the presence of a Y chromosome the
deficient haplo IV males become viable.
The lability of this last class indicates that
the Y chromosome is genetically active, can
compensate for the autosomal deficiency,
and thus alter the progeny sex ratio.
In D. virilis the situation is somewhat
different from that observed by Cooper in
1956 in D. melanogaster. Baker (1956) has
shown that, in a translocation, males having
two Y chromosomes plus a Y marked with
a 5th chromosome peach are fertile. These
results seem to indicate a species difference
in the effect of the extra Y on fertility or
the Y with the inserted peach locus is not
a complete Y and in consequence the true
composition of these males is X + 2Y plus
a fragment of the Y. The X chromosomal
associations in the multichromosomal types
are shown to be by trivalents or by tetravalents. The segregation data indicate that
the pattern of disjunction of trivalents is a
function of the particular Y chromosome
involved. In X2Y males with normal Y's or
with one normal and one marked Y, the Y's
disjoin almost twice as frequently as they
do from trivalents with two identical Y's.
Tetravalent segregation is almost entirely
two by two, with no preference for any of
the three types of disjunction.
An odd situation was reported by Tokunaga (1958) in substrains of Aphiochaeta vanthina Speiser. When a male of the substrain was crossed to individuals bearing
3rd chromosome genes of the original
strains, the mutant genes for brown, and so
on, behaved as if they were partially sexlinked in the following generations. On the
other hand, when a female carrying the partially sex-linked genes on the X and Y
chromosomes. Abrupt or Occhi chiari, was
crossed to the male of the substrain the
characters segregated as though they were
autosomal. As a working hypothesis it was
suggested that in this species the Y chromosome had the major male determining
factors. The "special" male arose as a translocation of these factors to the third chromosome with the consequent change in linkage
relations. Data on the role of the X chromosome in sex determination in this species
have not yet been obtained but if they support the interpretation they indicate real
differences between this species and that of Drosophila in the location of the sex genes.
The results are reminiscent of those obtained by Winge in Lebistes as well as those
in Melandrium and other forms in which
the Y or W chromosomes may contain
strong sex genes for either sex. They would
further support the thesis that sex genes
may be distributed to almost any loci within
the inheritance complex.
E. Maternal Influences on Sex Ratio
Aside from chance and specific genetic factors, sex ratio is subject to effects from agents intrinsic in the cells of the mothers (Buzzati-Traverso, 1941; Magni, 1952). Strains of Drosophila bifasciata (Magni, 1952, 1953, 1957), D. prosaltans (Cavalcanti and Falcao, 1954), D. willistoni and D. paulistorum Spassky, 1956 have been isolated which were nearly all of the female sex even though the mothers were outbred to other strains having normal ratio bisexual progeny. Study of these strains by the above workers and Malogolowkin (1958) IVIalogolowkin and Poulson (1957), and Malogolowkin, Poulson and Wright (1959), as well as by Carson (1956) have shown inheritance strictly through the mother regardless of the genetic nature of the males to which they were bred. The unbalanced ratio was retained even when the original chromosomes had been replaced by homologous genomes from lines giving normal male and female progenies. Transmission of this unbalanced sex ratio was through the cytoplasm. Eggs fertilized by Y-bearing sperm died early in the course of development. Malogolowkin, Poulson and Wright (19591 have shown that the high female ratio and embryonic male deaths may be transferred from affected females to those which do not normally show the condition through injection of ooplasm from infected females. Ooplasm of this same type is, w^hen injected into males, suflficient to cause death to occur within 3 days. In the females a latent period of 10 to 14 days after ooplasm injection was apparently necessary to establish egg sensitization. Once established the condition could be transmitted through the female line for several generations. The ooplasm injections were not as efficient in establishing these lines as females found naturally infected. Unisexual broods may fail to appear, in some instances fail to transmit the condition or produce intermediate progeny ratios, as well as in some instances to skip a generation. The infectious agent was found to vary in different species. Magni (1953, 1954) for D. bifasciata found that temperature above normal tended to remove the cytoplasmic agent making it ineffective. This result has a parallel to the action of temperature on certain viruses, as for instance that involved in one of the peach tree diseases. On the other hand, Malogolowkin (1958) ior D . willistoni found no temperature effect. Malogolowkin further found that the cytoplasmic factor was not independent of chromosomal genes since some wildtype mutant strains induced reversions to normal sex ratio. Recently Poulson has established the complete correlation of the high female progeny characteristic with the presence of a Trepomena in the fly's lymph. As with mouse typhoid variations, lethality was genotype dependent.
These cases have interest from more than
the sex ratio viewpoint. Cytoplasmically
inherited susceptibility of some strains of
D. 7nelanogaster to poisoning by carbon dioxide as studied by L'Heritier (1951, 1955)
depended on the presence of some cytoplasmic entity which passed through the egg
cytoplasm and also, but less efficiently, by
means of the male sex cells to the progeny.
The carbon dioxide susceptibility was transmitted through injections of hemolymph or
transplantation of organs of susceptible
strains. The transmissible substance had a
further property of heat susceptibility. The
COo susceptibility differed from that of "sex
ratio" in being partially male transmitted.
It agrees with sex ratio" D. bifasciata in
being heat susceptible but differs from D.
willistoni "sex ratio" in this respect.
D. bifasciata may behave differently than
D. willistoni in that Rasmussen (1957) and
Moriwaki and Kitagawa (1957) both conducted transplantation experiments with
negative results. However, it is jiossible that
these experiments may be affected by a
different incubation period for the injected
material as contrasted with that for D.
willistoni. A range of possibilities evidently
exist for extrinsic effects in sex ratio.
Carson (1956) found a female producing strain of D. borealis Patterson, which carried on for a period of 8 generations, produced 1327 females with no males. The strain showed no chromosomal abnormalities. It had 3 inversions. The females would not produce young unless mated to males from other strains having biparental inheritance. This requirement, together with gene evidence, showing that the females were of biparental origin, is against the female progenies being derived by thelytokous reproduction.
F. Male-Influenced Type of Female Sex Ratio
Sturtevant in 1925 described exj^eriments with a stock of D. affinis in which a great deficiency of sons was obtained from certain males, regardless of the source of females to which such males were mated. The few males obtained from such matings were normal in behavior but some of the sons of females from such anomalous cultures again gave very few sons (]Morgan, Bridges and Sturtevant, 1925).
Gershenson (1928) in a sampling of 19
females caught in nature found two that
were heterozygous for a factor causing
strong deviations toward females (96 per
cent to 4 per cent males) whereas the normal D. pseudoobscura ratio was nearly 1 to
1. The factor was localized in the X chromosome and was transmitted like an ordinary
sex-linked gene. Its effect was sex limited
as it was not manifest in either heterozygous or homozygous females. It had no
effect on the development of zygotes already formed but strongly influenced the
mechanism of sex determination through the
almost total removal of the spermatozoa
with the Y chromosome from the fertilization process. Egg counts showed the divergent ratio toward females was not caused
by death of the male zygotes inasmuch as
there was no greater mortality from such
cultures than from controls giving 1 to 1 sex
ratios.
Sturtevant and Dobzhansky (1936) showed that an identical or nearly identical phenotypc to that obscM'ved in Drosophila obscura was present in D. pseudoobscura. The D. pseudoobscura carrying the factor were scattered over rather wide geographical areas. Comparable types also were found in two other species D. athabasca and D. azteca. This genie "sex ratio"' sr lies in the right limb of the X of races A and B of D. pseudoobscura. Like the other cases analyzed, males carrying sr have mostly daughters and few sons regardless of the genotypes of their mates. Structurally the female sex ratio came through modification of the development of the sex cells of the male to give a majority of X-bearing sperm. Cytologic study showed that in "sex ratio" males the X underwent equational division at each meiotic division, whereas the autosomes behaved normally. The Y chromosome lagged on the first spindle, remained much condensed, and its spindle attachment end was not attenuated. The Y chromosome was not included in either telophase group of the first meiotic division but was left behind. It was sometimes noted in one of the daughter cells where it formed a small nucleus. Among 64 spermatocytes examined, all had an X chromosome and none a Y chromosome in their main nuclei. The micronuclei containing Y played no further part in the division, became smaller and exceedingly contracted. The final fate of the micronuclci was uncertain, but there was no indication that any spermatids died or were abnormal.
G. High Male Sex Ratio of Genetic Origin
In 1920, Thompson described a recessive mutant in D. melanogaster which killed all homozygous females but changed the male phenotypes only slightly, the wings standing erect above the back. The locus of the mutant was 38 in the X chromosome. This mutant was a leader for a class in which the genes affect only one sex but are innocuous to the other.
Bobbed-lethal, a sex-linked gene found i)y Bridges, is a gene of this class but one in which the mechanism of protection to the other sex is known. It kills homozygous females but does not kill the males because of the wild type allele which the males have in their Y chromosomes. The presence of this bobbed-lethal in a population consequently leads to male ratios higher than wild type.
A recessive gene in chromosome II, discovered by Redfield (1926), caused the
early death of the majority of female zygotes and led to a sex ratio of about 1 female to 5.5 males. The effect was transmitted througli both males and females. The
missing females may have died largely in
the egg stage although disproportionate
losses also occurred in the larvae and pupae.
The maternal effect was attributed to an
influence exerted by the chromosome constitution of the mother on the eggs before
they left the mother's body. Because of this
maternal lethal, the families from these
mothers have the normal number of sons
but few or no daughters.
A culture was observed by Gowen and
Nelson (1942) which yielded only male
progeny, 136 in all. Some of the male progeny were able to transmit the male-producing characteristics to half of their daughters
without regard to the characteristics of the
mates to which they were bred. The inheritance was without phenotypic effect on
the males. When present in the heterozygous condition in females, the female's
own phenotype gave no indication of the
gene's presence. The inheritance was sex
limited in that it affected only the eggs laid
by the mothers carrying it. In these eggs it
acted as a dominant lethal for the XX zygotes. The gene was found located in the 3rd
chromosome between the marker genes for
hairy and for Dicheate at approximately 31.
The X eggs carrying the gene, Ne, died in the
egg stage at between 10 and 15 hours under
25°C. temperature. The gene was as effective in triploids as in diploids. One dose
of this gene in triploids caused them to have
only male progeny and male type intersexes.
The presence of this gene caused the elimination of any embryos with chromosomal
capacities for initiating and developing the
primary or secondary female sexual systems. Since the gene itself was heterozygous
in the female the meiotic divisions of the
eggs would cause the gene to pass into the
polar bodies as often as to remain in the
fertilization nucleus yet the lethal effects
are found in all eggs. The antagonism was
between the egg cytoplasm and the XX
fusion products. Supernumerary sperm of
the Y type were not sufficient to overcome
the lethal effects. An XY fertilization nucleus was necessary for survival.
This case has parallel features with that
observed by Sturtevant (1956) for a 3rd
chromosome gene that destroys individuals
bearing the first chromosome recessive gene for prune. The gene responsible was found
to be a dominant, prune killer, K-pn, which
was located in the end of the 3rd chromosome at 104.5. Prune killer was without
phenotypic effect so far as could be observed except that it acted as a killer for
flies containing any known allele of prune.
It was equally effective in either males or
females, hemizygous or homozygous, for
prune. The larvae died in the 2nd instar
but no gross abnormalities were detected
in the dying larvae. This case has interest
for the Ne gene in that the killer gene occupies a locus in the 3rd chromosome although removed by about 70 units from Ne,
and acts on a specific genotype, the prune
genotype, whereas Ne acts on the specific
XX genotype. The known base for action
of the prune killer is genetically much narrower than that for the female killer, Ne,
in that prune killer acts on an allele within
a specific locus in the X chromosome, and
Ne acts on a type coming as a product of
the action of two whole sex chromosomes. It
is, of course, conceivable that when ultimately traced each action may be dependent
upon specific changes of particular chemical syntheses. The action of these genes is
also of interest from another viewpoint.
In mice there is a phenotype caused by
the homozygous condition of a recessive
gene (Hollander and Gowen, 1959) which
acts on its own specific dominant allelic
type in its progeny so as to cause an increased number of deaths between birth
and two weeks of age, as well as causing the
long bones to break and the joints to show
large swellings. The lethal nature of this
interaction is not a product of the mother's
milk nor does it show humoral effects such
as those observed with erythroblastosis in
the human. The interallelic interaction is
sex limited in that it is confined to the
mother and is without effect when the male
has the same genotype.
H. Female-Male Sex Ratio Interactions
A case in D. affinis involving the interaction of a "female sex ratio" factor and an autosomal "male sex ratio" factor has been studied by Novitski (1947). Starting from stock which had a genetic constitution for the "sex ratio" X chromosome which ordinarily causes males carrying it to produce only daughters, he was able to establish a recessive gene in chromosome B in whose presence only male offspring resulted. The genetic constitution of the male was alone important. High sterility accompanied the "male sex ratio" males breeding performance. The "male sex ratio" parents yielded 95 fertile cultures having an average of 25 individuals per culture. Females appeared in 10 of these cultures with an average of 3 per culture for those producing females. The total sex ratio was 77 males per female. The males were morphologically normal. They carried an X chromosome of their mothers. Cytologic observation of spermatogonial metaphases of 3 progeny showed that the Fi males may or may not have carried a Y of their fathers. This agreed with the tendency for sterility in these "male sex ratio" males. The ventral receptacles of the females from 4 such sterile cultures when examined proved devoid of sperm. The occasional female offspring of the "male sex ratio" males had one X chromosome from each parent. Females mated to "male sex ratio" males show large numbers of sperm in the ventral receptacles (7 out of 8 cases) , although such females had usually produced no or very few offspring. The sperm, if capable of fertilization, must have had a lethal effect on the zygotes. The recessive nature of the factor in chromosome B indicated that its potential lethal effect originated during spermatogenesis rather than at the time of fertilization. This lethal period corresponds to that when the "female sex ratio" factor in the X chromosome is active.
The research on sex ratio in Drosophila
reviewed shows that through the interplay
of the sex chromosome-located and autosomal-located factors all types of sex ratios
from only females in the family to only
males in the family may be generated.
V. Sex Determination in Other Insects
Sciara, a fungus gnat, offers sex-determining mechanisms quite different from any yet offered in Drosophila. Sciara is unique, yet in its uniqueness, it illustrates basic facts that were rediscovered in other species only through the study of abnormal forms some of which were created as induced developmental, chromosomal, or hormonal abnormalities. The complexities responsible for the remarkable facts were analyzed by Metz and his collaborators. The following brief review is based upon Metz's (1938) summarization of the chromosome behavior problem to which the reader is referred for further information. Of Sciara species studied 12 out of 14 have their basic chromosome groups composed of three types of chromosomes: autosomes, sex chromosomes, and "limited" chromosomes. Two species, S. ocellaris Comstock and S. reynoldsi Metz, lack the "limited" chromosomes. Two sets of autosomes and three sex chromosomes are found in the zygotes of all Sciara species at the completion of fertilization. The behavior of these chromosomal types will be considered in sequence.
The autosomes behave normally in somatic mitosis and in oogenesis. There is
cytologic and genetic evidence for synapsis,
crossing over, random segregation, and regular distribution of chromosomes and genes
in the female.
In spermatogenesis the story is quite different. The first maturation division is unipolar. Genetic evidence shows a complete,
selective segregation of the maternally derived autosomes and sex chromosomes from
those of paternal origin. The paternal homologues move away from the pole, are
extruded, and degenerate. The second
spermatocyte division is likewise unequal
and one of the products, a bud, degenerates.
Thus a spermatogonial cell passing through
meiosis gives rise to only one sperm. At the
second spermatocyte division all of the
chromosomes (maternal homologues) except
the sex chromosome undergo an equational
division but both halves of the sex chromosome enter into the same nucleus. This nucleus becomes the sperm nucleus. The chromosomes at the opposite pole form a bud
and degenerate. Fertilization of the Sciara
egg is normally monospermic not polyspermic.
The sex chromosomes XXX of the fertilization nucleus are derived, two from the
sperm and one from the oocyte. Their destiny depends on whether the egg they are in develops into a male or a female imago.
In male early development, those nuclei
which are to become male soma lose the two
sex chromosomes XX, contributed by the
father's sperm, to give X + 2A soma. These
chromosomes fail to complete mitosis at
the 7th or 8th cleavage division and are left
to degenerate in the general cytoplasm when
there are no true cells in the soma region
and no membranes surround the nuclei.
Those embryos which are to become female
soma at the same cleavage cycle eliminate
but one paternal X chromosome. On a chromosome basis the soma cells respectively
become X, plus two sets of autosomes, give
rise to males on differentiation, or become
XX + 2A and develop female organs (Du
Bois, 1932). The germ line nuclei for each
sex, on the other hand, remain unrestricted
in their development. They retain their
XXX constitutions until the first day of
larval life, or about 6 hours before the formation of the left and right gonads (Berry,
1941), when they eliminate a single paternally derived X. The X which is rejected or
makes its own exit is always one of two
sister chromosomes contributed by the
father. The process of loss is strikingly different from that noted in the soma-building
nuclei. Both cell walls and nuclear membranes are present. The path of the X chromosome is through the nuclear membrane
into the cytoplasm where degeneration
eventually takes place. The loss occurs at
a time when there is no mitotic activity and
the chromosomes are separated from the
cytoplasm by an intact nuclear membrane.
Some Sciara species are characterized by
females which produce only unisexual families — practically all females or practically
all males. Genetically, the sex of progeny is
accounted for by sex chromosomes, X' and
X, which are so designated because of their
physiologic properties. Mothers having only
female progeny are characterized by always
having the X' chromosome in all their cells,
X'X + 2A, whereas mothers whose progeny
are all males are XX + 2A (Moses and
Metz, 1928). The all female and all male
broods are observed in about equal numbers. The male has no influence on sex ratio.
Normally the progeny in all female broods
will be X'X + 2A or XX + 2A in soma and germ lines, whereas the all male progeny broods will be only X + 2A in the soma
and XX + 2A in the germ line.
Studies of Grouse (1943, 1960a, b) show
nondisjunction may occur and eggs entirely
lacking sex chromosomes or having double
the normal number may be produced. When
the nondisjunctional eggs lacking X chromosomes are from X'X females and are fertilized by sperm contributing two sister X
chromosomes, one X (as expected of X'X
mothers, not two as expected of XX cells) is
eliminated at the 7th or 8th cleavage to give
cells which develop into "exceptional" males
with XO + 2A soma and XX + 2A germ
line, all X chromosomes being of paternal
origin. The casting out of the single X chromosome is consequently controlled by the
characteristics of the egg cytoplasm derived
from the X'X mother rather than by the
kind of chromosomes present in the nuclei
at the 7th or 8th cleavage stage of the eml)ryo.
Similarly when nondisjunctional eggs
having both XX chromosomes of the XX
mothers are fertilized by the normal spermbearing XX chromosomes, the zygotes are
XXXX. At the 7th or 8th cleavage both
paternal X chromosomes are eliminated and
the resulting soma develops into an exceptional" female, XX + 2A, often with 3 X's
in her germ line. These observations confirm the earlier interpretations that the X'X
or XX constitution of the mother conditions
her eggs respectively to cast out 1 or 2
parental X chromosomes at the 7th or 8th
cleavage according to the cytoplasmic constitution of the egg. Somatic sex development is visually determined only after this
stage when the chromosomes of the somatic
cells take on the familiar aspect of either
XX + 2A for the females or X + 2A for the
males.
Although at present there are few examples in other species, this cytoplasmic conditioning of early embryologic development
by the mother's genotype may be a common
rather than a rare occurrence of development. The expression of the events to which
this control may lead may be quite different in different cases. The spiral of snail
shells was an early recognized case (Sturtcvant, 1923) . In Drosophila, a gene acting
through the mother's genome allows only male embryos to survive (Gowen and Nelson, 1942), suggesting that this gene acts
through her cytoplasm in a manner comparable to that of the X' and X chromosomes of Sciara. In either case the genotype would not be considered a primary sex
factor, profound as the effect on sex may
ultimately be.
Sciara also has strains or species where
the progeny of single matings are of both
sexes, bisexual, yet the chromosomal elimination mechanisms remain as described
above save for the fact that they now operate within broods rather than between
broods. Sex ratios for the families within
these bisexual strains are extremely variable, 1:0 or 0:1 ratios not being infrequent
and 1 : 1 ratios being the exception. An instance of a bisexual strain arising from a
unisexual line has been analyzed by Reynolds (1938). The females were found to be
3X in their germ line and produced 2X eggs
which developed into daughters and X eggs
which became sons. Loss of the extra X
from this line resulted in the line's return to
unisexual reproduction.
In general, bisexuality as contrasted with
unisexuality seems to be caused by differences in the X'X-XX mechanisms, either
specific for the mechanism, or induced by
modifying genes which cause different embryos within the same progeny to cast out
sometimes one or sometimes two X chromosomes from all their somatic nuclei. This
casting out occurs uniformly for all nuclei
of a given embryo as is shown by the fact
that sex mosaics or gynandromorphs are as
seldom observed in bisexual as unisexual
strains.
Some species of Sciara have large chromosomes which are in essence "limited" to
the germ line since they are lost from the
somatic nuclei at the 5th or 6th cleavage
division. Other species lack these chromosomes, yet agree with the rest in the behavior of their remaining chromosomes and
in sex determination. So far as is now
known the "limited" chromosomes seem
empty of inheritance factors influencing either sex or unisexual vs. bisexual broods or
for that matter other characteristics (their
persistence seems to make this latter unlikely I. Their presence does lead to a question of the efficacv of desoxvribonucleic acid (DNA) as the all inclusive agent in inheritance for they are clearly DNA-positive.
These cytogenetic observations clear up
most of the events leading to sex differentiation and the delayed time in embryologic
development when it takes place, but, as
INIetz points out, other puzzling questions
are raised. Observed chromosome extrusions
from cell nuclei are rarities today even
though discovered for Ascaris 60 years ago.
Why should this mechanism be so well defined in Sciara? Why should the mode of
elimination be so accurately timed and yet
be different in method and time for the
soma and germ cell lines? Two of the three
X chromosomes in the fertilized egg are
sisters from the father and should be identical. What is special in the 7th or 8th cleavage that causes elimination in the soma
plasm but not in the germ plasm? Why
should the casting out of one of these same
paternal X's in the germ plasm of both
sexes be reserved for a much later stage in
cleavage and by a different procedure?
The observations of Grouse (1943, 1960a),
obtained from translocations and species
crosses, are critical in showing that the
chromosome first moA'ing in its entirety to
the first differentiating pole of the second
spermatocyte division is the X and is the
one taking part in the later postfertilization
sex chromosome elimination of the 7th or
8th cleavage. The X heterochromatic end of
the chromosome and not the centromere region seems to be responsible for the unique
behavior of this chromosome (Grouse,
1960b). Induced nondisjunctional eggs of
X'X (female-producing) mothers having as
a consequence no sex chromosomes, when
fertilized by the normal sperm bringing in
XX chromosomes, develop into embryos
eliminating one of these X's at the 7th or
8th cleavage, as expected of the eggs of
X'X mothers, and become "exceptional"
males with X + 2A soma and XX paternal
germ line. Imagoes of these embryos become
functional but partially sterile males transmitting the paternal X, a reversal of the
normal condition. Similarly the nondisjunctional eggs retaining both X's, XX
from male-producing mothers, and having
4 X's on fertilization eliminate the two paternal X's, as occurs normally in XXX eml)ryos, to become "exceptional" females with 2X or sometimes in species hybrids 3X soma.
Phenotypic sex in Sciara on this evidence
depends on the adjustment of the X chromosome numbers and their contained genes
with this adjustment regularly taking place,
not at fertilization as in most forms like
Drosophila, but at the 7th or 8th cell cleavage of embryologic development. The X' vs.
X chromosomes of the mother predispose
her haploid egg cytoplasm to time specific
chromosome elimination. The germ line cells
on the other hand may regularly have other
chromosome numbers than the soma or exceptionally tolerate other numbers as extra
X's (Grouse, 1943), or extra sets (Metz,
1959).
Triploids of 3X and 3A soma have not
been found in pure Sciara species. Salivary
gland cells of a triploid hybrid S. ocellaris x
aS. reynoldsi have two sets of S. ocellaris and
one of S. reynoldsi chromosomes (Metz,
1959). Ghromosome banding in 2X -I- 3A,
3X + 3A larval glands showed the »S. ocellaris homologues were heterozygous. The
3X + 3A type chromosomes were of typical
female appearance. The 2X:3A-type had
X chromosomes of typical male type, very
thick, pale and diffuse. The mosaic salivaries were a mixture of the two types of
cells. These results suggest that the 3X:3A
would be a typical female. The intersexes
2X -I- 3A conceivably have a range in male
and female organ development depending
on how the S. ocellaris and S. reynoldsi
chromosomes act. If the chromosomes are
equivalent, the gene expression would be
expected to be that of a 2X to 3A or a
phenotypic intersex. If they act separately
the S. ocellaris genes would be in balance for
female determination, whereas the S. reynoldsi autosomal genes would have no X S.
reynoldsi genes to balance them and presumably would be overwhelmingly male in
effect. The phenotypic outcome would be
in doubt.
The maternally governed cleavage and
chromosomal sequence occurring before the
8th embryonic cell division, although related to the subsequent events leading to the
particular sex, is unique to Sciara and its
relatives. Phenotypic sex expression apparently starts with the somatic nuclei having
either X + 2A or XX + 2A as their chromosomal constitutions. Sciara is like Drosophila and many other genera in its basic
sex-determining mechanism. There is a sexindifferent period when the maternal genotype may influence development through
cytoplasmic substances. This is followed by
active differentiation when the sex phenotypes may be influenced by various agencies
but when passed becomes fixed for phenotypic sex. The indifferent period may represent a fairly large number of early cell generations as in amphibia or fish. Somatic
and germ cells are labile but are normally
susceptible to control only by forces developed within the organism as present
knowledge indicates for Sciara. In Sciara,
only the somatic cells conform in chromosome number to their expected phenotypes.
The germ cells show not only a regulated instability of their chromosome number but
the number is different from that of the
supporting somatic cells. The fact that sex
mosaics in Sciara may develop both an
ovary and testis in the same animal (Metz,
1938; Grouse, 1960a, b) shows localized
phenotypic sex determination occurring
fairly late in development rather than the
generalized determination that would occur if sex were established at fertilization.
The germ line chromosomal differentiation from that of the somatic tissue has
parallels in a number of fairly widely dispersed species. The significance of these
types to soma line determination and to the
reversibility or irreversibility of differential gene activation in differentiated cells
subsequent to embryogenesis has been considered by Beermann (1956). Although
these types are yet in the fringe areas of
our knowledge surrounding the general
paths taken in the evolution of sex determining mechanisms, they promise to have
more generality as clarification of gene action becomes more exact.
B. Apis and Habrobracon
Hymenopteran interi:)retations of sex determination have largely turned on studies of the honey bee. Apis mellijera Linnaeus and Habrohracon juglandis Ashmead. Dzierzon early established that the drones of honey bees come from unfertilized eggs, whereas the females, queens, and workers come from fertilized eggs so that sex differentiation is marked by an N set of chromosomes for the male and 2N for the female. This mechanism was satisfactory until it was realized that on a balanced theory the doubling of a set of chromosomes, if truly identical, should not change the gene balance and consequently not the sex. Further doubt was cast on the N-2N hypothesis for sex determination by the discovery of diploid males by Whiting and "Whiting (1925). These difficulties were resolved by Whiting (1933a, b) with the suggestion that the two genomes in the female were not truly alike but actually contained a sex locus linked with the gene for fused which was occupied by different sex alleles as Xa and Xb . In this A'iew the heterozygous condition would lead to the production of females, whereas in the haploid or homozygous condition either of these genes would react to produce males. Difficulties with this hypothesis became evident in that the ratios of males to females in certain crosses were significantly divergent from those which were expected. Evidence was collected and trials made to interpret these difficulties (Whiting, 1943a, b) by assuming that on fertilization by Xa-bearing sperm the polar body spindle would turn so as to cast out the Xa chromosome and retain the complementary Xb in the majority of cases instead of in a random half. Snell (1935) offered the hypothesis that there could be several loci for sex genes, such that the X locus might be a master locus, but when this locus was in homozygous condition the sex would then be controlled by genes in other loci in the genome. As pointed out by Bridges (1939) this hypothesis would satisfy that of sex gene balances postulated in Drosophila and of change in emphasis on loci for sex genes as observed by Winge (1934) in Lebistes. However, the work of Bostian (1939) called both hypotheses into question. Consideration of these further results led to the multiple allele hypothesis of complementary sex determination for Habrobracon (Whiting, 1943a) taking a similar form to that of the sterility relations observed for a single locus in the white clover of INIelilotus. Under this system the sex locus would be occupied by multii)le alleles, any one of which or any combination of identical alleles would be male-producing, but any combination of two different alleles would l»e female determining. By having a sufficient number of these genes the jiroduction of diploid males would be curtailed to a point at which their various frequencies could be explained. In support of this liyi)othesis, multiple alleles to the number of at least 9 were found to exist for this single sex locus.
In principle at least, the honey bee could
have the Habrobracon scheme of sex determination. Rothenbuhler (1958) has recently collected the researches which test
this possibility. Tests of the multiple allele
hypothesis as applied to the honey bee were
made by Mackensen (1951, 1955) who interpreted evidence for inviable progeny produced by mating of closely related individuals as proof that this species as well as
Habrobracon juglandis follows the multiple
allelic system. The discovery of male tissue
of bii)arental origin in mosaic bees from related i)arents was considered as further evidence for the multiple allelic theory of sex
determination (Rothenbuhler, 1957).
Most recent cytologic evidence supports
the concept that there are 16 chromosomes
in the gonadal cells of the male and 32 in
those of the female (Sanderson and Hall,
1948, 1951; Ris and Kerr, 1952; Hachinohe
and Onishi, 1952; Wolf, 1960». Hachinohe
and Onishi (1952) found 16 chromosomes
as characteristic of the meiosis in the drone.
Wolf observed a nucleus in both bud and
spermatocyte of the only maturation (equational) division.
The greatest progress has been made in
understanding the mechanisms of sex mosaicism in the Hymenoptera species. These
mosaics, although ordinarily of rather rare
occurrence, have a direct bearing on sex
determination and development. In Apis,
iiolyspermy furnishes the customary basis
for their formation. One sperm fertilizes
the haploid egg nucleus and another sperm,
which has entered the egg instead of degenerating as it ordinarily does, enters into
mitotic cleavage and eventually forms islands of haploid cells of paternal origin
among the diploid cells derived from the
fertilized egg (Rothenbuhler, Gowen and
Park, 1952). Evidence showing that genetic
influences affect the sperm nucleus toward
stimulating its independent cleavage is
found to exist in Apis material (Rothenbuhler, 1955, 1958). Tliousands of gynandromorphs have been observed in Apis, all
but a small number of which have been produced in this manner. This method of initiating sex mosaics also exists for Habrobracon (Whiting, 1943b) but is rather rare.
In Habrobracon the frequent mode has a
different origin. The gynandromorph is
formed from the cleavage products of the
normal fertilization of the egg nucleus combined with those of a remaining nuclear
product of oogonial meiosis. Under these
conditions the female tissue is 2N of biparental origin and the male tissue is N of
maternal origin (Whiting, 1935, 1943b).
This type is less frequent in Apis but one
specimen has been described by Mackensen
(1951).
A number of other ways in which sex
mosaics may occur are occasionally expressed in these species. Three different
kinds of male tissue have been observed in
individual honey bee mosaics produced by
doubly mated queens — haploid male tissue
from one father, haploid male tissue from
the other genetically different father, and
diploid male tissue of maternal-paternal
origin. In other cases, the diploid, biparental
tissue was female and associated with two
kinds of male tissue (Rothenbuhler, 1957,
1958) . Cases where the haploid portions of
the sex mosaics are of two different origins,
one paternal and the other maternal, while
the female portion is representative of the
fertilized egg, are known in Habrobracon
(Whiting, 1943b). Similarly, Taber (1955)
observed females which were mosaics for
two genetically different tissues and which
he accounted for as the result of binucleate
eggs fertilized by two sperm. Mosaic drones
of yet another type were observed by
Tucker (1958) as progeny of unmated
queens. They were interpreted as the cleavage products from two of the separate nuclei formed in meiosis. These cases represent
a number of the possible types that arise
through meiotic or cleavage disfunctions
under particular environmental or hereditary conditions.
Tucker (1958) studied the method by
which impaternate workers were formed
from the eggs of unmated queens. For this
purpose he used genetic markers, red, chartreuse, ivory, and cordovan. Observations Avore nuult' i)ii 237 workers from hotcrozygous mothers. For the chartreuse loeus. 12 to
20 per cent were homozygous, for x\w i\ory
locus 1.8 per cent, and (he cordovan locus
on lesser numbers per t-ent. An egg which
is destined to become an automictic worker,
a gynandromorph with somatic male tissue
or a mosaic male, is retained within the
queen for an unusually long time during
which meiosis is suspended in anaphase 1.
Normal reorientation of first division
spindle is i)ossibly inhibited by this aging
so that after the egg is laid meiosis II occurs
with two second division spindles on sejiarate axes as Goldschmidt conjectured for
rarely fertile rudimentary Drosophila
(19")7i. Two polar l)odies and two egg
prontich^i are formed. The polar bodies take
no fiu'ther part in develoinnent. In most of
the unusual eggs the two egg pronuclei
unite to form a diploid cleavage nucleus
which develops into a female. Rarely the
two egg pronuclei develop separately as two
haploid cleavage nuclei to form a mosaic
male. Two unlike haploid cleavage nuclei,
one descending from each of the two secondary oocytes after at least one cleavage
di\ision. unite to form a dijiloid cleavage
nucleus which develops together with the
remaintler of the haploid cleavage nuclei
to produce a gynandromorph with mosaic
male tissues. The male and female tissues
within these unusual gynandromor[ihs or
female types were identical with normal
drone or normal female tissues so were
probably haploid and diploid resiiectively.
Genetic segregation observed within the
mothers of automictic workers allows the
estimation of the distance between the locus
of the gene and its centromere. "With random
recombination and "central union" Tucker
estimates this distance for the chartreuse
locus to centromere as 28.8 units and for the
ivory to its centromere 3.6 units. Four lines
of bees of diverse origin all showed a low
percentage of automictic or gynandromorphic types produced from queens in each
line. Various chance environmental conditions apparently influence the rate of production of these types. However, there were
some females in two of the lines with higher
frequencies inciicating that innate factors
may have significant eft'ects on their frequencies. Observations on Drosophila species — I), porthenogeuetica (Stalker, 1956b),
I). ))ia)njabeirai (INIurdy and Carson, 1959)
and D. nielanogaster (Goldschmidt, 1957) —
strongly suppt)i-t this A-iew.
The problem of sex determination in Hal)rol)i'acon presently stands as a function of
multii)le alleles in one locus, the heterozygotes being female and the azygotes and
homozygotes male. The occasionally diploid
males are regularly produced from fertilized
eggs in two allele crosses after inbreeding.
These diploid males are of low viability and
are nearly sterile. Their few daughters are
triploids, their sperm being dijiloid. Apis
probably follows the same scheme, as a
few cases of mosaics with diploid male tissue are known and close inbreeding results
in a sufficient number of deaths in the egg
to account for diploid males which might be
formed. ^lormoniella (Whiting, 1958), however, shows that this scheme for sex determination does not hold for all Hymenoptera.
In this form diploid males may occur
through some form of mutation. They may
then develop from unfertilized eggs laid by
triploid females. In contrast to Habrobracon the dijiloid males are highly viable and
fertile. Their sperm are diploid and their
numerous daughters triploid. Virgin triploid
females produce 6 kinds of males, 3 haploid
and 3 dij^loid. Similarly ]\Ielittobia has still
a different and as yet unexplained form of
sex determination. Haploid eggs develop
into males. After mating many eggs are laid
which develop into nearly 97+ per cent females. The method of reproduction is close
inbreeding but no dijiloid males or "bad"
eggs are formed. The prol)lem of the sex
determining mechanism remains open
(Schmieder and Whitiuii, 1947).
The silkworm, Bombyr mori, differs from Drosophila, Lymantria and the species thus far discussed in having a single region in the ^^' chromosome (Hasimoto, 1933; Tazima, 1941, 1952) occupied by a factor or factors of high female potency. The strong female potency has thus far been connnon to all races. The chromosome patterns of the sexes are like those of Abraxas and Lymantria: males ZZ + 2A and females ZwV 2A. The diploid chromosome number is 56 in both sexes. Extensive, well executed studies have revealed no W chruniosome loci for genes expressed as morphologic traits. From radiation-treated material it has been possible to pick up a translocation of chromosome II to the W chromosome as well as a cross-over from chromosome Z. This chromosome together with tests of hypoploids and hyperploids have materially aided in understanding how the normal chromosome complexes determine sex. The sex types resulting from different chromosome arrangements have been summarized by Yokoyama 1 1959) and are presented in Table 1.3.
Whenever the W chromosome was absent a male resulted. Extra Z or A chromosomes did not influence the result. Similarly
with a W chromosome in the fertilized egg
a female developed. Again extra Z or A
chromosomes did not influence the result.
A full Z chromosome was essential to survival. Hypoploids deficient for different
amounts of the Z chromosome in the presence of a normal W chromosome all died
without regard to the portion deleted. Hyperploids for the Z chromosome, on the other
hand, when accompanied with a W chromosome all lived and showed no abnormal sexual cliaracteristics. Parthenogenesis led to
the pioduction of both sexes, although the
males were more numerous than the females. Diploidy was necessary for the eml)ryo to go beyond the blastoderm stage.
Triploid and tetraploid cells were often
found. High temperature treatments led to
merogony (Hasimoto, 1929, 1934). The exceptional males were homozygous for a sexlinked recessive gene and were explained
by assuming that the egg nuclei were inactivated by the high temperature and
the exceptional males developed from the
union of two sperm nuclei. This conclusion
was supported by cytologically observed
l)olyspermy (Kawaguchi, 1928) and by
cytologic observation of the union of two
sperm nuclei by Sato ( 1942 ) . Binucleate eggs
were also believed to occur, which when
fertilized by different sperm may each construct half of the future body. This type of
mosaicism was influenced by heredity
iGoldschmidt and Katsuki, i927, 1928,
1931 ). Polar body fertilization was also believed to occur, one side of the embryo originating from the ordinary fertilized egg nucleus and the other side from the union of
TABLE 1.:^ Sex in Bombyx iitori (Summarized by T. Yokoyama, 1959.)
Sex
Chromosome Types and Numbers
W
z
A
Male
W
II.W.ZL
w w
WW WW
zz
zz
zzz
z
z
zz zzz zz zz
AA
Male
Male
Female
Female
Female
Female
Female
Female
AAA
AAA
AA
AA
AAA
AAAA
AAA
AAAA
nuclei of two of the polar bodies. Similarly, dispermic merogony was noted following the formation of one part of the body from the fertilization nucleus, the other part from the union of two sperm nuclei, the result being a gynandromorph or mosaic.
VI. Sex Determination in Dioecious Plants
A. Melandrium T Lychnis
Over the last 20 years studies on several species of dioecious plants have made notable advances in unclerstanding the mechanisms by which sex is determined.
Melandrium album has been shown to have the same chromosome arrangement as Drosophila. The male has an X and Y plus 22 autosomes, whereas the female has XX plus 22 autosomes. Sex-linked inheritance is known for genes borne in the X chromosomes as well as for genes born in the Y chromosome. The X and Y chromosomes are larger than any of the autosomes with the Y chromosome about 1.6 times that of the X in the materials studied by Warmke ( 1946) . Separate male and female plants are characteristic of the species. By use of colchicine and other methods, Warmke and Blakeslee (1939), Warmke (1946), and Westergaard (1940) have made various l^olyploid types from which they could derive other new X, Y and A chromosome combinations from which information was obtained on the location of the sex determining elements. The Y chromosome carries the male determining elements, the X chromosome the female determining elements. The guiding force of the elements in the Y chromosome during development is sufficient to override the female tendencies of several X chromosomes. Data derived by each of these investigators are shown in Table 1.4.
From these data Warmke (1946) concluded that the balances between the X and the Y chromosomes essentially determined sex with the autosomes of relatively little importance. Where no Y chromosomes were present but the numbers of X chromosomes ranged from 1 to 5, only females were observed, even though the autosomes varied in number from two to four sets. When a Y chromosome was present the individual was of the male type unless the Y was balanced by at least 3 X chromosomes when an occasional hermaphroditic blossom was formed.
TABLE L4
Numhers of X, Y, chromosomes and A, autosome sets
and the sex of the various Melandrium plants
(Data from H. E. Warmke, 1946; and
M. Westergaard, 1953.)
Ratio
Chromosome
Warmlce
Westergaard
Constitution
X/A
4A 5X
Female
1.3
4A 4X
Female
1.0
Female
4A 3X
Female
0.8
4A 2X
Female
0.5
3A 3X
Female
1.0
Female
3A 2X
Female
0.7
Female
2A 3X
Female
1.5
2A 2X
Female
1.0
Female
Bisexual*
X/Y
4A 4X Y
4.0
Male
4A 3X Y
Malet
3.0
Male
4A 2X Y
Malet
2.0
Male
4A X Y
Male
1.0
3A 3X Y
Malet
3.0
3A 2X Y
Malet
2.0
Male
3A X Y
Male
1.0
2A 2X Y
Malet
2.0
2A X Y
Male
1.0
Male
4A 4X YY
Malet
2.0
4A 3X YY
Male
1.5
4A 2X YY
Male
1.0
Male
2A X YY
Male
0.5
- Occasional staminate but never carpellate
blossom.
t Occasional licrina])hr(i(lit ic blossom.
When 4 X chromosomes were present together with a Y, the plants were hermaphroditic but occasionally had a male blossom. Two Y chromosomes almost doubled the male effect. Two Y chromosomes balanced 4 X chromosomes to give a majority of male plants. Only an occasional plant showed an hermaphroditic blossom. Autosomal sex effects, if present, were only observed when plants had 4 sets and 3 or 4 X chromosomes balanced by a Y chromosome. Warmke used the ratio of the numbers of X to Y chromosomes as a scale against which to measure clianges from complete male to hermaphroditic types. No mention is made of quantitative measures of the sex character changes with increasing X chromosome dosages. This is of interest since in many forms changes in chromosome balance are accompanied by changes of phenotype which are unrelated to sex. That such phenotypic changes do accompany changes in autosomal balance in Melandrium are proven, however, by further observations of Warmke in 4 trisomic types coming from crosses of triploids by diploids. Of 36 such trisomies analyzed, 5 or 6 of them were of different growth habits and morphologic types. These differences did not affect the sex patterns since all were females. Warmke and Blakeslee in 1940 observed an almost complete array of chromosome types from 25 to 48 in progeny derived from crosses of 3N x 3N, 4N X 3N, and 3N x 4N. Out of about 200 plants studied, only 4 were found to show indications of hermaphroditism. These types were 2XY and 3XY. As noted from the table, even the euploid plants would occasionally be expected to have an hermaphroditic blossom. Of the 200 plants, all with a Y (XY, 2XY, 3XY) were males and all plants without the Y (2X, 3X, 4X) were females. In an 8-year period up to 1946. Warmke was able to observe only one male trisomic. From these facts he concluded that the autosomes are unimportant in the sex determining mechanism utilized by this species. In their crosses they were unsuccessful in getting a 5XY plant, the point at which the female factor influence of the X chromosomes might be expected to nearly equal or slightly surpass that of the single Y. From the j^hysiologic side the obscrvation of Strassburger in 1900, as quoted by both Warmke and Westergaard, that the fungus Ustilago violacea when it infects Melandrium will cause diseased plants to produce mature blossoms with well developed stamens (filled with fungus spores) as well as fertile pistils, shows that these females have the potentialities of both male and female development. The case suggests that sex hormone-like substances may be produced by the fungus which acting on the developing Melandrium sex structures cause sex reversions. Should this be true, Melandrium cells would have a parallel with those of fish where sex hormones incor|)orated in the developing organism in sufficient quantities can cause the soma to develop a phenotype opposite to that expected of their chromosomal type. For other aspects see Burn's chapter and Young's chapter on hormones.
Westergaard's studies (1940) with European strains of Melandrium were in progress at the same time as those of Warmke and Blakeslee. In their broad aspects both sets of data are concordant in showing the l^rimary role of elements found within the Y chromosome in determining the male sex and of elements in the X chromosomes for the female sex. Examination of Table 1.4 shows that the strain used by Westergaard has a Y chromosome containing elements of greater male sex potentialities than the strain used by Warmke.
A similar difference appeared in the sex potencies of the autosomes of European strains. Instead of obtaining essentially only male and female plants in crosses involving aneuploid types, Westergaard obtained from 3N females (3A + 3X) x 3N males (3A + 2XY) 10 plants which were more or less hermaphroditic, 21 females, and 15 males. Studies of the offspring of these hermaphrodites through several generations showed that their sex expression required effects by both the X chromosomes and certain autosome combinations which under special conditions counterbalanced the female suppressor in the Y chromosome. Increasing the X chromosomes from 1 to 4 increased the hermaphrodites from to 100 per cent in the presence of a Y chromosome. However, in euploids these types would be all males. The significance of the autosomes is further shown by the fact that among 205 aneuploid 3XY plants, 72 were males and 133 were hermaphrodites.
As pointed out for Drosophila, quantitative studies on the effects of sex chromosomes and autosomes in Melandrium are
handicapped by not having a suitable scale
for the evaluation of the different sex types.
The data presented by Westergaard and by
Warmke make this difficulty become particularly evident. In the interest of quantizing
the X, Y, and A chromosome on sex the
author has assigned a value of 1 for the
male type, 3 for the female type, and 2
when the types are said to be hermaphroditic. When the types are mixed, as for example, in the data of Warmke where he says
a particular type is male with a few blossoms, the type is assigned a value of 1.05 or
1.10, depending on the numbers of these
blossoms. His bisexual type which comes
as a consequence of Y, 4X and 4A chromosome arrangement is given a value of 2, although possibly the value should be somewhat higher as it may well be that the fully
bisexuals are further along in the scale toward female development than the hermaphroditic types. The data are treated on the
additive scale both as between chromosomal
types and within chromosomal type. This is
apparently unfair if we examine the work
of Westergaard in which it looks as if particular autosomes rather than autosomes in
general make a contribution to sex determination. The results, when these methods are
used, are as follows:
Westergaard in Tables 1 to 5 of his 1948
paper gives information on sex types with
a determination of the numbers of their
different kinds of chromosomes. Analysis
of these data by least square methods shows
that the sex type may be predicted from
the equation
Sex type = - 1.37 Y + 0.10 X
+ 0.01 A + 2.34
This equation fits the data fairly well considering that the correlation between the variables and the sex type is 0.87. This analysis again shows that the Y chromosome has a strong effect toward maleness. The X chromosomes are next in importance with an effect of each X only about 1/13 that of the Y and in the direction of femaleness, the autosomes have one tenth the effect of the X chromosomes but they too have a composite effect toward femaleness. It is to be remembered that the Y chromosome variation is limited to 2 chromosomes whereas the X chromosomes may total 4, and the autosomes may range from 22 up to 42, so that the total effect of the autosomes is definitely more than their single effects. These data are for aneuploids. Examining Westergaard's data for 1953 for the euploids and assigning the value of 1.5 for the type observed when there was one Y chromosome, four X, and four sets of autosomes, we have the following equation:
Sex value = -1.29 Y + 0.10 X - 0.01
(autosome sets) + 2.53
In these data, as distinct from those above, the autosomes are treated as sets of autosomes since they are direct multiples of each other so the value of the individual autosome is but 1/11 that given in the equation.
This equation shows no pronounced difference from that when the aneuploids were utilized. The Y chromosomes have slightly less effect toward the male side. The X chromosomes have practically identical effects but there has been a shift in direction of the autosomal effects on sex, although the value is small. The constants are subject to fairly large variations arising through chance.
In Table 10 of Westergaard's 1948 paper
he presented data on the chromosome constitution and sex in the aneuploids which
carried a Y chromosome. These data are of
particular interest as the plants are counted
for the i)roi)ortions of those which are male
to those which are hermaphroditic. The
plants with a Y chromosome plus an X are
all males. Those which have either one, two
or three Y chi'omosomes balanced by two
X chromosomes have 89 per cent males.
The plants with three X chromosomes and
one oi- two Y's have 36 p(T cent males and
those which have one or two ^■ chroiiiosoincs
and four X chromosomes have no males.
The woik iiiv()l\-cd in getting these data is,
of course, large indeed and is definitely
handicapped by the diiiicuHies in obtaining certain types. Thus the XXYYY and the
4X + 2Y types depend only on one plant.
There are eight observations but the fitting
of the data for the X and Y constitutions
eliminates three degrees of freedom from
that number so that statistically the observations are few. The data do have the advantage that the sex differences can be
measured on an independent quantitative
scale. The equation coming from the results
is:
Percentage of males = 13.2 Y - 36.4 X
+ 134.4
These results show that the Y chromosome increases the proportion of males and the X chromosome increases the proportion of intersexes. The data are not comparable with those analyzed earlier as these data are describing simply the ratio between the males and intersexes, instead of the relations betw'een the males, intersexes, and females. The equation fits the observations rather well, as indicated by the fact that the correlation between the X's and Y's and the percentages of males is 0.98, but there are large uncertainties.
As a contrast to these data we have those presented by Warmke (1946) in his Tables 2 and 3. These data give the numbers of X and Y chromosomes found within the plants but not the numbers of autosomes, the autosomes being considered as 2, 3, and 4 genomes. Analyzing these data in the same manner as those of Westergaard's Table 1, we find that the Sex type = -1.05 Y + 0.22 X -0.04 A
-f2.25
As indicated for Westergaard's data, the A effect is now in terms of the diploid type e(iualing 2, the trijiloid 3, and the tetral)loid 4.
The Y chromosome has a i)ronounccd effect toward maleness, the effect l)eing someW'hat less in Warmke's data than that of Westergaard's. The X chromosomes on the other hand, have nearly twice the female infhience in Warmke's data that they do in the phiiits grown by Westergaartl. A difference in sign exists for the effect of the A rhi'oinosnnie genom(\s as well as a difference in [\\v (|uantitati\'e effect. Th(^ values for both sets of data are small and toward the male side. As Westergaard points out, the strains used by these investigators are of different geographic origins. The evolutionary history of the two strains may have a bearing on the lesser Y and greater X effects on the sex of the American types. Chromosome changes seem to have occurred in the strains before the studies of Warmke and Westergaard and will be discussed.
The location of the sex determiners has
been studied by both investigators utilizing
techniques by which the Y chromosome becomes broken at different places. These
breakages may occur naturally and at fairly
high rates in individuals which are Y + 2X
+ 2A. These facts suggest that the breakage of the Y chromosome occurs in meiosis
since the breakage comes in selfed individuals of highly inbred stocks where heterozygosity is not to be expected, in the Y
chromosome which has no homologue thus
does not synapse, and in the second meiotic
E. Sex in Man: Chromosomal Basis
A surprise even to its discoverers, Tjio and Levan (1956), came with the observation that the somatic number of chromosomes in cultures of human tissue was 46 rather than the previously supposed 48. Search for the true number has been going on for more than half a century. In early investigations the numbers reported varied widely. Difficulties of proper fixation and spreading of the chromosomes of human cells accounted for most of this variation and the numerous erroneous interpretations. Among the observations that of de Winiwarter (1912) was of particular interest in showing the chromosome number as 46 autosomes plus one sex chromosome with the Y being absent. This number was also found later by de Winiwarter and Oguma (1926). Observations by Painter (1921, 1923) showed 46 chromosomes plus an X and a Y, a total of 48. This number was subsequently reported by a series of able investigators, Evans and Swezy (1929), Minouchi and Ohta (1934), Shiwago and Andres (1932), Andres and Navashin (1936), Roller (1937), Hsu (1952), Mittwoch (1952), and Darlington and Haque (1955). As Tjio and Levan indicated, the acceptance of 48 as the correct number, with X and Y as the sex chromosome arrangement, was so general that when Drs. Eva Hanson-Melander and S. Kullander had earlier found 46 chromosomes in the liver cells of the material they were studying they temporarily gave up the study. In the few years since 1956, the acceptance of 46 chromosomes as the normal complement of man has become nearly universal. There are 22 paired autosomes plus the X and Y sex chromosomes.
The reasons which have warranted this
change of viewpoint are no doubt many,
but three improvements in technique are
certainly significant. The first came as a
consequence of simplifying the culture of
human somatic cells. The second followed
Hsu's (1952) recognition that pretreatment
of these cells before fixation with hypotonic
solutions tended to better spreads of the
chromosomes on the division plates when
subsefiuently stained by the squash techniciuo. Pretreatment of the cultures with colchicine made the studies more attractive
by increasing the numbers of usable cells
that were in the metaphase of cell division.
Ford and Hamerton (1956) in an independent investigation, closely following
that of Tjio and Levan, observed that the
human cell complement contained 46 chromosomes. They, too, agreed with Painter
and others that followed him that the male
was XY and the female XX in composition.
A flood of confirming evidence soon followed: Hsu, Pomerat and Moorhead (1957),
Bender (1957), Syverton (1957), Ford,
Jacobs and Lajtha (1958), Tjio and Puck
(1958), Puck (1958), Chu and Giles (1959),
and a number of others.
In most instances the results of the different investigators were surprisingly consistent in showing that the individual cell
chromosome counts nearly always totaled
46. This was no doubt due in part to the
desirability of single layers of somatic cells
for identifying and separating the different
chromosomes into distinct units. Chu and
Giles' results illustrate this consistency.
For 34 normal human subjects, including
29 American whites and 4 American Negroes, and one of unknown race, and regardless of sex, age, or tissue, the diploid
chromosome number of the somatic cells
was overwhelmingly 46. In only five individuals were other numbers observed in
isolated cells. Out of 620 counts, 611 had
46 chromosomes; two individuals, whose
majority of cells showed 46, had 3 cells with
45 chromosomes; three other individuals,
the majority of whose cells showed 46, had
6 cells with 47 chromosomes. Average cell
plates counted per individual was nearly 20.
The only recent observations at variance
with these results were those of Kodani
(1958) who studied spermatogonial and
first meiotic metaphases in the testes from
15 Japanese and 8 whites. In these studies
at least several good spermatogonial metaphases in which the chromosomes could be
counted accurately, and secondly at least
15 spermatocyte metaphases in which the
structure of individual chromosomes could
be observed clearly, were made on each
specimen. The numbers of cells studied in
metaphase were generally above these numbers, one reaching 60 metaphases. Some variation was noted within individuals. Among individuals, numbers of 46, 47, and 48 were
observed. Among 15 Japanese, 9 had 46, 1
had 47, and 5 had 48 chromosomes, whereas
among the whites 7 had 46, and 1 had 48.
Sixteen of the 23 individuals had 46 chromosomes. Karyotype analyses indicated
that the numerical variation was caused by
a small supernumerary chromosome. On the
basis of these observations it would appear
that individuals within races may vary in
chromosome number and yet be of normal
phenotype. However, in view of the extensive observations by others, it seems unlikely that the variation between individuals is as large as that indicated. It will
require much further study to establish any
other number than 46 as the normal karyotype of man. This is particularly true in
view of the work of Makino and Sasaki
(1959) and Alakino and Sasaki cited by
Ford (1960), in which they studied the human cell cultures of 39 Japanese and found
without exception 46 chromosomes, and the
earlier work of Ford and Hamerton (1956)
on spermatogonial material where they, too,
found 46 chromosomes in that tissue. The
best features of these human chromosome
studies will come in the identification of
the individual chromosomes making up the
human group. The chromosome pairs may
be ordered according to their lengths. The
longest chromosome is about 8 times the
length of the smallest. The chromosomes
may be classified according to their centromere positions. The chromosomes are said
by most observers to be fairly easily separated into 7 groups. Separation of the individual chromosome pairs from each other
and designation of the pairs so that they
can be identified by trained investigators
in all good chromosome preparations is not
possible according to some ciualified cytologists and admitted difficult by all students.
However, standardized reporting in the
rapidly growing advances in human cell
studies should refine observations, reduce
errors, and encourage better techniques.
With this in mind, 17 investigators working
in this field met in Denver in 1959 in what
has come to be called the "Denver conference" (Editorial, 1960). From an examination of the available evidence on chromosome morphologies an idiogram was set up
as a standard for the somatic chromosome complement of the normal human genome.
A reproduction of this standard is presented
in Figure 1.1, as kindly loaned by Dr. Theodore T. Puck for this purpose.
Fig. 1.1. Id
The autosomes were first ordered in relation to their size and such attributes as would help in their positive identification. Numbers were given to each chromosome as a means of permanent identification. Basically, identification is assisted by the ratio of the length of the long arm to that of the short arm; the centromeric index calculated from the ratio of the length of the shorter arm to the whole length of the chromosome ; and the presence or absence of satellites. Classification is assisted by dividing the chromosome pairs into seven groups. Groups 1-3. Large chromosomes with approximatel}^ median centromeres. The three chromosomes are readily distinguished from each other by size and centromere position. Group 4-6. Large chromosomes with submedian centromeres. The two chromosomes are difficult to distinguish, but chromosome 4 is slightly longer. Group 6-12. Medium sized chromosomes with submedian centromeres. The X chromosome resembles the longer chromosomes in this group, especially chromosome 6, from which it is difficult to distinguish. This large group is the one which presents major difficulty in identification of individual chromosomes. Group 13-15. INledium sized chromosomes with nearly terminal centromeres ("acrocentric" chromosomes). Chromosome 13 has a prominent satellite on the short arm. Chromosome 14 has a small satellite on the short arm. No satellite has been detected on chromosome 15. Group 16-18. Rather short chromosomes with approximately median (in chromosome 16) or sul>median centromeres. Group 19-20. Short chromosomes with approximately median centromeres. Group 21-22. Very short, acrocentric chromosomes. Chromosome 21 has a satellite on its short arm. The Y chromosome belongs to this group. Separations of the human chromosome pairs into the seven groups is not as difficult as designating the pairs within groups (Patau, 1960). The svstem is a notable advance in summarizing visually the current information in the hope that availability of such a standard will promote further refinements, lessen misclassification, and contribute to a better understanding of the problems by cytologists and other workers in the field.
1. Xuclear Chromatin, Sex Chromatin
Sexual dimorphism in nuclei of man (Barr, 1949-59) and certain other mammals may be detected by the observable presence of nuclear chromatin adherent to the inner surfaces of the nuclear membrane. The material is about 1 /x in diameter. It frequently can be resolved into two components of equal size. It has an affinity for basic dyes and is Feulgen and methyl green positive. Nuclear chromatin can be recognized in 60 to 80 per cent of the somatic nuclei of females and not more than 10 per cent of males. It is known to be identifiable in the females of man, monkey, cat, dog, mink, marten, ferret, raccoon, skunk, coyote, wolf, bear, fox, goat, deer, swine, cattle, and opossum, but is not easily usable for sex differentiation in rabbit and rodents because these forms have multiple large particles of chromatin in their nuclei. The tests can be made quickly and easily on skin biopsy material or oral smears. Extensive utilization of the presence or absence of nuclear chromatin in cell samples of man has been made for assigning the presumed genetic sex to individuals who are phenotypically deviates from normal sex types. (See also chapters by Hampson and Hampson, and by Money.) Numerous studies on normal individuals seem to support the test's high accuracy. However, in certain cases involving sexual modification, questions have arisen which are only now being resolved. In male pseudohermaphroditism, sex, determined by nuclear chromatin, is male, thus agreeing with the major aspects of the phenotype. For female pseudohermaphroditism, individuals with adrenal hyperplasia or those without adrenal hyperplasia give the female nuclear chromatin test. For cases listed as true hermaphrodites Grumbach and Barr (1958) list 6 of the male type and 19 of the female type. For the syndrome of gonadal dysgenesis they list 90 as male and 12 as female among the proved cases and 15 more as female among those that are suspected. In the syndrome of seminiferoustubule dysgenesis where there is tubular fibrosis, 9 are listed as male and 18 female. Where there is germinal aplasia, 15 are listed as male and 1 as female. The seeming difficulties in assigning a sex constitution to some of these types are now being dissipated through the study of the full chromosome complements which are responsible for these different disease conditions. As observations on different chromosome types have been extended, evidence has accumulated to show that the numbers of sex nuclear chromatins, for at least some of the nuclei making up the organism, often equals (n — 1) times the number of X chromosomes. The majority of male XY nuclei are chromatin negative as are most of the Turner XO type. Female nuclei XX have a single chromatin positive element as do the XXY and XXYY types. The XXX and XXXY have 14 and 40 per cent respectively with two Barr bodies in cases for which quantitative data are available. However, a child with 49 chromosomes, but whose cultured cell chromosomes appear as single heteropycnotic masses making identification of the individual chromosomes difficult, showed 50 per cent of the cell nuclei with three Barr elements (Fraccaro and Lindsten, 1960) . The chromosome constitution of these nuclei was interpreted as trisomic for 8, 11, and sex chromosomes. Sandberg, Crosswhite and Gordy (1960) report the case of a woman 21 years old having various somatic changes which does not fit this sequence. The chromosome number was 47 and the nuclei were considered trisomic for the sixth largest chromosome. Two chromatin positive bodies were ])rosent in the nuclei.
2. Chrotnosome Complement and Phenotyppe in Man
Experience of the past 50 years has emphasized that genes and trisomies or other types of aneuploid chromosome complexes may lead to the development of abnormal phenotypes expressing a variety of characteristics. Drosophila led the way in illustrating how the different gene or chromosome arrangements may affect sex expression. Investigations of human abnormal types, particularly those with altered sex differentiation, have reccntly .^liown that man follow.- other species in this regard. The Y carries highly potent male influencing factors. Gene differences often lead to characteristic phenotypes of unique form.
3. Testicular Feminization
The testicular feminization syndrome illustrates one of these types. As described by Jacobs, Baikie, Court Brown, Forrest, Roy, Stewart and Lennox (1959), "In complete expression of this syndrome the external genitalia are female, pubic and axillary hair are absent or scanty, the habitus at puberty is typically female, and there is primary amenorrhoea. The testes can be found either within the abdomen, or in the inguinal canals, or in the labia majora, and as a rule the vagina is incompletely developed. An epididymis and vas deferens are commonly present on both sides, and there may be a rudimentary uterus and Fallopian tubes. The condition is familial and is transmitted through the maternal line." A sex-linked recessive, a sex-limited dominant, and chromosome irregularities of the affected persons have been postulated as mechanisms causing the apparent inheritance of this condition. Chromosome examinations of the cells of affected persons have shown 46 as the total number and X and Y as the sex complement. The karyotype analysis agrees with the Barr nuclear chromatin test in that the cells are chromatin-negative but both are at variance with the sex phenotypes in the sense that aside from suppressed testes the patients are so completely female. Genetically, Stewart (1959) has described two color-blind patients with the testicular feminization syndrome in the first five patients he reported. The limited data from these cases suggest that the genie basis for this condition is either independent or but loosely linked with color blindness. This evidence does not exclude sex-linkage but does make it less probable. The third hypothesis of autosomal inheritance may take one of several forms. A recessive gene which affects only the male phenotypes when in homozygous condition is apparently untenable because the matings from which these individuals come are of the outbreeding type and the ratios apparently do not differ from the one-to-one ratio expected of a heterozygous dominant instead of that required for an autosomal recessive. The hypothesis advanced by Witschi, Nelson and Segal (1957), that the presence of an autosomal gene in the mother converts all her male offspring into phenotypes of more or less female constitution, in a manner comparable to that of the Ne gene in Drosophila (Gowen and Nelson, 1942) which causes the elimination of all the female type zygotes, is also made unlikely by the ratios of normal to testicular feminization phenotypes observed in the progenies of these affected mothers. The evidence favors a simple autosomal dominant, acting only in the male zygotes and perhaps balanced by some genes of the X chromosome, which have sufficient influence on the developing male zygote to guide it toward an intermediate to nearly female phenotype. The observations of Puck, Robinson and Tjio ( 1960) indicate that the action of a gene for this condition may not be entirely absent in the female, because in heterozygous condition in an XX individual it seemed to delay menarche as much as 8 years. If this delay be diagnostic for the heterozygote, it will further assist in the genetic analysis of this problem. Evidence on this point should be a part of the genetic studies.
Cases closely similar to those described
by Jacobs, Baikie, Court Brown, Forrest,
Roy, Stewart and Lennox (1959) are
presented by Sternberg and Kloepfer
(1960). The patients show no trace of masculinity. They are remarkably uniform in
anatomic expression. Except for failure to
menstruate due to lack of uteri they undergo normal female puberty. Cryptorchid
testes, usually intra-abdominal, if removed
precipitate menopause symptoms. Four unrelated cases were found in this one study
with 7 additional cases traced through pedigree information. A total of 11 affected individuals was found in 6 sibships having
26 siblings of whom 5 were normal males.
In each kindred the inheritance was compatible with that of a sex-linkecl recessive
gene. A chromosomal study of a thyroid
tissue culture from one case revealed 46
chromosomes with normal XY male configuration. The individuals observed were
designated as simulant females."
4. Superfemale
The human superfemale has been recognized by Jacobs, Baikie, Court Brown, MacGregor, Maclean and Harnden (1959) in a girl of medium height and weight, breasts underdeveloped, genitalia infantile, vagina small, and uterocervical canal 6 cm. in length. Ovaries appeared postmenopausal with normal stroma, and as indicated by a biopsy specimen, deficient in follicle formation. Menstruation was thought to have begun at age 14, but was irregular, occurring every 3 to 4 months and lasting 3 days. The last spontaneous menstruation was at 19. Estrogen therapy caused some development of the breasts and external genitalia, vagina, and uterus with slight uterine bleeding. The patient's parents were above 40 years of age, mother 41, at time of her daughter's birth.
Examination of sternal marrow cultures
showed 47 chromosomes in over 80 per cent
of the cells examined. The extra chromosome was the X, the chromosomal type
being XXX plus 22 pairs of autosomes.
Buccal smears showed 47 per cent of nuclei
contained a single chromatin body and 14
per cent contained 2 chromatin bodies as
expected of a multiple XX or XXX genotype. In comparison, 25 smears from 20 normal women had 36 to 51 per cent chromatin
positive cells but none of these contained
2 chromatin bodies. Two chromatin bodies
were seen in some cells of the ovarian stromal tissue. The patient showed a lack of
vigor, mentally was subnormal, was underdeveloped rather than overly developed in
the phenotypic sexual characteristics. Examination of the patient's mother showed
her to be XX plus 22 pairs of autosomes,
the normal 46 chromosomes.
Other cases show that types with XXX
plus 22 pairs of autosomes are of female
l)henotype but may vary in fertility and
development of the secondary sexual characteristics from nonfunctional to functional
females bearing children ( Stewart and Sanderson, 1960; Eraser, Campbell, MacGillivray, Boyd and Lennox, 1960). The triplo
X condition in man has a greater range of development and fertility than in Drosophila.
In man ovaries may develop spontaneously.
In Drosophila they require transplantation to a diploid female host where they may attach to the oviducts and release eggs for
fertilization (Beadle and Ephrussi, 1937).
These cases present confirmation of two
facts already mentioned for Drosophila.
They show that when the X chromosome
has primarily sex determining genes, the
organism generally becomes unbalanced
when 3 of these X chromosomes are matched
against two sets of autosomes. The resulting phenotypes are female but relatively
undeveloped rather than overdeveloped.
The second is that the connotations evoked
by the prefix "super" are by no means applicable to this human type or to the Drosophila type.
The characteristics of the patient also
suggest that the autosomes may be carrying
sex genes opposing those of female tendencies as observed in both Drosophila and
Rumex genie imbalance.
5. Klinefelter Syndrome
In the Klinefelter syndrome there is male differentiation of the reproductive tracts with small firm descended testes. Meiotic or mitotic divisions are rare, sperm are ordinarily not found in the semen. The type is eunuchoid in appearance with gynecomastia, high-pitched voice, and sparse facial hair growth. Seminiferous tubules showing an increased number of interstitial cells are atrophic and hyalinized. Urinary excretion of pituitary gonadotrophins is generally increased, whereas the level of 17-ketosteroids may be decreased. The nuclear chromatin is typically female. Of the dozen or more cases studied (Jacobs and Strong, 1959; Ford, Jones, Miller, Mittwoch, Penrose, Ridler and Sha])iro, 1959; Bergman and Reitalu quoted by Ford, 1960), only one, having but 5 metaphase figures, had less than 47 chromosomes in the somatic cells and XXY sex chromosomes. That case was thought to have typical female chromosomes XX + 22 AA. Two other cases were of particular interest as indicating further chromosome aberration. Ford, Jones, Miller, Mittwoch, Penrose, Ridler and Shapiro (1959) studied one patient who displayed both the Klinefelter and Mongoloid syndromes. The chromosome number was 48, the sex chromosomes being XXY and the 48tli chromosoinc being small acrocentric.
This individual had evidently developed
from an egg carrying 2 chromosomal aberrations, one for the sex chromosomes and the
second for one of the autosomes. The other
case, Bergman and Reitalu as cited by
Ford (1960), had 30 per cent of its cells
with an additional acrocentric chromosome
which had no close counterpart in the normal set.
Data where the Klinefelter syndrome occurs in families showing color blindness
(Polani, Bishop, Ferguson-Smith, Lennox,
Stewart and Prader, 1958; Nowakowski,
Lenz and Parada, 1959; and Stern, 1959a)
further test the XXY relationship and give
information on the possible position of the
color blindness locus with reference to the
kinetochore. Polani, Bishop, FergusonSmith, Lennox, Stewart and Prader (1958)
tested 72 sex chromatin-positive Klinefelter
patients for their color vision and found
that none was affected by red-green color
blindness. Nowakowski, Lenz and Parada
( 1959) tested 34 cases and detected 3 affected persons, 2 of whom were deuteranomalous and one protanopic. Stern (1959a)
l^oints out that these cases and their ratios
are compatible with the interpretation of
the Klinefelter syndrome as XXY. One of
the deuteranomalous cases had a deuteranomalous mother and a father with normal
color vision. This case could have originated
from a nondisjunctional egg carrying 2
maternal X chromosomes fertilized by a
sperm carrying a Y chromosome. The other
two cases had normal fathers with heterozygous mothers. There are several explanations by which the color-blind Klinefelter
progenies could be obtained. The heterozygotes might manifest the color-blind condition. The second hypothesis, which is
favored, is that of crossing over between the
kinetochore and the color-blind locus at the
first meiotic division to form eggs each
carrying 2 X chromosomes, one homozygous
for color blindness, and the other for normal
vision. An equational nondisjunction would
form eggs homozygous for color blindness
which on fertilization by the Y chromosomes of the male would give the necessary
XXY constitution for the color-blind male
which is Klinefelter in phenotype. A third
possibiHty is that these exceptions may
arise without crossing over as the result of nondisjunction at the second meiotic division.
If the hypothesis of crossing over is accepted, the color-blind locus separates freely
from its kinetochore and would suggest that
the position of the locus is at some distance
from the kinetochore of the X chromosome.
A disturbed balance between the X and
the Y chromosomes alters the sexual type.
A single Y chromosome, contributing factors important to male development, is able
to alter the effects of two sets of female
influencing X chromosomes. Yet two Y
chromosomes in a complex of XXYY plus
44 autosomes seem to have little or no more
influence than one Y (Muldal and Ockey,
1960). The locations of the sex-influencing
genes in man are thus more like those of the
plant Melandrium than of Drosophila in
which the male-determining factors occur
in the autosomes. The relative potencies of
the male sex factors compared with those of
the female, however, are much less than
those in Melandrium.
6. Turner Syndrome
Turner's syndrome or ovarian agenesis further substantiates the female influence of the X chromosomes. The cases occur as the developmental expression of accidents in the meiotic or mitotic divisions of the chromosomes. These accidents lead to adults unbalanced for the female tendencies of the X chromosome. The gonads consist of connective tissue. The rest of the reproductive tract is female. Growth stimuli of puberty are lacking, resulting in greatly reduced female secondary sexual development. Patients are noticeably short and may be abnormal in bone growth. In its more extreme form, designated as Turner's syndrome, the individuals may show skin folds over the neck, congenital heart disease, and subnormal intellect, as well as other metabolic conditions. Earlier work (Barr, 1959; Ford, Jones, Polani, de Almeida and Briggs, 1959) shows that 80 per cent of the nuclear chromatin patterns are of the male type. Evidence from families having both this condition and color blindness suggested that at least some of the Turner cases would be found to have 45 chromosomes, the sex chromosome being a lone X (Polani, Lessof and Bishop, 1956). Work of Ford, Jones, Polani, de Almeida and Briggs, (1959) has confirmed this hypothesis and added the fact that some of these individuals are also mosaics of cells having 45 and 46 chromosomes. The 45 chromosome cells had but one X, whereas the 46 had two X's. This finding may explain the female-chromatin cell type observed in about 20 per cent of the cases having the Turner syndrome. Such mosaics of different chromosome cell types could also be significant in reducing the severity of the Turner syndrome and in increasing the range of symptoms which characterize this chromosome-caused disease as contrasted with those characterizing Turner's disease. Further cases observed in other investigations, Fraccaro, Kaijser and Lindsten (1959), Tjio, Puck and Robinson (1959), Harnden, and Jacobs and Stewart cited by Ford (1960) have all shown 45 chromosome cells and a single X chromosome. As with the XXX plus 44 autosome super females, the Turner type, X plus 44 autosomes, also shows a rather wide range in development from sterility with extensive detrimental secondary effects to nearly normal in all respects. Bahner, Schwarz, Harnden, Jacobs, Hienz and Walter (1960) report a case which gave birth to a normal boy. Other cases have been described (Hoffenberg, Jackson and jVIuller, 1957; Stewart, 19601 in which menstruation was established over a period of years. The XO type in man and Melandrium is morphologically female. In Drosophila on the other hand, the XO type is phenotypically nearly a perfect male. It is further to be noted that the X chromosome of Drosophila appears to have a less pronounced female bias than that of man when balanced against its associated autosomes, inasmuch as the XO + 2A type in Drosophila is male as contrasted with the XO + 2A type in man which is female. At the same time it seems that the autosomes in the human may be influential in that the female gonadal development is suppressed instead of going to completion as it does in the XX type.
7. Hermaphrodites
Hermaphroditic phenotypes in man, to the number of at least 74 (Overzier, 1955), have been observed and recorded since 1900. Types with a urogenital sinus predominated. The uteri were absent in some cases, even when complete external female genitalia were present. Ovotestes were found on the right side of the body in over half the cases; separate left ovaries or testes were about equally frequent ; in three cases separate testis and ovary were indicated. The left side of the body showed a different distribution of gonad types; about onefourth had ovotestes, another fourth ovaries, and one-twelfth testes. Unilateral distribution of gonad types was most frequent. The presence or absence of the prostate seemed to have significance because it is sometimes absent in purely female types. In recent literature similar cases have been called true hermaphrodites. This is an exaggeration in terms of long established practice in plants and animals where true hermaphroditism includes fully functioning gametes of each sex.
Hungerford, Donnelly, Nowell and Beck
(1959» have reported on a case of a Negro
in which the culture cells had the chromosome complement of a normal female 46,
with XX sex chromosomes. Unfortunately,
the possibility that this case may be a chromosome mosaic was not tested by karyotype samples from several parts of the body.
Harnden and Armstrong (1959) established separate skin cultures from both sides
of the body of another hermaphroditic type.
The majority of the cells were apparently of
XX constitution with a total of 46 chromosomes. However, in one of the 4 cultures
established, some 7 per cent of cells had an
abnormal chromosome present, suggesting
that the case might involve a reciprocal
translocation between chromosomes 3 and
4 when the chromosomes were ordered according to size. All the other cell nuclei were
normal. The fact that the majority of the
cells in these two cases were XX and with
46 chromosomes seems to predicate against
the view that either changes in chromosome
number or structure of the fertilized egg are
necessary for the initiation of hermaplu'odites.
Ferguson-Smith (1960) describes two
cases of gynandromorphic type in which the
reproductive organs on the left side were
female and on the right side were male. The
recognizable organs were Fallopian tube,
ovary with primordial follicles only, immature uterus in one case, none in the other,
rudimentary prostate, small testis and epididymis, vas deferens, bifid scrotum, phallus, perineal urethra, pubic and axillary
hair, breasts enlarging at 14 years. Testicular development with hyperplasia of Leydig
cells, germinal aplasia, and hyalinization of
the tubules was suggestive of the Klinefelter
syndrome. Nuclear-chromatin was positive
in both cases. Modal chromosome number
was 46. The sex chromosomes were interpreted as XX. The 119 cell counts on one
patient showed a rather wide range; 7 per
cent had 44 chromosomes, 13 per cent had
45, 62 per cent had 46, and 18 per cent had
47 chromosomes. The extra chromosome
within the cells containing 47 chromosomes
was of medium size with submedian kinetochore as generally observed for chromosomes of group 3.
It is surprising that the male differentiation in these four and other hermaphroditic
cases (Table 1.5) is as complete as it is.
Other observations show that the Y chromosome contains factors of strong male
potency, yet in its absence the hermaphrodites develop an easily recognized male
system. It is not complete but the degree of
gonadal differentiation is as great as that
observed in the XXY 4- 2A Klinefelter
types. The bilateral sex differentiation in
hermaphrodites would seem to require other
conditions than those heretofore considered.
Another case of hermaphroditism is that
presented by Hirschhorn, Decker and
Cooper (1960). The patient's j:)henotype
was intersexual with phallus, hypospadias,
vagina, uterus. Fallopian tubes, two slightly
differentiated gonads in the position of ovaries. The child was 4 months old. Culture
of bone marrow cells showed that the individual was a mosaic of two types. About
60 per cent of the cells had 45 chromosomes
of XO karyotype, and 40 per cent had 46
chromosomes with a karyotA'^pe XY. The
Y chromosome when present was larger
than Y chromosomes of normal individuals.
The change in size may be related to the
association of the XO and XY cells and
be similar etiologically to the case discussed
by yivtz (1959) in Sciara triploids.
There are mosaics in Drosophila formed
from the loss by the female in some cells of
one of lioi- X chromosomes, as for instance in ring chromosome types, which may display primary and secondary hermaphroditic
development. For this to happen the altered
nuclei apparently find their way into the
region of the egg cytoplasm which is to
differentiate into the reproductive tract. As
seen in the adults, organ tissue of one chromosome type is cell for cell sharply differentiated from that of the other chromosome
type with regard to sex. These observations
indicate that for these mosaics the basic
chromosome structure of the cell itself
determines its development. In fact most
mosaics of this species show this cell-restricted differentiation. Several problems
arise when these well tested observations
are considered in comparison with those
now arising in the chromosome mosaics of
the sex types in man. It would seem unlikely
that the bone marrow cells or for that
matter any somatic cells not a part of the
reproductive tract would operate to modify
the adult sex or a part thereof. Rather the
developmental secjuence should start from
cell differences within the early developing
reproductive tract. Circulatory cells or substances would be of dubious direct significance from another viewpoint. All cells of
the body would ultimately be about equally
affected by any cells or elements circulating
in the blood. With strong male elements
and strong female elements the result expected would be a reduction in sex development of either sex instead of the sharply
differentiated organ systems which are observed. This raises the question, are the
chromosomally differentiated cell sex mosaics primary to or secondarily derived
from the tissues of the ultimate hermaphrodites? Study of the cell structure of the sex
organs themselves as well as much other
information will be necessary to clear up
this problem.
There are, however, other types of controlled sex development, as by various
genes, which lead to the presence of both
male and female sexual systems. Genes
for these phenotypes are relatively rare, but
once found are transmitted as commonly
expected. The inherited hermaphroditic
cases in Drosophila are certainly relevant
to the testicular feminization syndrome in
man. Are they equally pertinent to the
highly sporadic hermaphroditic forms just considered for man? If so, they indicate a
genie basis for these types which would
probably be beyond the range of the microscope to detect. The low frequency of true
hermaphrodites in the human, together with
lack of information on possible inheritance
mitigates against the genie explanation ; although genie predisposition acting in conjunction with rare environmental events as
occurs in our Balb/Gw mice (Hollander,
Go wen and Stadler, 1956 ) could explain the
rare hermaphrodites observed in that particular line of mice and a limited number of
its descendants.
8. XX XY + U Autosome Type
The XXXY -I- 44 autosome type in the human has been studied by Ferguson-Smith, Johnson and Handmaker (1960) and Ferguson-Smith, Johnston and Weinberg (1960). The two cases described were characterized by primary amentia, microorchidism and by two sex chromatin bodies in intermitotic nuclei. The patients were similar in having disproportionately long legs; facial, axillary, and abdominal hair scant; pubic hair present; penes and scrota medium to well developed ; small testes and prostates; vasa deferentia and epididymides normally developed on both sides of the body and no abnormally developed Miillerian derivatives. Testes findings were like those in Klinefelter cases with chromatinpositive nuclei, small testes with nearly complete atrophy, and hyalinization of seminiferous tubules and islands of abnormal and pigmented Leydig cells in the hyalinized areas. The few seminiferous tubules present were lined with Sertoli cells but were without germinal cells. Nuclear chromatin was of female type. About twofifths of the nuclei had double and twofifths single sex chromatin. The modal chromosome count for bone marrow cells was 48, 75 per cent of the cells having this number. Chromosome counts spread from 45 to 49. This type, XXXY plus 44 autosomes, may be looked upon as a superfemale plus a Y or a Klinefelter plus an X chromosome. In either case the male potency of the genes in the Y chromosome is able to dominate the female tendencies of XXX to develop nearly complete male phenotypes.
TABLE 1.5
Chromosome kinds and numbers for different recognized sex types in man
External Type
Male
Female
Female (rare hemophilic)
Eunuchoid female Female
Female Female Female
Female Female
Male . Male. Male.
Male. Male.
Male . . . Female. Female .
Female . Female .
Male. Male.
Hermaphrodite
Hermaphrodite. Intersex
Numbers of Chromosomes
chromosoil rearrange
1 + X
fragment
1 2 2
2 2 Interpreted a.s XX trisomic for Sand 11 or as XXXX 4X + Yl
3X+ Yj 1 3 3
2l /mosaic
Missing or Extra Chromosomes
? Chr mosome 3
orT(X;A)
Small autosome
? Large chromosome or
T(X;A)
X or Y
(X or Y) +21
46,47,48 cliromosomes X, Y X or Y -fA set
Sex Chromatm
Negative Positive
Negative Negative Positive
Negative Positive Negative
Negative
Negative Negative
Negative ±
Negative Positive Positive
Triple positi
Negative Double posit Double posit
Negative
Double posit Negative
Negative Negative
Designating Term
Normal male Normal female
Pure gonadal dysgenesis CJonadal dysgenesis
Testicular feiuinizati
Turner
Turner type female
Tinner? gave birth to boy Turner
Turner
Klinefelter
Klinefelter mongoloid
Klinefelter Klinefelter
Klinefelter
Klinefelter Sujierfemale
Superfemale gave birtli to children
Testicular deficiency
Precocious puberty
Triploid
Hermaphrodite or intersex dei)cnding on definition
Investigators*
2 3, 4, 5,
5, 12, 13, 14, 40, 41
41
16, 17, 18, 40, 41
19
40
23, 40
24, 25
41
26
29, 30, 31,. 32, 33 35, 39, 40
34
41
Tjio and Levan, 1956.
Nilsson, Bergman, Reitalu and Waldenstrom, 1959. Harnden and Stewart, 1959. Stewart, 1960b. Stewart, 1960a.
Elliott, Sandler and Rabinowitz, 1959.
Jacobs, Baikie, Court Brown, Forrest, Roy, Stewart and Lennox, 1959. Stewart, 1959
Lubs, Vilar and Bergenstal, 1959. Sternberg and Kloepfer, 1960. Puck, Robinson and Tjio, 1960. Ford, Jones, Polani, de Almeida and Briggs, 1959. Fraccaro, Kaijser and Lindsten, 1959. Fraccaro, Kaijser and Lindsten, 1960a.
15. Bahner, Schwarz, Harnden, Jacobs, Hienz and Walter, 1960.
16. Jacobs and Strong, 1959.
17. Bergman, Reitalu, Nowakowski and Lenz, 1960.
18. Nelson, Ferrari and Bottura, 1960.
19. Ford, Jones, Miller, Mittwoch, Penrose, Hidlor and Shapiro, 1959.
20. Ford, Polani, Briggs and Bishor), 1959.
21. Crooke and Hayward, 1960.
22. Muldal and Ockey, 1960.
23. Jacobs, Baikie, Court Broun, MacCJregor, Maclean and Harnden, 1959.
24. Eraser, Campbell, .MacCiillivray, Boyd and Lennox, 1960.
25. Stewart and Sanderson, 1960.
26. Jacobs, Harnden, Court Brown, Cohl.stein, Close, MacGregor, Maclean and Strong, 1960.
27. Ferguson-Smith, Johnston and Handiiiaker, 196(
28. Book and Santesson, 1960.
29. Harnden and Armstrong, 1959.
30. Hungerford, Donnelly, Nowell and Beck, 1959.
31. Ferguson-Smith, Johnston and Weinberg, 1960.
32. deAssis, Epps, Bottura and Ferrari, 1960.
33. Gordon, O'Gorman, Dewhurst and Blank, 1960
34. Hirschhorn, Decker and Cooper, 1960.
35. Sasaki and Makino, 1960.
36. Bloise, Bottura, deAssis, and Ferrari, 1960,
37. Fraccaro, Kaijser and Lindsten, 1960c.
37a. Fraccaro and Lindsten, 1960.
38. Fraccaro, Ikkos, Lindsten, Luft and Kaijser, 1960.
39. Harnden, 1960.
40. Ferguson-Smith and Johnston, 1960.
41. Sandberg, Koepf, Crosswhite and Hauschka, 1960.
42. Hayward, 1960.
Both cases had severe mental defects but it should be remembered that they were sought in institutions for which this is a criterion of admittance. Their mental ability was distinctly less than that of Klinefelter XXY cases which have come under study. The pattern of the XXXY effects on the reproductive tract, however, was comparable with that observed in the XXY genotypes. The effects of one Y chromosome were balanced by either two or three X chromosomes to give nearly equal phenotypic effects.
9. XXY + 66 Autosome Type
XXY + 66 autosome type was established by Book and Santesson (1960) for an infant boy having several somatic anomalies which may or may not be relevant to the sex type. Externally the genitalia were normal for a male of his age, penis and scrotum with testes present in the scrotum. Again the Y chromosome demonstrates its male potencies over two X's even in the presence of three sets of autosomes. The case is of particular significance since further development may indicate what male potencies an extra set of autosomes may possess.
10. Summary of Types
Other types of sex modifying chromosomal combinations and their contained genes have been observed particularly as mosaics or as chromosomal fragments added or substracted from the normal genomes. No doubt other types will be discovered during the mushroom growth of this period. Time can only test the soundness of the observations for the field of human chromosomal genetics and cytology is difficult at best requiring special aptitudes and experience. Mistakes, no doubt, will be made. The status of the subject is summarized in Table 1.5.
Other cases not related to sex or only secondarily so were scrutinized during the course of these studies. The information gained from them is valuable as it strengthens our respect for the mechanisms involved. The sex types which are dependent on loss or gain of the X and/or Y chromosomes belong to the larger category of monosomies or trisomies. Numbers of autosomal monosomic and trisomic syndromes have also been identified in the course of these investigations. Similarly, not all cases that have been studied have turned out to be associated with chromosomal changes. This in itself is important since it lends confidence in those that have, as well as redirects research effort toward the search for other causes than chromosomal misbehavior. The first trisomic in man was identified through the study of Mongolism. The condition affects a number of primary characteristics but not those of sex, for males and females occur in about equal numbers. The broad spectrum of these effects points to a loss of balance for an equally extensive group of genes in the two sexes. The common association of characteristics making up these Mongoloids, together with their sporadic appearance and their change in frequency with maternal age, all suggest the findings which Lejeune, Gautier and Turpin (1959a, b) and Lejeune, Turi)in and Gautier (1959a, b) were able to demonstrate so successfully. They established that the tissue culture cells of Mongoloid imbeciles had 47 chromosomes and that the extra chromosome was in the small acrocentric group. Lejeune, Gautier and Turpin (1959a, b) have now confirmed these observations on not less than nine cases. Jacobs, Baikie, Court Brown and Strong (1959), Book, Fraccaro and Lindsten (1959) and Fraccaro (cited by Ford, 1960) as well as later observers have substantiated the results on more than ten other cases. The well known maternal age effect, whereby women over 40 have a chance of having IMongoloid offspring 10 to 40 times as frequently as those of the younger ages, would seem to point to non-disjunction in oogenesis as the most important cause of this condition. Some women who have had previous JMongoloid progeny have an increased risk of having others. This is an important consideration in that genetic factors may materially assist in bringing about nondisjunction in man as they are known to do in Drosophila (Gowen and Gowen, 1922; Gowen, 1928). The products of the nondisjunctions approach those expected on random distribution of the chromosomes (Gowen, 1933) so that occasionally more than one type of chromosome disjunction will appear in a given individual. Such a case is that illustrated by Ford, Jones, Miller, Mittwoch, Penrose, R idler and Shapiro (1959) in which the nondisjunctional type included not only that for the chromosome important to Mongolism but also the sex chromosomes significant in determining the Klinefelter condition. This individual showed 48 chromosomes, 22 pairs of normal autosomes, 3 sex chromosomes XXY, and a small acrocentric chromosome matching a pair of chromosomes, the 21st, within the smallest chromosomes of the human idiogram. The analysis of Mongolism showed the way for the separation of the various human sex types through chromosome analyses.
Chromosome translocations furnish another means of establishing an anomaly
that may then continue on an hereditary
basis as either the male or female may
transmit the rearranged chromosomes. Polani, Briggs, Ford, Clarke and Berg (1960),
Fraccaro, Kaijser and Lindsten (1960b),
Penrose, Ellis and Delhanty (1960) and
Carter, Hamerton, Polani, Gunalp and
Weller (1960) have studied Mongoloid
cases which they interpreted in this manner.
In some cases the rearranged chromosomes
have been transmitted for three generations.
Several of the translocations were considered to include chromosomes 15 and 21.
Another trisomic autosomal type was rc])ortcd by Patau, Smith, Therman, Inhorn
and Wagner (1960). The patient was female and had 47 chromosomes. The extra
chromosome was a medium-sized acrocentric autosome belonging to the D group.
Despite extensive malformations affecting
several organs the patient lived more than
a year. Another female iiortraying the same syndrome has since been found, so other
cases may be expected. Among the characteristics are mental retardation, minor motor seizures, deafness, apparent micro or
anophthalmia, horizontal palmar creases,
trigger thumbs, Polydactyly, cleft i)alate,
and hemangiomata.
The third trisomic type was also reported
by Patau, Smith, Therman, Inhorn and
Wagner (1960). Six individuals have been
observed. The characters affected are mental retardation, hypertonicity (5 patients),
small mandible, malformed ears, flexion of
fingers, index finger overlaps third, big toe
dorsiflexed (at least 4), hernia and/or diaphragm eventration, heart anomaly (at
least 4), and renal anomaly (3). The sexes
were two males and four females. The extra chromosome was in the E group and was
diagnosed as number 18. Edwards, Harnden,
Cameron, Crosse and Wolff (1960) have
described a similar case but they consider
the trisomic to be number 17. Ultimate comparisons of these types no doubt will decide
if this is a 4th trisomic or if all the cases
belong in the same group.
The Sturge-Weber syndrome apparently
is caused by another trisomic. Locomotor
and mental abilities are retarded. Hayward
and Bower (1960) interpret the 3 chromosomes responsible as the smallest autosomes,
number 22, of the human group.
Trisomic frequencies should be matched
by equal numbers of monosomies. Turpin,
Lejeune, Lafourcade and Gautier (1959)
have reported polydysspondylism in a child
with low intelligence, dwarfing, and multil)le malformations of spine and sella turcica.
The somatic cell chromosome count was
only 45 but one of the smallest acrocentric
chromosomes appeared to have been translocated, the greatest part of this chromosome being observed on the short arm of one
of the 3 longer acrocentric chromosomes.
Th(> condition appears to be unique and not
likely to be found in other unrelated famiVws. However, the phenotyj^ic effects were
so severe that all members of the proband's
family would seemingly be worthy of careful sur\'('y for their chromosome characteristics.
The comi)lex pattern of multiple anomalies renders each syndrome distinct from the othei's. Chromosome losses or gains from the normal diploid would be expected to lead to the complex changes. Mongolism is influenced by age of the mother and probably to some extent by her inheritance. It is to be expected that the other trisomies may show parallel relations. Other trisomies may be expected although, as the chromosomes increase in size, a group of them will have less opportunity to survive because of loss of balance with the rest of the diploid set. Thus far most of these conditions affect the sex phenotypes. This is in accord with the results in Drosophila. Changes in the balance of the X chromosomes are less often lethal than the gain or loss of an autosome. Other animals show like effects. In plants, loss or gain of a chromosome, although generally detrimental, often causes less severe restrictions on life. Harmful effects are observed but do not cause early deaths. This may be because many aneuploids are within what are presumably polyploid plant species.
Ford (1960) has collected the data on 13 different phenotypes that could come under suspicion of chromosomal etiology as examined by a number of workers. Careful cytologic examination of patients suffering from one or another of these diseases has shown that the idiograms were normal in both number and structure of the chromosomes. The disease conditions were: acrocephalosyndactyly, arachnodactyly
(Marfan's syndrome), chondrodystrophy, Crouzon's disease, epiloia, gargoylism, Gaucher's disease, hypopituitary dwarfism, juvenile amaurotic idiocy, Laurence-MoonBiedl syndrome. Little's disease, osteogenesis imperfecta, phenylketonuria, and anencephalic types. To this list Sandberg, Koepf , Crosswhite and Hauschka (1960) have now been added neurofibromatosis, Lowe's syndrome, and pseudohypoparathyroidism.
F. Sex Ratio in Man
Sex ratio studies on human and other animal populations have always been large in volume. The period since 1938 is no exception. Geissler's (1889) data on family sex ratios, containing more than four million births, have been reviewed and questions raised by several later analysts. Edwards (1958) has reanalyzed the clata from this population and considered these points and reviewed the problems in the light of the following questions: (1) Does the sex ratio vary between families of the same size? (2) Do parents capable of producing only unisexual families exist? (3) Can the residual deviations in the data be satisfactorily explained? Probability analyses were based on Skellam's modified binomial distribution, a special case of the hypergeometrical. The following conclusions were drawn. The probability of a birth being male varies between families of the same size among a complete cross-section of this 19th century German population. There is no evidence for the existence of parents capable of producing only unisexual families. With the assumption that proportions of males vary within families, the apparent anomalies in the data appear to be explicable. These studies have a bearing on the variances observed in further work dealing with family differences such as that of Cohen and Glass ( 1959) on the relation of ABO blood groups to the sex ratio and that of Novitski and Kimball (1958) on birth order, parental age, and sex of offspring. Novitski and Kimball's data are of basic significance, for the interpretations are based on a large volume of material covering a one-year period in which improved statistical techniques were utilized in the data collection, in showing that within these data sex ratio variation showed relatively little dependence on age of mother, whereas it did show dependence on age of father, birth order, and interactions between them. These observations have direct bearing on the larger geographic differences observed in sex ratios as discussed by Russell (1936) and have recently been brought to the fore through the studies of Kang and Cho (1959a, b). If these data stand the tests for biases, they are of significance in showing Korea to have one of the highest secondary sex ratios of any region, 113.5 males to 100 females, as contrasted with the American ratio of about 106 males to 100 females. Of similar interest is the lower rate of twin births, 0.7 per cent in Korea vs. about 1 per cent in Caucasian populations and the fact that nearly twothirds of these tW'in births in Korean peoples are identical, whereas those in the Caucasian groups are only about half that number. The reasons for these differences must lie in the relations of the human X and Y chromosomes and autosomes and the balance of their contained genes. Little or nothing is known about how these factors operate in the given situations.
Acknowledgment
In formulating and organizing the material on which this paper is based I have been fortunate in the helpful discussions and analytical advice contributed so generously by Doctors H. L. Cai'son, K. W. Cooper, H. V. Grouse, C. W. Metz, S. B. Pipkin, W. C. Rothenbuhler, and H. D. Stalker, and others having primary research interests in this field. To them, and particularly to my research associates Doctors S. T. C. Fung and Janice Stadler, our secretary Gladys M. Dicke and to my wife, ]\Iarie S. Gowen, I tender grateful acknowledgment for their contributions to the manuscript. Direct reference is made to some 300 investigations in the body of the paper but the actual number, many of which could equally well have been incorporated, that furnished background to these advances is much larger, more than 1600. To this vast effort toward understanding the foundations for sex we are all indebted.
VII. References
AiDA, T. 1921. On the inheritance of color in a fresh-water fish Aplocheilus latipes (Temmick and Schlegel), with special reference to sexlinked inheritance. Genetics, 6, 554-573.
AiDA, T. 1936. Sex reversal in Aplocheilus latipes and a new explanation of sex differentiation. Genetics, 21, 136-153.
Andersson, T. 1956. Intersexuality in pigs. Kungl. Skogs-och Lantbruksakad. tidskrift Arg., 95, 257-262.
Andke-s, A. H., AND Navashin, M. S. 1936. Morphological analysis of human chromosomes. Proc. Maxim Gorkv Med. Genet. Res. Inst., Moscow, 4, 506-524."
Aronson, M. M. 1959. New evidence for genetic activity of the Y chromosome in D. melanogasler. Records Genet. Soc. Am., 28, 59.
AsDELL, S. A. 1936. Hermaphroditism in goats. Dairy Goat J., 14, 3-4.
Bahner, F., Schwarz, G., H.arnden, D. G., J.\cobs, P. A., HiENz, H. A., AND Walter, K. 1960. A fertile female with XO sex chromosome constitution. Lancet, 2, 100-101 (7141).
Baker, W. K. 1956. Chromosome segregation in D. virilis males with multiple sex chromosomes. Genetics, 41, 907-914.
Baker, W. K., and Spofford, J. B. 1959. Heterochromatic control of position-effect variega
tion in Drosophila. Biol. Contr., Univ. Texas
Publ., 5914, 135-154.
Baltzer, F. 1935. Experiments on .sex-development in Bonellia. The Collecting Net, 10, 3-8.
Bamber, R. C, and Herdman, E. C. 1932. A report on the progeny of a tortoiseshell male cat, together with a discussion of his gametic constitution. J. Genet., 26, 115-128.
Barr, M. L. 1959. Sex chromatin and phenotype in man. Science, 130, 679-685.
Barr, M. L., and Bertram, E. G. 1949. A morphological distinction between neurones of the male and female, and the behavior of the nucleolar satellite during accelerated nucleoprotein synthesis. Nature, London, 163, 676677.
Be.\dle, G. W. 1930. Genetical and cytological studies of Mendelian asynapsis in Zea mays. Cornell Univ. Agric. Exper. Sta., 129, 3-23.
Beadle, G. W., and Ephrussi, B. 1937. Ovary transplants in Drosophila melanogaster: meiosis and crossing over in superfemales. Proc. Nat. Acad. Sc, 23, 356-360.
Beerm.^nn, W. 1956. Nuclear differentiation and functional morphology of chromosomes. Cold Spring Harbor Symposia Quant. Biol, 21, 217-232.
Bellamy, A. W., and Queal, M. L. 1951. Heterosomal inheritance and sex determination in Platypoecilus maculatus. Genetics, 36, 93-107.
Bemis, W. p., and Wilson, G. B. 1953. A new hypothesis explaining the genetics of sex determination in Spinacia oleracea L. J. Hered., 44, 91-95.
Bender, M. A. 1957. X-ray induced chromosome aberrations in normal human tissue cultures. Science, 126, 974-975.
Berg, R. L. 1937. The relative frequency of mutations in different chromosomes of Drosophila melanogaster. II. Sterility mutations. Genetics, 22, 241-248.
Bergman, S., Reitalu, J., Nowakowski, H., and Lenz, W. 1960. The chromosomes in two patients with Klinefelter syndrome. Ann. Human Genet., 24, 81-88.
Berry, R. O. 1941. Chromosome behavior in the germ cells and development of llic gonads in Sciara ocellaris. J. Morphol., 68, 547-576.
Blakeslee, a. F. 1904. Se.xual reproduction in the mucorineae. Proc. Am. Acad. Sc, 40, 203322.
Bloise, W., Bottura, C, de As.sis, L. M., and Ferrari, I. 1960. Gonadal dysgenesis (Turner's .syndrome) with male phenotype and XO chromosomal constitution. Lancet, 2, 1059-60 (7159).
Book, J. A., Fraccvro, M., and Lindsten, J. 1959. Cvtogenetical ()l).scrvations in Mongolism. Acta Paediat., 48, 453-468.
Book, J. A., and Santesson, B. 1960. Malformation syndrome in man associated with triploidv (69 chromosomes). Lancet, 1, 868 (7129).
Bostian, C. H. 1939. Multiple alleles and sex determination in Habrobracon. Genetics, 24, 770-776.
Breider, H. 1942. ZW-Mannchen iind WWWeibchen bei Platypoecilus maculatus. Biol. Zentralbl., 62, 183-195.
Bridges, C. B. 1916. Non-disjunction as proof of the chromosome theory of heredity. Genetics, 1, 1-52; 107-163.
Bridges, C. B. 1921. Triploid intersexes in Drosophila melanogaster. Science, 54, 252-254.
Bridges, C. B. 1922. The origin of variations in sexual and sex-limited characters. Am. Naturalist, 56, 51-63.
Bridges, C. B. 1932. The genetics of sex in Drosophila. In Sex and Internal Secretions, 1st ed., E. Allen, Ed., pp. 55-93. Baltimore: The Williams & Wilkins Company.
Bridges, C. B. 1939. Cytological and genetic basis of sex. In Sex and Internal Secretions, 2nd ed., E. Allen, C. H. Danforth, and E. A. Doisy, Eds., pp. 15-63. Baltimore: The Williams & Wilkins Company.
Bridges, C. B., and Breh.me, K. S. 1944. The mutants of Drosophila melanogaster. Carnegie Inst. Washington Publ., 552, 1-257.
Brosseau, G. E., Jr. 1960. Genetic analysis of the male fertility factors on the Y chromosome of Drosophila melanogaster. Genetics, 45, 257-274.
Burdette, W. J. 1940. The effect of artificially produced tetiaploid regions of the chromosomes of Drosophila melanogaster. Univ. Texas Publ., 4032, 157-163.
Buzzati-Traverso, a. 1941. An extreme case of sex-ratio in D. bilineata. Drosophila Information Service, 14, 49.
Carson, H. L. 1956. A female-producing strain of D. borealis Patterson. Drosophila Information Service, 30, 109-110.
CARSON, H. L., Wheeler, M. R., and Heed, W. B. 1957. A parthenogenetic strain of D. mangabeirai Malogolowkin. Univ. Texas Publ., 5721, 115-122.
Carter, C. O., Hamerton, J. L., Polani, P. E., GuNALP, A., AND Weller, S. D. V. 1960. Chromosome translocation as a cause of familial mongohsm. Lancet, 2, 678-680 (7152).
Cav.alcanti, a. G. L., and Falcao, D. N. 1954. A new type of sex-ratio in Drosophila prosaltans Duda. In Proceedings 9th International Congress Genetics, Part II, pp. 1233-1235. (Caryologia, Vol. VI. Suppl.)
Chang, C. Y., and Witschi, E. 1955. Breeding of sex-reversed males of Xenopus laevis Daudin. Proc. Soc. Exper. Biol. & Med., 89, 150152.
Chang, C. Y., .and Witschi, E. 1956. Genetic control and hormonal reversal of sex differentiation in Xenopns. Proc. Soc. Exper. Biol. & Med., 93, 140-144.
Chu, E. H. Y., and Giles, N. H. 1959. Human chromosome complements in normal somatic cells in culture. Am. J. Human Genet., 11, 63-79.
Cohen, B. H., .and Glass, B. 1959. Further observations on the ABO blood groups and the sex ratio. Am. J. Human Genet.. 11, 274-278.
Cole, L. J., and Hollander, W. F. 1950. Hybrids of pigeon by ringdove. Am. Naturalist, 84, 275308.
Cooper, K. W. 1952. Studies on spermatogenesis in Drosophila. Am. Philos. Soc. Yrbk., 1951, 146-147.
Cooper, K. W. 1956. Phenotypic effects of Y chromosome hyperploidy in Drosophila melanogaster, and their relation to variegation. Genetics, 41, 242-264.
Cooper, K. W. 1959. Cytogenetic analysis of major heterogametic elements (especially Xh and Y) in Drosophila melanogaster and the theory of "heterochromatin." Chromosoma, 10, 535-588.
Crooke, a. C, and Hayward, M. D. 1960. Mosaicism in Klinefelter's Svndrome. Lancet, 1, 1198 (7135).
Crouse, H. V. 1943. Translocations in Sciara; their bearing on chromosome behavior and sex determination. Missouri Agric. Exper. Sta. Res. Bull., 379, 1-75.
Crouse, H. V. 1960a. The nature of the influence of X-translocations on sex of progeny in Sciara coprophila. Chromosoma, 11, 146-166.
Crouse, H. V. 1960b. The controlling element in sex chromosome behavior in Sciara. Genetics, 45, 1429-1443.
Crow, J. F. 1946. The absence of a primary sex factor on the A" chromosome of Drosophila. Am. Naturahst, 80, 663-665.
Danforth, C. H. 1932. Genetics of sexual dimorphism in plumage. In Congress of Genetics, Ithaca, vol. 2, pp. 34-36.
DARLINGTON, C. D., AND Janaki Ammal, E. K. 1945. Chromosome Atlas oj Cultivated Plants. London: George Allen & Unwin, Ltd.
Darlington, C. D., and Haque, A. 1955. Chromosomes of monkevs and men. Nature, London, 175, 32.
de Assis, L. M., Epps, D. R., Bottura, C, and Ferrari, I. 1960. Chromosomal constitution and nuclear sex of a true hermaphrodite. Lancet, 2, 129-130 (7142).
Demerec, M. (Editor). 1950. Biology of Drosophila. New York: John Wiley & Sons, Inc.
de Winiavarter, H. 1912. Etudes sur la spermatogenese humaine. I. Cellule de Sertoli. II. Heterochromosome et mitoses de I'epithelium seminal. Arch. Biol., 27, 91-190.
DE Winiavarter, H., and Oguma, K. 1926. Nouvelles recherches sur la spermatogenese humaine. Arch. Biol., 36, 99-166.
DOBZH.ANSKY, T., AND BRIDGES, C. B. 1928. The reproductive system of triploid intersexes in Drosophila melanogaster. Am. Naturalist 62, 425-434.
DoBZHANSKY, T., AND HoLZ, A. M. 1943. A reexamination of the problem of manifold effects of genes in Drosophila melanogaster. Genetics, 28, 295-303.
DoBZHANSKY, T., AND ScHULTZ, J. 1934. The distribution of sex-factors in the X chromosome of Drosopliila melanogaster. J. Genet., 28, 349-386.
DoBZH.^x.SKY, T., AND Spassky, B. 1941. Intersexes in Drosophila pseiidoobscura. Proc. Nat. Acad. Sc, 27, 556-562.
Dressler, O. 1958. Zytogenetische Untersuchungen an diploidem und polyploidem spinat (Spinacia oleracea L.) unter besonderer Beriicksichtigung der Geschlechtsvererbung als Grundlage einer Imzucht-Heterosis-Zuchtimg. Ztschr. Pflanzenzucht., 40, 385-424.
DuBois, A. M. 1932. A contribution to the embryology of Sciara (Diptera). J. Morphol., 54, 161-195.
Eatox, O. N. 1943. An anatomical study of hermaphroditism in goats. Am. J. Vet. Res., 4, 333-343.
Eaton, 0. N. 1945. The relation between polled and hermaphroditic characters in dairy goats. Genetics, 30, 51-61.
Editorial. 1960. A proposed standard system of nomenclature of human mitotic chromosomes. Lancet, 1, 1063-1065 (7133).
Edwards, A. W. F. 1958. An analysis of Geissler's data on the human sex ratio. Ann. Human Genet., 23, 6-15.
Edwards, J. H., Harnden, D. G., Cameron, A. H., Crosse, M., and Wolff, O. H. 1960. A new trisomic syndrome. Lancet, 1, 787-790 (7128).
Elliott, G. A., Sandler, A., and Rabinowitz, D.
1959. Gonadal dysgenesis in three sisters. J. Clin. Endocrinol., 19, 995.
Esser, K., and Straub, J. 1958. Genetische Untersuchungen an Sordaria macrospora Auersw.. Kompensation und Induktion bei genbedingten Entwicklungsdefekten. Ztschr. indukt. Abstammungs. Vererb., 89, 729-746.
Evans, H. M., and Swezy, O. 1929. The chromosomes in man. Mem. Univ. California, 9, 11.
Fankhauser, G., and Humphrey, R. R. 1959. The origin of spontaneous hcteroploids in the progeny of diploid, triploid, and tetraploid axolotl females. J. Exper. Zool., 142, 379-422.
Ferguson-Smith, M. A. 1960. Cytogenetics in man. Arch. Int. Med., 105, 627-639.
Ferguson -Smith, M. A., and Johnston, A. W.
1960. Chromosome abnormalities in certain diseases of man. Ann. Int. Med., 53, 359-71.
Ferguson-Smith, M. A., Johnston, A. W., and Handmaker, S. D. 1960. Primary amentia and micro-orchidism associated with an XXXY sex chromosome constitution. Lancet, 2, 184187(7143).
Ferguson-Smith, M. A., Johnston, A. W., and Weinberg, A. R. 1960. The chromosome complement in true hermaphroditism. Lancet, 2,126-128(7142).
Ford, C. E. 1960. Human cytogenetics: its present place and future possibilities. Am. J. Human Genet., 12, 104-117.
Ford, C. E., and Hamerton, J. L. 1956. Tlic chromosomes of man. Natm-e, London, 178, 1020-1023.
Ford, C. E., J.'^cobs, P. A., and Lajtha, L. G. 1958. Human somatic clu-omosomes. Nature. London, 181, 1565-1568.
Ford, C. E., Jones, K. W., Miller, O. J., MittwocH, U., Penrose, L. S., Ridler, M., and Sh.-^piro, a. 1959. The chromosomes in a patient showing both mongolism and the Klinefelter syndrome. Lancet, 1, 709-710 (7075).
Ford, C. E., Jones, K. W., Polani, P. E., de Almeida, J. C, and Briggs, J. H. 1959. A sex-chromosome anomaly in a case of gonadal dysgenesis (Turner's syndrome). Lancet, 1, 711-713 (7075).
Ford, C. E., Pol.ani, P. E., Briggs, J. H., and Bishop, P. M. F. 1959. A presumptive human XXY/XX mosaic. Nature, London, 183, 1030-1032.
Fraccaro, M., Ikkos, D., Lindsten, J., Luft, R., and Kaijser, K. 1960. A new type of chromosomal abnormality in gonadal dysgenesis. Lancet, 2, 1144 (7160).
Fraccaro, M., Kaijser, K., and Lindsten, J.
1959. Chromosome complement in gonadal dysgenesis (Turner's syndrome). Lancet, 1, 886(7078).
Fraccaro, M., Kaijser, K., and Lindsten, J. 1960a. Somatic chromosome complement in continuously cultured cells of two individuals with gonadal dysgenesis. Ann. Human Genet., 24, 45-61.
Fraccaro, M., K.aijser, K., and Lindsten, J. 1960b. Chromosomal abnormalities in father and Mongol child. Lancet, 1, 724-727 (7127).
Fr.accaro, M., Kaijser, K., .\nd Lindsten, J. 1960c. A child with 49 chromosomes. Lancet, 2, 899904 (7156).
Fraccaro, M., .-vnd Lindsten, M. 1960. A child with 49 chromosomes. Lancet, 2, 1303 (7163).
Eraser, J. H., Campbell, J., M.^cGillivray, R. C, Boyd, E., .^nd Lennox, B. 1960. The XXX syndrome frequencj^ among mental defectives and fertility. Lancet, 2, 626-627 (7151).
Frost, J. N. 1960. The occurrence of partially fertile triploid metafemales in Drosophila melanogaster. Proc. Nat. Acad. Sc, 46, 47-51.
Fung, S. T. C, and Gowen, J. W. 1957a. The developmental effect of a sex-limited gene in Drosophila melanogaster. J. Exper. Zool., 134, 515-532.
Fung, S. T. C, and Gowen, J. W. 1957b. Pigment-inducing potentialities of testes, ovaries and hermaphroditic (Hr) gonads. J. Exper. Zool., 135, 5-18.
Fung, S. T., and Gowen, J. W. 1960. Role of autosome IV in Drosophila melanogaster sex balance. Rec. Genet. Soc. Am., 29, 70-71.
Gallien, L. 1954. Inversion experimentale du sexe, sous Taction des hormones sexuelles, chez le triton Pleurodeles waltlii Michah. Analyse des consequences genetiques. Bull. Biol. France et Belgique, 88, 1-51.
Gallien, L. 1955. Descendance unisexuee d'une fcmelle de Xenopus laevis Daud. ayant subi, pendent sa phase larvaire. Taction gynogenese (ill benzoate d'oestradiol. Compt. rend. Acad. Sc, 240, 913-915.
Gallien, L. 1956. Inversion experimentale du .sexe chez im Anoure inferieur Xenopus laevis Daudin. Bull. Biol. France et Belgique, 90, 163-183.
Geissler, a. 1889. Beitrage zur Frage des Geschlects verhaltnisses der Geborenen. Ztschr. Sachsischen Statist. Bureaus, 35, 1-24.
Gershenson, S. 1928. A new sex ratio abnormality in Drosophila obscura. Genetics, 13, 488-507.
GoLDSCHMiDT, R. B. 1934. Lymantria. Bibl. Genet., 11, 1-186.
GoLDSCHMiDT, R. B. 1948. New facts on sex determination in D rosopliila mclanogaster. Proc. Nat. Acad. Sc, 34, 245-252.
GoLDSCHMiDT, R. B. 1949. The Beaded Minuteintersexes in Drosophila rnelanogaster Meig. J. Exper.Zool., 112, 233-301.
GoLDSCHMiDT, R. B. 1951. The maternal effect in the production of the Bd-M-intersexes in Drosophila rnelanogaster. J. Exper. Zool., 117, 75110.
GoLDSCHMiDT, R. B. 1955. Theoretical Genetics. Berkeley: Uni\ersitj- of California Press.
GoLDSCHMiDT, R. B. 1957. A remarkable action of the mutant "rudimentary" in Drosophila rnelanogaster. Proc. Nat. Acad. Sc, 43, 731-736.
GOLDSCHMIDT, R. B., AND Katsuki, K. 1927. Erblicher Gynandromorphismus imd somatische Mosaikbildung bei Bombijx niori. Biol. Zentralbl., 47, 45-54.
GOLDSCHMIDT, R. B., AND Katsuki, K. 1928. Cytologie des erblichen Gynandromorphismus von Bombyx mori L. Biol.Zentralbl., 48, 685-699.
GoLDSCHMiDT, R. B.,.'\ndK.'\tsuki, K. 1931. Vierte Mitteilung liber erblichen Gynandromorphismus und somatische Mosaikbildung bei Bombyx mori L. Biol. Zentralbl., 51, 58-74.
Goodrich, H. B. 1916. The germ-cells in Ascaris incurva. J. Exper. Zool., 21, 61-100.
Gordon, M. 1946. Interchanging genetic mechanisms for sex determination in fishes under domestication. J. Hered., 37, 307-320.
Gordon, M. 1947. Genetics of Platypoecilus maculatus. IV. The sex determining mechanism in two wild populations of the Mexican platyfish. Genetics, 32, 8-17.
Gordon, R. R., O'Gorman, F. J. P., Dewhurst, C. J., .\ND Blank, C. E. 1960. Chromosome count in a hermaphrodite with some features of Klinefelter's svndrome. Lancet, 2, 736-739 (7153).
CrOWEN, J. W. 1928. On the mechanism of chromosome behavior in male and female Drosophila. Proc. Nat. Acad. Sc, 14, 475-477.
GowEN, J. W. 1933. Meiosis as a genetic character in Drosophila melanogaster. J. Exper. Zool., 65, 83-106.
GowEN, J. W. 1942. On the genetic basis for hermaphroditism. Anat. Rec, 84, 458 (Abst.).
GowEN, J. W. 1947. Sex determination in Drosophila melanogaster. Rec Genet. Soc Am., 16, 33 (Abst.).
GowEN, J. W., AND Fung, S. T. C. 1957. Determination of sex through genes in a major sex locus in Drosophila mekmogaster. Heredity, 11, 397-402.
GowEN, J. W., AND G.'Vy, E. H. 1933. Eversporting as a function of the Y chromosome in Drosophila melanogaster. Proc Nat. Acad. Sc, 19, 122-126.
GowEN, M. S., AND GowEN, J. W. 1922. Complete linkage in Drosophila melanogaster. Am. Naturalist, 56, 286-288.
GowEN, J. W., AND Nelson, R. H. 1942. Predetermination of sex. Science, 96, 558-559.
Grumbach, M. M., and Barr, M. L. 1958. Cytologic tests of chromosomal sex in relation to sexual anomalies in man. Recent Progr. Hormone Res., 14, 255-324.
H.^CHiNOHE, Y., AND Onishi, N. 1952. On the meiosis of the drone of Honey bee Apis melliUca L. Bull. Nat. Inst. Agric. Sc. (Japan), (G)3, 83-87.
Hammond, J. 1912. A case of hermaphroditism in the pig. J. Anat. Physiol., 46, 307-312.
H.'Vnnah, A. M. 1955. The effect of aging the maternal parent upon the sex ratio in Drosophila melanogaster. Ztschr. indukt. Abstammungs. Vererb., 86, 574-599.
Harnden, D. G. 1960. Abnormalities of chromosome constitution and defects of sex development. Proc. Roy. Soc. Med., 53, 493-494.
Harnden, D. G., and Armstrong, C. N. 1959. Chromosomes of a true hermaphrodite. Brit. M. J., 2, 1287-1288.
Harnden, D. G., and Stewart, J. S. S. 1959. Chromosomes in a case of pure gonadal dysgenesis. Brit. M. J., 2, 1285-1287.
Hartmann, M. 1956. Das Wesen und die Grundsetzlichkeiten des Geschlechts und der Geschlechtsbestimmung im Tier-und Pflanzenreich. In Die Sexualitdt, 2nd ed. Stuttgart: Gustav Fischer.
Hartmann, M., and Bauer, H. 1953. Allgemeine Biologic. Stuttgart: Gustav Fischer.
Hasimoto, H. 1929. Some experiments on the mosaics of Bombyx mori. Japan. J. Genet., 4(3), 113-114.
Hasimoto, H. 1933. Genetical studies on the tetraploid female in the silkworm. I. Bull, sericult. Exper. Sta. Japan, 8, 359-381.
Hasimoto, H. 1934. Genetical studies on the tetraploid female in the silkworm. Bull, sericult. Exper. Sta. Japan, 8, 505-523.
Hayward, M. D. 1960. Sex-chromosome mosaicism in man. Heredity, 15, 219-235.
Hayward, M. D., and Bower, B. D. 1960. Chromosomal trisomy associated with the SturgeWeber syndrome. Lancet, 2, 844-846 (7155).
Henking, H. 1891. LTntersuchungen libsr die ersten Entwichlungsvorgange in den Eiern der Insekten II. Uber spermatogenese und deren Beziehung zur Eintwickelung bei Pyrrhocoris apterus M. Ztschr. wiss. Zool., 51, 685-736.
Hixtox, C. W. 1959. A cytological study of w" chromosome instability in cleavage mitoses of Drosophila ynelanogaster. Genetics, 44, 923933.
HiRSCHHORN, K., Decker, W. H., and Cooper, H.
L. 1960. True hermaphroditism with XY/
XO mosaicism. Lancet, 2, 319-320 (7145).
HOFFENBERG, R., JaCKSOX, W. P. U., AND MULLER, W. H. 1957. Gonadal dysgenesis with menstruation. J. Chn. Endocrinol., 17, 902-907.
HoLL.ANDER, W. F. 1944. Mosaic effects in domestic birds. Quart. Rev. Biol., 19, 285-307.
Hollander, W. F., and Gowen, J. W. 1959. A single-gene antagonism between mother and fetus in the mouse. Proc. Soc. Exper. Biol. & Med., 101, 425-428.
Hollander, W. F., Gowen, J. W., and Stadler, J.
1956. A study of 25 gynandromorphic mice of the Bagg albino strain. Anat. Rec, 124, 223-244.
Hsu, T. C. 1952. Mammalian chromosomes in vitro. 1. The karyotype of man. J. Heredity, 43, 167-172.
Hsu, T. C, Po.MERAT, C. M., and Moorhe.ad, p. S. 1957. Mammalian chromosomes in vitro. VIII. Heteroploid transformation in human cell strain Maves. J. Nat. Cancer Inst., 19, 867-873.
Hughes, W. 1929. The freemartin condition in swine. Anat. Rec, 41, 213-247.
Humphrey, R. R. 1945. Sex determination in ambystomid salamanders : a study of the progeny of females experimentally converted into males. Am. J. Anat., 76, 33-66.
Humphrey, R. R., .\nd Fankhauser, G. 1956. Structure and functional capacity of the ovaries of higher polyploid (4N, 5N) in the Mexican axolotl {Siredon or Amhystoma mexicanum). J. Morphol., 98, 161-198.
Hungerford, D. A., Donnelly, A. J., Nowell, P. C, AND Beck, S. 1959. The chromosome constitution of a human phenotypic intersex. Am. J. Human Genet., 11, 215-236.
Irwin, M. R., and Cole, L. S. 1936. Immunogenetic studies of species and of species hybrids from the cross of Columba livia and Sireptopelia risoria. J. Exper. Zool., 73, 309318.
Ishihara, T. 1956. Cytological studies of tortoiseslu'll male cats. Cytologia, 21, 391-398.
Jacobs, P. A., Baikie, A. G., Court Brown, W. M., Forrest, H. Roy, J. R., STEWaRT, J. S. S., and Lennox, B. 1959. Chromosomal sex in the syndrome of testicular feminisation. Lancet, 2,591-592(7103).
JacoBs, P. A., B.\iKiE, A. G., Court Brown, W. M., MacGrkgok, T. N., Maclean, N., and Harnden, D. G. 1959. P]vidence for the existence of the human "super female." Lancet, 2, 423425 (7100).
JacoBS, P. A., Baikie, X. C;., Court Brown, W. M., AND Strong, J. A. 1959. The somatic chromosomes in mongolism. Lancet, 1, 710 (7075).
Jacobs, P. A., Harnden, D. G., Court Brown, W. M., Goldstein, J., Close, H. G., MacGregor, T. N., Maclean, N., and Strong, J. A. 1960. Abnormalities involving the A^-chromosome in women. Lancet, 1, 1213-1216 (7136).
Jacobs, P. A., and Strong, J. A. 1959. A case of human intersexuality having a possible XXY sex-determining mechanism. Nature, London, 183, 302-303.
Jacobsex, p. 1957. The sex chromosomes in Humulus. Hereditas, 43, 357-370.
Janick, J. 1955. Environmental influences on sex expression in monoecious lines of spinach. Proc. Am. Soc. Horticult. Sc, 65, 416-422.
Janick, J., and Ellis, J. R. 1959. Identification of the sex chromosome in Sjyinacia oleracea. (Abst.) Rec. Genet. Soc. Am., 28, 78.
Janick, J., Mahoney, D. L., and Pfahler, P. L. 1959. The trisomies of Spinacia oleracea. J. Hered., 50, 47-50.
Jaxick, J., axd Stevexson, E. C. 1955a. The effects of polyploidy on sex expression in spinach. J. Hered., 46, 151-156.
Janick, J., and Stevenson, E. C. 1955b. Genetics of the monoecious character in spinach. Genetics, 40, 429-437.
Jennings, H. S. 1939. Genetics of Paramecium hursaria. I. Mating types and groups, their interrelations and distribution; mating behavior and self sterility. Genetics, 24, 202233.
Johnston, E. F., Zeller, J. H., and Cantwell, G. 1958. Sex anomalies in swine. J. Hered., 49, 255-261.
Kang, Y. S., and Cho, W. K. 1959a. Data on the biology of Korean populations. Human Biol., 31, 244-251.
K.\ng, Y. S., and Cho, W. K. 1959b. The sex ratio at birth of the Korean population. Eugenics Quart., 6, 187-195.
Kawaguchi, E. 1928. Zytologische Untersuchungen am Seidenspinner und seine Verwandten. I. Gametogenese von Bombyx mori L. und Bom,byx rnandarina M. und ihre Bastarde. Ztschr. Zellforsch., 7, 519-533.
Keller, K., and Tandler, J. 1916. Uber das Verhalten der Eihaeute bei der Zwellingstriichtigkeit des Rindes. Wien. tieriirztl. Monatsschr., 3, 513-526.
KiHARA, H. 1925. Chromosomes of Rinncx acctosella L. Botan. Mag., Tokyo, 39, 353-360.
KiHARA, H., AND Ono, T. 1923. Cytological studies on Rumex L. I. Chromosomes of Rumex acetosa L. Botan. Mag., Tokyo, 37, 84-91.
KiHARA, H., AXD Yamamoto, Y. 1935. Chromosomenverhiiltnisse bei Aucidia chinensis Benth. Agric. & Horticult., Japan, 10, 2485-2496.
Kimball, R. F. 1939. Mating tvpes in Euplotes. Am. Naturalist, 73, 451-456.
KoDANi, M. 1958. The supernumerary chromosome of man. .\in. J. Human Genet.. 10, 125140.
KoLLER, P. C. 1937. The genetical and mechanical properties of sex chromo-somes. III. Man. Proc. Roy. Soc. Edinburgh, 57, 194-214.
KoMAi, T. 1952. On the origin of the tortoiseshell male cat — a correction. Proc. Japan Acad., Tokyo, 28, 150-155.
KoMAi, T., AXD Ishihara, T. 1956. On the origin of the male tortoiseshell cat. J. Hered., 47, 287291.
IvoNDO, K. 1952. Studies on intersexuality in milk goats. Japan. J. Genet., 27, 131-141.
KoNDO, K. 1955a. Studies on intersexuality in milk goats. III. On the homozygous recessive males and the methods of eliminating intersexual gene. Bull. Nat. Inst. Agric. Sc, Japan, ser. G, 10, 107-117.
KoNDO, K. 1955b. The frequency of occurrence of intersexes in milk goats. Japan. J. Genet., 30, 139-146.
IvRivsHENKO, J. 1959. New evidence for the homology of the short euchromatic elements of the X and Y chromosomes of Drosophila busckii with the microchromosome of Drosophila melayiogaster . Genetics, 44, 1027-1040.
Lebedeff, G. a. 1934. Genetics of liermaphroditism in Drosophila virilis. Proc. Nat. Acad. Sc, 20, 613-616.
Lejeune, J., Gautier, M., and Turpin, R. 1959a. Les chromosomes humaines en culture de tissus. Compt. rend. Acad. Sc, 248, 602-603.
Lejeune, J., Gautier, M., and Turpin, R. 1959b. Etude des chromosomes somaticjues de neuf enfants mongoliens. Compt. rend. Acad. Sc, 248, 1721-1722.
Lejeune, J., Turpin, R., and G.^utier, M. 1959a. Le Mongolisme, premier exemple d 'aberration autosomique humaine. Ann. Genet., 2, 41-19.
Lejeune, J., Turpin, R., and Gautier, M. 1959b. Le mongolisme, maladie chromosomique (trisomie). Bull. Acad. Nat. Med., 143, 256-265.
Lewis, D. 1942. Evolution of sex in flowering plants. Biol. Rev., Cambridge Phil. Soc, 17, 46-67.
L'Heritier, p. 1951. The CO2 sensitivity problem in Dro.sophila. Cold Spring Harbor Symposia Quant. Biol., 16, 99-112.
L'Heritier, P. 1955. Les \'irus integres et I'unite cellulaire. Ann. Biol., 31, 481-496.
LiLLiE, F. R. 1917. The freemartin; a study of the action of sex hormones in the foetal life of cattle. J. Exper. Zool., 23, 371.
Love, A. 1944. Cj'togenetic studies on Rumex subgenus Acetosella. Hereditas, 30, 1-136.
Love, A. 1957. Sex determination in Rumex. Proc Genet. Soc. Canada, 2, 31-36.
Love, A., and S.\rk.ar, N. 1956. Cytotaxonomy and sex determination of Rumex paucifolius. Canad. J. Botany, 34, 261-268.
LuBS, H. A., Jr., Vilar, O., and Bergenst.al, D. M. 1959. Familial male pseudohermaphroditism with labial testes and partial feminization: endocrine studies and genetic aspects. J. Clin. Endocrinol., 19, 1110-1120.
Mackensen, O. 1951. Viability and sex determination in the honev bee {Apis ynellijcra L.). Genetics, 36, 500-509.
Mackensen, O. 1955. Further studies on a lethal series in the honey bee. J. Hered., 46, 72-74.
Magni, G. E. 1952. Sex-ratio in Drosophila bifasciata. 1st. Lombardi di Scienze e Lettere, 85,391^11.
Magni, G. E. 1953. 'Sex-ratio,' a non-Mendelian character in Drosophila bifasciata. Nature, London, 172,81.
Magni, G. E. 1954. Thermic cure of cytoplasmic sex-ratio in Drosophila bifasciata. In Proceedings 9th International Congress of Genetics, Bellagio, pp. 1213-1216.
Magni, G. E. 1957. Analisi della trasmissione delle particelle citoplasmatiche "sex-ratio" in Drosophila bifasciata. Atti. 3 Riun. A.G.I., Ric Sci. 27, 67-70. (Suppl.)
Making, S. 1950. Contribution of the sex chromosome in an intersex goat. A preliminary note. Iden-Sogo-Kenkya, 1, 1-3.
Making, S. 1951. Chromosome Numbers in Animals, 2nd ed. Ames: Iowa state College Press.
Making, S., and Sa.saki, M. 1959. On the chromosome nimiber of man. Proc. Japan Acad , 35, 99-104.
Malogolowkin, C. 1958. Maternally inherited sex-ratio conditions in Drosophila unllistoni and Drosophila paulistorum. Genetics, 43, 276-286.
Malogolowkin, C, and Pgulson, D. F. 1957. Infective transfer of maternally inherited abnormal sex -ratio in Drosophila willistoni. Science, 125, 32.
Malogolowkin, C, Poulson, D. F., and Wright, E. Y. 1959. Experimental transfer of maternally inherited abnormal sex-ratio in Drosophila uiillistoni. Genetics, 44, 59-74.
Mampell, K. 1941. Female sterility in interracial hybrids of Drosophila pseudoobscura. Proc Nat. Acad. Sc, 27, 337-341.
McClung CE. The Accessory Chromosome-Sex Determinant?. (1902). The Biological Bulletin. 3 (1–2): 43–84. doi:10.2307/1535527
Metz, C. W. 1938. Chromosome behavior, inheritance and sex determination in Sciara. Am. Naturalist, 72, 485-520.
Metz, C. W. 1959. Chromosome behavior and cell lineage in triploid and mosaic salivary glands of species hvbrids in Sciara. Chromosoma, 10, 515-534.
Minouchi O. and Ohta T. On the number of chromosomes and the type of sex chromosomes in man. (1934) Cytologia 5: 472-90.
Mittwoch, U. 1952. The chromosome complement in a mongolian imbecile. xA.nn. Eugenics, London, 17, 37.
Morgan, L. V. 1925. Polyploidy in Drosophila melanogaster with two attached A' chromosomes. Genetics, 10, 148-178.
Morgan, L. V. 1929. Contributions to the genetics of Drosophila simulans and Drosophila ynelanogaster . VIII. Composites of Drosophila melanogaster. Publ. Carnegie Inst. Washington, 399, 223-296.
Morgan, T. H. 1914. Heredity and Sex, 2nd ed. New York : Columbia University Press.
Morgan, T. H., and Bridges, C. B. 1919. Contributions to the genetics of Drosophila melanogaster. I. The origin of gynandromorphs. Publ. Carnegie Inst. Washington, 278, 1-122.
Morgan, T. H., Bridges, C. B., and Sturtevant, A. H. 1925. The genetics of Drosophila. Bibliog. Genet., 2, 1-262.
MoRiWAKi, D., and Kitag.^wa, 0. 1957. Sexratio-J in Drosophila bifasciata. I. A preliminary note. Japan. J. Genet., 32, 208-210.
Moses, M. S., and Metz, C. W. 1928. Evidence that the female is responsible for the sex ratio in Sciara (Diptera). Proc. Nat. Acad. Sc, 14, 928-930.
MULD.4L, S., AND OcKEY, C. H. 1960. The "double male," a new chromosome constitution in Klinefelter syndrome. Lancet, 2, 492 (7148).
MuRDY, W. H., AND Carson, H. L. 1959. Parthenogenesis in Drosophila mangabeirai Malog. Am. Naturalist, 93, 355-363.
Nelson, L., Ferrari, I., and Bottura, C. 1960. Chromosomal constitution in a case of Klinefelter's syndrome. Lancet, 2, 319 (7145).
Neuhaus, M. E. 1939. A cytogenetic study of the i' chromosome of Drosophila melanogaster. J. Genet., 37, 229-254.
Newby, W. W. 1942. A study of intersexes produced by a dominant mutation in Drosophila virilis. Blanco Stock. Univ. Texas Publ., 4228, 113-145.
NiLSsox, I. M., Bergman, S. Reit.\lu, J., and W.^lDENSTROM, J. 1959. Haemophilia A in a "girl" with male sex-chromatin pattern. Lancet, 2, 264-266 (7097)
NoviTSKi, E. 1947. Genetic analysis of an anomalous sex ratio condition in Drosophila affinis. Genetics, 32, 526-534.
NoviTSKi, E., AND Ki.MBALL, A. W. 1958. Birth order, parental ages, and sex of offspring. Am. J. Human Genet., 10, 268-275.
NowAKOwsKi, H., Lenz, W., and Parada, J. 1959. Diskrepenz zwisclien Chromatinbefund und genet ischem Gcschlecht l^eim Klinefeltersyndrom. Acta Endocrinol., 30, 296-320.
Ono, T. 1930. Further investigations on the cytologv of Rumex VI-VIII. Botan. Mag., Tokyo, 44, 168-176.
Ono, T. 1935. Chromosomen und sexualitat von Rumex acetosa. Sc. Repts., Tohoku Univ. Biol., ser. IV, 10, 41-210.
Ono, T. 1940. Vorkommen der triploiden intersexe bei Humidus japnnicns. Japan. J. Botan. & Zool., 8, 1632-1634.
OvERZiER, Claus. 1955. Hermaphroditismus verus. Acta Endocrinol., 20, 63-80.
P.AiNTER, T. S. 1921. The i' chromosome in mammals. Science, 53, 503-504.
Painter, T. S. 1923. Studies in mammalian spermatogenesis. II. The spermatogenesis of man. J. Exper. Zool., 37, 291-334.
PaTAU, K. 1960. The identification of individual chromosomes, especially in man. Am. J. Human Genet., 12, 250-276.
PaVTAU, K., Smith, D. W., Therman, E., Inhorn, S. L., .-vno Wagner, H. P. 1960. Multiple congenital anomaly caused by an extra autosome. Lancet, 1,790-793 (7128)."
Patterson, J. T. 1938. Sex differentiation. Al)crrant forms in Drosophila and sex differentiation. Am. Natm-alist, 72, 193-206.
Patterson, J. T., Stone, W. S., and Bedechek, S. 1937. Further studies on X chromosome balance in Drosophila. Genetics, 22, 407-426.
Penrose, L. S., Ellis, J. R., and Delhanty, J. D. A. 1960. Chromosomal translocations in mongolism and in normal relatives. Lancet, 2, 409(7147).
Pipkin, S. B. 1940. Multiple sex genes in the X chromosome of Drosophila melanogaster. Univ. Texas Publ., 4032, 126-156.
Pipkin, S. B. 1941. Intersex modifying genes in wild strains of Drosophila melanogaster. Genetics, 26, 164-165.
Pipkin, S. B. 1942. Intersex modifying genes in wild strains of Drosophila melanogaster. Genetics, 27, 286-298.
Pipkin, S. B. 1947. A search for sex genes in the second chromosome of Drosophila melanogaster using the triploid method. Genetics, 32, 592-607.
Pipkin, S. B. 1959. Sex balance in Drosophila melanogaster: Aneuploidy of short regions of chromosome 3, using the triploid method. Univ. Texas Publ., 5914, 69-88.
Pipkin, S. B. 1960. Sex balance in Drosophila melanogaster : Aneuploidy of long regions of chromosome 3, using the triploid method. Genetics, 45, 1205-1216.
PoLANi, P. E., Bishop, P. M. F., Ferguson-Smith, M. A., Lennox, B., Stew.\rt, J. S. S., and Prader, a. 1958. Colour vision studies and the A^ chromosome constitution of patients with Klinefelter's syndrome. Natuie, London, 182, 1092-1093.
PoLANi, P. E., Briggs, J. H., Ford, C. E., Clarke, O. M,, AND Berg, J. M. 1960. A Mongol girl with 46 chromosomes. Lancet, 1, 721-724 (7127).
PoLANI, P. E., LeSSOF, M. H., AND BiSHOP, P. M. F. 1956. Colour-blindness in "ovarian agenesis" (gonadal dysplasia). Lancet, 2, 118-120 (6934).
PooLE, H. K., AND Olsen, M. W. 1958. Incidence of parthenogenetic development in eggs laid bv 3 strains of dark Cornish chickens. Proc. Soc. Exper. Biol. & Med., 97, 477-478.
PouLSON, D. F. 1940. The effects of certain X chromosome deficiencies on the development of D. melanogaster. J. Exper. Zool., 83, 271326.
Price, J. B., Jr., 1949. The location of a sex limited factor, Ix", in D. virilis. Univ. Texas Publ., 4920, 22-23.
Puck, T. T. 1958. Genetic studies on somatic mammalian cells in vitio. J. Cell. Physiol. Suppl. 1, 52, 287-311.
Puck, T. T., Robinson, A., .and Tjio, J. H. 1960. Familial primary amenorrhea due to testicular feminization: a human gene affecting sex differentiation. Proc. Soc. Exper. Biol. & Med., 103, 192-196.
Raper, J. W. 1960. Control of SOX in fungi. Am. J. Botan., 47, 794-808.
Rasmussen, I. E. 1957. Tentativi di infezione sperimentale con il carattere "sex ratio" in Drosophila bifasciata. Atti III Rium. A. Genet. Ital. T>a Ric. Sc, Suppl., 27, 71-74.
Ravix, A. \V. 1960. The origin of bacterial species, genetic recombination and factors limiting it between bacterial populations. Bact. Rev., 24, 201-221.
Redfield, H. 1926. The maternal inheritance of a sex-limited lethal effect in Drosophila melanogaster. Genetics, 11, 482-502.
Reynolds, J. P. 1938. Sex determination in a bisexual strain of Sciara coprophila Lintner. Genetics, 23, 203-220.
Rick, C. M., and Hanna, G. C. 1943. Determination of sex in Asparagus officinalis L. Am. J.Botan., 30, 711-714.
Ris, H., AND Kerr, W. E. 1952. Sex determination in the honey-bee. Evolution, New York, 6, 444-445.
RoTHENBUHLER, W. C. 1955. Hereditary aspects of gynandromorph occurrence in honey bees (.4/j^s mellifica). Iowa State Coll. J. Sc, 29, 487-488.
RoTHENBUHLER, W. C. 1957. Diploid male tissue as new evidence on sex determination in honey bees {Apis mellifera). J. Hered., 48, 160-168.
RoTHENBUHLER, W. C. 1958. Progress and problems in the analysis of gynandromorphic honey bees. In Proceedings 10th International Congress of Entomology, vol. 2, pp. 867-874.
RoTHENBUHLER, W. C., GoWEN, J. W., AND PaRK, O. W. 1952. Androgenesis with zygogenesis in gynandromorphic honey bees {Apis mellifera L.). Science, 115, 637-638. Russell, W. L., Russell, L. B., and Gow^er, J. S.
1959. Exceptional inheritance of a sex-linked gene in the mouse explained on the basis that X/0 sex-chromosome constitution is female. Proc. Nat. Acad. Sc, 45, 554-560.
Russell, W. T. 1936. Statistical study of the sex ratio at birth. J. Hyg., 36, 381-401. Sandberg, a. a., Crosswhite, L. H. and Gordy, E. 1960. Trisomy of a large chromosome. Association with mental retardation. J. A. M. A., 174, 221-225.
Sandberg, A. A., Koepf, G. F., Crosswhite, L. H., AND Hauschka, T. 1960. The chromosome constitution of human marrow in various developmental and blood disorders. Am. J. Human Genet., 12, 231-249.
Sanderson, A. R., and Hall, D. W. 1948. The cytology of the honey bee, Apis mellifica L. Nature^ London, 162, 34-35.
Sanderson, A. R., and Hall, D. W. 1951. Sex in the honey bee. Endeavour, 10, 33-39.
Sasaki, M., and Making, S. 1960. The chromosomal constitution of a human hermaphrodite. Texas Rep. Biol. & Med. 18, 493-500.
Satina, S., and Blakeslee, a. F. 1925. Studies on biochemical differences between (-I-) and ( — ) sexes in mucors. Proc. Nat. Acad. Sc, 11, 528534.
Sato, H. 1942. Cytological studies of the eggs of silkworm exposed to high temperature. I, II. Sanshigaku-zassi, 14(1).
Schmieder, R. G., and Whiting, P. W. 1947. Reproductive economy in the chalcidoid wasp Melittobia. Genetics, 32, 29-37.
Schrader, F. 1928. Die Geschlechtschrornosomen {The sex chromosomes). Berlin: Gebriider Borntraeger.
Schrader, F., and Sturtevant, A. H. 1923. A note on the theorv of sex determination. Am. Naturalist, 57, 379-381.
Shen, T. H. 1932. Zytologische Untersuchungen iiber sterilitat bei Mannchen von Drosophila mchuiogasler und bei F. Mannchen der Kreuzung zwischen D. simulans Werbchen und D. )uelanogaster Mannchen. Ztschr. Zellforsch., 15, 547-580.
Shiwago, p. I., AND Andres, A. H. 1932. Die Geschlechtschromosomen in der spermatogenese des Menschen. Ztschr. Zellforsch., 16, 413-431.
Smith, B. W. 1955. Sex chromosomes and natural polyploidy in dioecious Rumex. J. Hered., 46, 226-232.
Smith, D. W., Patau, K., Therivun, E., and Inhorn, S. L. 1960. A new autosomal trisomy syndrome ; multiple congenital anomalies caused bv an extra chromosome. J. Pediat., 57, 338345.
Smith, S. G. 1955. Cytogenetics of obligatory parthenogenesis. Canad. EntomoL, 87, 131135.
Smith-White, S. 1955. The life history and genetic system of Leucopogon juniperinus. Heredity, 9, 79-91.
Sneep, J. 1953. The significance of andromonoecy for the breeding of Asparagus officinalis L. Euphytica, 2, 89-95.
Snell, G. D. 1935. The determination of sex in Habrobracon. Proc Nat. Acad. Sc, 21, 446453.
SoNNEBORN, T. M. 1937. Sex, sex inheritance and sex determination in Paramecium aurelia. Proc. Nat. Acad. Sc, 23, 378-385.
SoNNEBORN, T. M. 1947. Recent advances in the genetics of Paramecium and Euplotes. Advances Genet., 1, 264-358.
SoNNEBORN, T. M. 1949. Ciliated protozoa: Cytogenetics, genetics, and evolution. Ann. Rev. Microbiol., 3, 55-80.
SoNNEBORN, T. M. 1957. Breeding systems, reproductive methods, and species in protozoa. In The Species Problem, pp. 163-324. Washington : American Association for the Advancement of Science.
Sprackling, L. S. 1960. The chromosome complement of the developing eggs produced by Drosophila parthenogenetica Stalker virgin females. Genetics, 45, 243-256.
Spurway, H., and Haldane, J. B. S. 1954. Genetics and cytology of Drosophila suhohscura. IX. An autosomal recessi\-e mutant transforming homogametic zygotes into intersexes. J. Genet., 52, 208-225. "
Stalker, H. D. 1942. Triploid intersexuality in D. americana Spencer. Genetics, 27, 504-518.
Stalker, H. D. 1954. Parthenogenesis in Drosophila. Genetics, 39, 4-34.
Stalker, H. D. 1956a. On the evolution of parthenogenesis in Lonchoptera (Diptera). Evolution, 10, 34S-359.
Stalker, H. D. 1956b. Selection within a unisexual strain of Drosophila (Abstr.). Rec. Genet. Soc Am., 25, 662, Genetics, 41, 662.
Stern, C. 1929. Untersuchungen iiber Aberrationen des Y chromosoms \-on Drosophila melanogaster. Ztschr. indukt. Abstammungs. Vererb., 51, 253-353.
Stern, C. 1936. Somatic crossing-over and segregation in Drosophila melanogaster. Genetics, 21, 625-730.
Stern, C. 1959a. Color-blindness in Klinefelter's syndrome. Nature, London, 183, 1452-1453.
Stern, C. 1959b. Use of the term "superfemale". Lancet, 2, 1088 (7111).
Stern, C, and Hadorn, E. 1938. The determination of sterility in Drosophila males without a complete Y chromosome. Am. Naturalist, 72, 42-52.
Sternberg, W. H., and Ivloepfer, H. W. 1960. Genetic and pathologic study of "simulant" females (testicular feminization syndrome). Am. Soc. Human Genet., 11-12 (Abstr.).
Stewart, J. S. S. 1959. Testicular feminization and colour-blindness. Lancet, 2, 592-594 (7103).
Stew.art, J. S. S. 1960a. Gonadal dj'sgenesis. The genetic significance of unusual variants. Acta Endocrinol., 33, 89.
Stewart, J. S. S. 1960b. Genetic mechanisms in Inmian intersexes. Lancet, 1, 825-826 (7128).
Stewart, J. S. S., and Sanderson, A. R. 1960. Fertility and ohgophrenia in an apparent triplo-X-female. Lancet, 2, 21-22 (7140).
Stone, W. S. 1942. The Ix" factor and sex determination. Univ. Texas Publ., 4228, 146152.
Sturtevant, a. H. 1920a. Intersexes in Drosophila simulans. Science, 51, 325-327.
Sturtevant, A. H. 1920b. Genetic studies on Drosophila simulans. I. Introduction. Hybrids with Drosophila melanogaster. Genetics, 5, 488-500.
Sturtevant, A. H. 1921. Genetic studies on Drosophila simulans. III. Autosomal genes; general discussion. Genetics, 6, 179-207.
Sturtevant, A. H. 1923. Inheritance of direction of coiling in Limnaea. Science, 58, 269270.
Sturtevant, A. H. 1945. A gene in Drosophila melanogaster that transforms females into males. Genetics, 30, 297-299.
Sturtevant, A. H. 1946. Intersexes dependent on a maternal effect in hybrids between Drosophila repleta and D. neorepleta. Proc. Nat. Acad. Sc, 32, 84-87.
Sturtevant, A. H. 1949. The Beaded Minute combination and sex determination in Drosophila. Proc. Nat. Acad. Sc, 35, 311-313.
Sturtevant, A. H. 1956. A highly specific complementary lethal system in Drosophila melanogaster' Genetics, 41, 118-123.
Sturtevant, A. H., and Dobzhanskv, T. 1936. Geographical distribution and cytology of "sex ratio" in Drosophila pseudoobscura and related species. Genetics, 21, 473-490.
Suomalainen, E. 1950. Parthenogenesis in animals. Advance Genet., 3, 193-253.
SuoMALAiNEN, E. 1954. The cytology of the parthenogenetic curculionids of Switzerland. Chromosoma, 6, 627-655.
Syverton, J. T. 1957. Comparative studies of normal and malignant human cells in continuous culture. In Cellular Biology, Nucleic Acids and Viruses. New York Acad. Sc, special publ., 5, 331-340.
Taber, S., 3rd. 1955. Evidence of binucleate eggs in the honey bee. J. Hered., 46, 156.
Tanaka, Y. 1952. Genetics of the Bombyx mori. Tokyo : Shokado Company.
Tanaka, Y. 1953. Genetics of the silkworm, Bombyx mori. Advance Genet., 5, 239-317.
TaYL0R, J. H. 1957. The time and mode of duplication of chromosomes. Am. Naturalist, 91,209-221.
TaziMA, Y. 1941. Larval striations as a simple means for the differentiation of sex. J. sericult. Sc. Japan., 12(3).
TaVziMA, Y. 1952. Sex determination. In Genetics of the Bombyx mori, Y. Tanaka, Ed., pp. 351372. Tokyo : Shokado Company.
Thompson, D. H. 1920. A new type of sex-linked lethal mutant in Drosophila. Anat. Rec, 20, 215.
Tjio, J. H., AND Levan a. 1956. The chromosome number of man. Hereditas, 42, 1-6.
Tjio, J. H., and Puck, T. T. 1958a. Genetics of somatic mammalian cells. II. Chromosomal constitution of cells in tissue culture. J. Exper. Med., 108, 259-268.
Tjio, J. H., and Puck, T. T. 1958b. The somatic chromosomes of man. Proc. Nat. Acad. Sc, 44, 1229-1237.
Tjio, J. H., Puck, T. T., and Robinson, A. 1959. The somatic chromosomal constitution of some human subjects with genetic defects. Proc. Nat. Acad. Sc, 45, 1008-1016.
Tokunaga, C. 1958. The Y chromosome in sex determination of Aphiochaeta xanthina. In Proceedings 10th International Congress of Genetics, Montreal, Vol. 2, pp. 295-296 (Abstr.).
Tucker, K. W. 1958. Automictic parthenogenesis in the honey bee. Genetics, 43, 299-316.
TuRPiN, R., Lejeune, J., Lafourcade, J., AND Gautier, M. 1959. Aberrations chromosomiques et maladies humaines. La polydysspondylie a 45 chromosomes. Compt. rend. Acad. Sc, 248, 3636-3638.
Warmke, H. E. 1946. Sex determination and sex balance in Melandrium. Am. J. Botan., 33, 648-660.
W.ARMKE, H. E., AND Blakeslee, A. F. 1939. Sex mechanisms in polyploids of Melandrium. Science, 89, 391-392.
Warmke, H. E., D.widson, H., and LeClerc, G. 1945. Polyploidy investigations. In Annual Report, Director of Department of Genetics, Carnegie Institute Year Book, vol. 44, pp. 113115.
Weir, J. A. 1958. Sex ratio related to sperm source in mice. J. Hered., 49, 223-227.
Weishons, W. J., AND Russell, L. B. 1959. The }' chromosome as the bearer of male determining factors in the mouse. Proc. Nat. Acad. Sc, 45, 560-566.
Westergaard, M. 1940. Studies on cytology and sex determination in polyploid forms of Melandrium album. Dansk Botau. .Vik., 10, 1131.
We.stergaard, M. 1946a. Structural changes of the ?? chromosome in the offspring of polyploid Melandrium. Hereditas, 32, 60-64.
Westergaard, M. 194613. Aberrant ?? chromosomes and sex expression in Melandrium album. Hereditas, 32, 419^43.
WESTERcaARD, M. 1948. The relation between chromosome constitution and sex in the offspring of triploid Melandrium. Hereditas, 34, 257-279.
Westergaard, M. 1953. Uber den Mechanismus der Geschlechtsbestimmung bei Melandrium album. Naturwissenschaften, 40, 253-260.
Westergaard, M. 1958. The mechanism of sex determination in dioecious flowering plants. Advance Genet., 9, 217-281.
White, M. J. D. 1954. Animal Cytology and Evolution, 2nd ed. Cambridge: Cambridge University Press.
Whiting, P. W. 1933a. Selective fertilization and sex determination in Hymenoptera. Science, 78, 537-538.
Whiting, P. W. 1933b. Sex determination in Hymenoptera. Biol. Bull., 65, 357-358.
Whiting, P. W. 1935. Sex determination in bees and wasps. J. Hered., 26, 263-278.
Whiting, P. W. 1943a. Multiple alleles in complementary sex determination of Habrobracon. Genetics, 28, 365-382.
Whiting, P. W. 1943b. Androgenesis in the parasitic wasp Habrobracon. J. Hered., 34, 355-366.
Whiting, P. W. 1958. Diploid males and triploid females in Habrobracon and Mormoniella. Bull. A. Southeastern Biol., 5, 16.
Whiting, P. W., and Whiting, A. R. 1925. Diploid males from fertilized eggs in Habrobracon. Science, 63, 437-438.
WiESE, L. 1960. Die diplogenotypische Geschlechtsbestimmung. Fortschr. Zool., 12, 295335.
Wilson, E. B. 1928. The Cell in Development and Heredity, 3rd ed. New Yoik: The Macmillan Company.
WiNGE, O. 1922. A pecuhar mode of inheritance and its cytological explanation. Compt. rend, tr^.v. lab.Carlsberg, 17, 1-9.
WiNGE, 0. 1934. The experimental alteration of sex chromosomes into autosomes and vice versa, as illustrated by Lebistes. Compt. rend, trav. lab. Carlsberg, ser. physioL, 21, 1-49.
WiNGE, 0., AND DiTLEVSEN, E. 1948. Colour inheritance and sex determination in Lebistes. Compt. rend. trav. lab. Carlsberg, ser. phvsiol., 24, 227-248.
WiTSCHi, E., Nelson, W. O., and Segal, 8. J. 1957. Genetic developmental and hormonal aspects of gonadal dysgenesis and sex inversion in man. J. Clin. Endocrinol., 17, 737-753.
Wolf, E. B. 1960. Eine Nachuntersuchung zur Cytologie der Honigbiene {Apis m,ellifica L.). Zool. Beitr., neue folge, 5, 373-391.
Yamamoto, T.-O. 1953. Artificially induced sexreversal in genotypic males of the medaka {Oryzim latipes). J. Exper. Zool., 123, 571594.
Yamamoto, T.-O. 1957. Esterone-induced intersex of genetic male in the medaka, Oryzias latipes. J. Fac. Sc. Hokkaido Univ., ser. VI, Zool., 13, 440-444.
Yaalamoto, T.-O. 1959a. A further study on induction of functional sex-re\'ersal in genotypic males of the medaka {Oryzias latipes) and progenies of sex-reversals. Genetics, 44, 739-757.
Yam.amoto, T.-O. 1959b. The effect of estrone dosage level upon the percentage of sexreversals in genetic male {XY) of the medaka (Oryzias latipes). J. Exper. Zool., 141, 133-153.
Yamamoto, Y. 1938. Karyogenetische I'ntersuchungen bei der Gattung Rumex. VI. Gesclilochtsbestimmung bei eu- und aneuploiden Pflanzen von Rumex acetosa L. Mem. Coll. Agric. Kyoto Imp. Univ., 43, 1-59.
Yao, T., and Olsen, M. W. 1955. Microscopic observations of parthenogenetic embryonic tissues from virgin turkeys. J. Hered., 46, 133-134.
YoKOYAMA, T. 1959. Silkworm Genetics Illustrated. Tokyo: Japan Society for Promotion of Science.
Zoschke, U. 1956. Untersuchungen iiber die Bestimmung des Geschlechtes beim Spinat unter besonderer Beriicksichtigung der Zuchtung eines monozischen oder gleichzeitig schossenden Spinats. Ztschr. Pflanzenziicht., 35, 257-296.
