Identification of a novel sex determining chromosome in cichlid fishes that acts as XY or ZW in different lineages

Sex determination systems are highly conserved among most vertebrates with genetic sex determination, but can be variable and evolve rapidly in some. Here, we study sex determination in a clade with exceptionally high sex chromosome turnover rates. We identify the sex determining chromosomes in three interspecific crosses of haplochromine cichlid fishes from Lakes Victoria and Malawi. We find evidence for different sex determiners in each cross. In the Malawi cross and one Victoria cross the same chromosome is sex-linked but while females are the heterogametic sex in the Malawi species, males are the heterogametic sex in the Victoria species. This chromosome has not previously been reported to be sex determining in cichlids, increasing the number of different chromosomes shown to be sex determining in cichlids to 12. All Lake Victoria species of our crosses are less than 15,000 years divergent, and we identified different sex determiners among them. Our study provides further evidence for the diversity and evolutionary flexibility of sex determination in cichlids, factors which might contribute to their rapid adaptive radiations.

     

Comparative chromosome painting between two marsupials: origins of an XX/XY1Y2 sex chromosome system

Cross-species chromosome painting was used to investigate genome rearrangements between tammar wallaby Macropus eugenii (2n = 16) and the swamp wallaby Wallabia bicolor (2n = 10♀/11♂), which diverged about 6 million years ago. The swamp wallaby has an XX female:XY₁Y₂ male sex chromosome system thought to have resulted from a fusion between an autosome and the small original X, not involving the Y. Thus, the small Y₁ should represent the original Y and the large Y₂ the original autosome. DNA paints were prepared from flow-sorted and microdissected chromosomes from the tammar wallaby. Painting swamp wallaby spreads with each tammar chromosome-specific probe gave extremely strong and clear signals in single-, two-, and three-color FISH. These showed that two tammar wallaby autosomes are represented unchanged in the swamp wallaby, two are represented by different centric fusions, and one by a tandem fusion to make the very long arms of swamp wallaby Chromosome (Chr) 1. The large swamp wallaby X comprises the tammar X as its short arm, and a tandemly fused 7 and 2 as the long arm. The acrocentric swamp wallaby Y₂ is a 2/7 fusion, homologous with the long arm of the X. The small swamp wallaby Y₁ is confirmed as the original Y by its painting with the tammar Y. However, the presence of sequences shared between the microdissected tammar Xp and Y on the swamp wallaby Y₂ implies that the formation of the compound sex chromosomes involved addition of autosome(s) to both the original X and Y. We propose that this involved fusion with an ancient pseudoautosomal region followed by fission proximal to this shared region. Received: 16 October 1996/Accepted: 30 January 1997     

Reversion of XO to XY sex chromosome mechanism in a phasmid

Summary The sex chromosomes of the male phasmid Isagoras schraderi Rehn comprise an X and a Y, — each with a submedian kinetochore, and one euchromatic and one heterochromatic arm. At meiosis X and Y form an unequal sex bivalent in which the euchromatic arms are terminally associated. Relatively recent reversion from the XO-XX mechanism characteristic of the Phasmidae is indicated by the presence of the euchromatic arm in both X and Y. The diploid number of the male is 34.Unequal autosomal bivalents are found at meiosis in two other species of Isagoras — Isagoras subaquiles Rehn and Isagoras sp. — and in Pseudophasma menius Westwood. The chromosome complements of these species are described.     

An XY sex chromosome mechanism in a mantid with achiasmatic meiosis

An Australian mantid, Ima fusca, with 2n male equals 34, shows achiasmatic meiosis in the male, as in other Australian members of the subfamily Iridopteryginae. It is, however, unique among approximately 104 mantid species that have been studied cytologically, in having an XY sex chromosome mechanism. The X and Y chromosomes are not associated as a bivalent in first metaphase, but arrange themselves opposite one another on the spindle and regularly pass to different poles at first anaphase.     

Evolution of sex chromosomes ZW of Schistosoma mansoni inferred from chromosome paint and BAC mapping analyses

Chromosomes of schistosome parasites among digenetic flukes have a unique evolution because they exhibit the sex chromosomes ZW, which are not found in the other groups of flukes that are hermaphrodites. We conducted molecular cytogenetic analyses for investigating the sex chromosome evolution using chromosome paint analysis and BAC clones mapping. To carry this out, we developed a technique for making paint probes of genomic DNA from a single scraped chromosome segment using a chromosome microdissection system, and a FISH mapping technique for BAC clones. Paint probes clearly identified each of the 8 pairs of chromosomes by a different fluorochrome color. Combination analysis of chromosome paint analysis with Z/W probes and chromosome mapping with 93 BAC clones revealed that the W chromosome of Schistosoma mansoni has evolved by at least four inversion events and heterochromatinization. Nine of 93 BAC clones hybridized with both the Z and W chromosomes, but the locations were different between Z and W chromosomes. The homologous regions were estimated to have moved from the original Z chromosome to the differentiated W chromosome by three inversions events that occurred before W heterohcromatinization. An inversion that was observed in the heterochromatic region of the W chromosome likely occurred after W heterochromatinization. These inversions and heterochromatinization are hypothesized to be the key factors that promoted the evolution of the W chromosome of S. mansoni.     

Diversification of a ZZ/ZW sex chromosome system in Characidium fish (Crenuchidae, Characiformes)

Karyotype and other chromosomal markers of Characidium cf. gomesi were analyzed using conventional (Giemsa-staining, Ag-NOR and C-banding) and molecular (Fluorescent in situ hybridization (FISH) with 18S and 5S rDNA biotinylated probes) techniques. Both sexes had invariably diploid chromosome number 2n = 50 while karyotypes of males and females differed. That of male consisted of 32 metacentric + 18 submetacentric chromosomes and that of female consisted 31 metacentric + 18 submetacentric + 1 subtelocentric chromosomes. The Z chromosome was medium-sized metacentric, while W was highly heterochromatinized subtelocentric element. NORs as revealed by Ag-staining were situated at 2–7 telomeric regions while FISH with 18S probes showed consistently 10 signals at telomeric regions. FISH with 5S rDNA probe showed constantly signals at one metacentric pair. Distribution of centromeric heterochromatin was mostly in all chromosome pairs, besides some telomeric sites. The common origin of the sex chromosome system of ZZ/ZW type in the karyotypes of other representatives of the genus analyzed so far might be hypothesized based on biogeography and partial phylogeny of the group.     

A highly differentiated ZZ/ZW sex chromosome system in a Characidae fish, Triportheus guentheri.

Seventeen specimens of Triportheus guentheri, a fish of the family Characidae, were submitted to chromosomal analysis, with a highly differentiated heteromorphic ZW pair being detected. Chromosome W is much smaller than chromosome Z and mostly heterochromatic. Chromosome Z is the largest in the karyotype, with heterochromatin occurring in the telomeric and centromeric regions only. The W chromosome also varies somewhat in size, the variations being probably due to its long arm. In addition to two other autosomal pairs, chromosome Z shows also an occasional Ag-NOR. Aspects of the ZW system differentiation and of the NOR presence in chromosome Z are discussed. The Characidae family includes a great deal of neotropical freshwater fish species and Triportheus appears to represent the only genus in this family having sex chromosome differentiation at a cytological level.     

Interspecific hybridization between Oryzias latipes and Oryzias curvinotus causes XY sex reversal

The teleost fish, Oryzias curvinotus, is a closely related species to the medaka, Oryzias latipes, and both species have the DMY gene, which is required for male development in O. latipes. It suggests that the molecular function of the DMY gene and the following molecular events of sex differentiation are conserved between these two species. In the present study, we obtained interspecific hybrids between O. curvinotus and O. latipes and demonstrated sex-reversed XY females in the hybrids. The incidence of sex-reversed females in F1 XY hybrids between O. curvinotus females and O. latipes males, and hybrids between O. latipes females and O. curvinotus males were 21% and 100%, respectively. These results indicate that DMY does not always determine maleness in hybrid fish even though it is able to specify normal male development on its native genetic background and suggest that there are some differences between DMY(latipes) and DMY(curvinotus) alleles. Appearance of XY females in F1 hybrids also suggests that an autosomal or X-liked gene(s) from the maternal species interferes in the function of the paternal DMY gene in the male-determining process of the hybrid fish. These hybrid fish would supply a new experimental approach for investigating the genetic and molecular pathway of testis determination and differentiation.     

XX:XY sex chromosome system with X heterochromatinization: an early stage of sex chromosome differentiation in the Neotropic electric eel Eigenmannia virescens

An early stage of sex chromosome differentiation is reported to occur in the electric eel Eigenmannia virescens (Pisces, Sternopygidae) from populations of two tributaries of the Paraná river system (Brazil). Cytogenetic studies carried out in the two populations showed that the Mogi-Gua?u population is characterized by 2n = 38 chromosomes and undifferentiated sex chromosomes and the Tietê population presents 2n = 38 both for males and females and an XX:XY sex chromosome system. The X-chromosome is acrocentric, easily recognized by the presence of a conspicuous heterochromatin block in its distal portion; the Y-chromosome is probably one of the medium sized acrocentrics present in the male karyotype. BrdU induced R-bands of the two populations did not reveal any difference in the euchromatic regions of the chromosomes. AluI and HaeIII restriction enzyme digestion patterns and chromomycin A3 staining of the X-chromosome are presented. The possible role of heterochromatinization in the evolution of sex chromosomes in fish is discussed.     

Molecular cytogenetics and characterization of a ZZ/ZW sex chromosome system in Triportheus nematurus (Characiformes, Characidae)

Chromosomes of Triportheus nematurus, a fish species from family Characidae, were analyzed in order to establish the conventional karyotype, location of C-band positive heterochromatin, Ag-NORs, GC- and AT-rich sites, and mapping of 18S and 5S rDNA with fluorescence in situ hybridization (FISH). The diploid number found was 2n = 52 chromosomes in both males and females. However, the females presented a pair of differentiated heteromorphic chromosomes, characterizing a ZZ/ZW sex chromosome system. The Z chromosome was metacentric and the largest one in the karyotype, bearing C-positive heterochromatin at pericentromeric and telomeric regions. The W chromosome was middle-sized submetacentric, appearing mostly heterochromatic after C-banding and presenting heterogeneous heterochromatin composed of GC- and AT-rich regions revealed by fluorochrome staining. Ag-NORs were also GC-rich and surrounded by heterochromatic regions, being located at the secondary constriction on the short arms of the second chromosome pair, in agreement with 18S rDNA sites detected with FISH. The 18S and 5S rDNA were aligned in tandem, representing an uncommon situation in fishes. The results obtained reinforce the basal condition of the ZZ/ZW sex system in the genus Triportheus, probably arisen prior to speciation in the group.     

ZZ/ZW sex chromosome system in an undescribed species of the genus Apareiodon (Characiformes, Parodontidae)

The chromosomes of an undescribed species of the genus Apareiodon (Characiformes, Parodontidae) from the Verde River, a headwater affluent of the Tibagi River (Paraná State, Brazil), were investigated using conventional Giemsa and Ag stainings, C-banding, CMA(3) fluorescence and fluorescent in situ hybridization (FISH) using 18S and 5S rDNA probes. The diploid chromosome number was 2n = 54, with the karyotype composed of 48 meta/submetacentric and six subtelocentric chromosomes in males, and 47 meta/submetacentric + seven subtelocentric chromosomes in females. The difference is hypothesized to be due to a ZZ/ZW heteromorphic sex chromosome system, a cytotaxonomic characteristic previously observed only in some species of the genus Parodon (family Parodontidae). The presence of similar and/or identical heteromorphic sex chromosome systems might suggest that species of the genera Parodon and Apareiodon bearing ZZ/ZW heteromorphic sex chromosomes likely constitute a monophyletic group, a hypothesis to be tested by a robust phylogeny of the family.     

Structural differences between XX and ZW sex lampbrush chromosomes inRana rugosa females (Anura: Ranidae)

In this study we investigated the morphology and pairing behavior of sex lampbrush chromosomes of XX and ZW females of Rana rugosa from five localities in Japan. Whereas lampbrush chromosomes of XX females from Hiroshima and Isehara had subterminally located centromeres and showed remarkable similarity, those of XX females from Hamakita had the centromeres in the middle. Analysis of landmark configurations revealed that chromosome Xq of Hamakita females closely resembled a part of Xq of Hiroshima and Isehara females, whereas Xp of Hamakita females was inverted compared with the other part of Xq of Hiroshima and Isehara females. Z chromosomes from Kanazawa and Niigata closely resembled the Hiroshima X, whereas the W closely resembled the Hamakita X. XX pairings from Hiroshima, Isehara, and Hamakita were found to be joined by one to four chiasmata at various points all along the axis in both the short and long arms, whereas chromosomal pairs from Kanazawa and Niigata showed only one chiasma between Zp and the distal region of Wq. From these findings we conclude that (1) both the W and the Hamakita X must have evolved from the more primitive Hiroshima and Isehara X chromosomes by a series of pericentric inversions; and (2) females distributed in Hamakita possess two X chromosomes similar to the W, suggesting that either sex-determining or sex-modifying genes on the Hamakita X are clearly different from those on the Kanazawa and Niigata W chromosome. Received: 27 February 1996; in revised form: 22 May 1996 / Accepted: 25 May 1996     

Relationships between breeding soundness and sex drive classifications in beef bulls

Prior to use in single-sire mating trials, ninety-two beef bulls were evaluated to determine the relationships between. Breeding Soundness Examination (BSE) and sex drive assessment categories. A BSE score was given to each bull based on scrotal circumference, spermatozoal motility and morphology. Eighty bulls were classified as satisfactory potential breeders, while 12 were classified as questionable potential breeders. Each bull was exposed to two, 10 min modified libido/serving capacity tests. Sixty-nine bulls were classified as high libido and 23 were classified as medium libido. Differences in scrotal circumference and spermatozoal motility did not influence (P > 0.05) the BSE classifications. However, differences in primary, secondary and total spermatozoal abnormalities were the major factors affecting the BSE classification. Bulls classified as high libido serviced three times more (P < 0.01) during the two test periods than bulls classified as medium libido. High libido bulls were superior in all sex drive traits. However, bulls in the two libido categories did not differ (P > 0.05) in semen quality or scrotal circumference. There was no relationship (P > 0.05) between BSE and sex drive traits (r = -0.16 to 0.24).     

Sex chromatin and sex chromosome systems in nonditrysian Lepidoptera (Insecta)

Eleven representatives of the superorder Amphiesmenoptera (Trichoptera + Lepidoptera) were examined for sex chromatin status. Three species represent stenopsychoid, limnephiloid and leptoceroid branches of the Trichoptera; eight species belong to the primitive, so-called nonditrysian Lepidoptera and represent the infra-orders Zeugloptera, Dacnonypha, Exoporia, Incurvariina, Nepticulina and Tischeriina. The female-specific sex chromatin body was found in the interphase somatic nuclei of Tischeria ekebladella (Bjerkander 1795) (Lepidoptera, Tischeriina). The sex chromatin was absent in all investigated Trichoptera species as well as in all representatives of the nonditrysian Lepidoptera except Tischeria ekebladella . The sex chromosome mechanism of Limnephilus lunatus Curtis 1834 (Trichoptera, Limnephilidae) is Z/ZZ. The sex chromosome mechanism of Tischeria ekebladella (Lepidoptera, Tischeriina) is ZW/ZZ including the W chromosome as the largest element in the chromosome set. The data obtained support the hypothesis that the Z/ZZ sex chromosome system, the female heterogamety and the absence of the sex chromatin body in interphase nuclei are ancestral traits in the superorder Amphiesmenoptera. These ancestral characters are probably kept constant in all the Trichoptera and in the most primitive Lepidoptera. The W sex chromosome and the sex chromatin evolved later in the nonditrysian grade of the Lepidoptera. It is proposed that the sex chromatin is a synapomorphy of Tischeriina and Ditrysia.     

An early stage of ZW/ZZ sex chromosome differentiation in Poecilia sphenops var. melanistica (Poeciliidae,Cyprinodontiformes)

The mitotic and meiotic chromosomes of the American cyprinodont fish Poecilia sphenops var. melanistica were analysed. All 46 chromosomes are telocentric. By specific staining of the constitutive heterochromatin with C-banding and various AT-specific fluorochromes, the homomorphic chromosome pair 1 could be identified as sex chromosomes of the ZW/ZZ type. All female animals exhibit a W chromosome with a large region of telomeric heterochromatin that is not present in the Z chromosome. These sex chromosomes cannot be distinguished by conventional staining; they represent the first demonstration of sex chromosomes in fishes in an early stage of morphological differentiation. The W heterochromatin and the telomeric heterochromatin in the two autosomes 18 show a very bright fluorescence when stained with AT-specific fluorochromes. This allows the direct identification of the chromosomal sex by examining the interphase nuclei: females exhibit three, males only two brightly fluorescent heterochromatic chromocenters in their nuclei. The significance of these ZW/ ZZ sex chromosomes and their specific DNA sequences, the dose compensation of the Z-linked genes, and the experimental possibilities using sex-reversed ZW males are discussed.