_??_1992 The Japan Mendel Society Cytologia 57: 369-372, 1992

ChromosomeStudies in Hypoptopomatinae (Pisces,Siluriformes, ): I. XX/XYSex ChromosomeHeteromorphism in tietensis

Artur Antonio Andreata1, Lurdes Foresti de Almeida-Toledo1, Claudio Oliveira2 and Silvio de Almeida Toledo Filho1 1 Departamento de Biologia, Instituto de Biociencias,USP, Sao Paulo, Caixa Postal 11.461,05499, Sao Paulo, SP, Brasil 2Departamento de Morfologia, Instituto de Biociencias,UNESP, Campus de Botucatu, Brasil. AcceptedApril 13, 1992

The subfamily Hypoptopomatinae belongs to family Loricariidae, one of the largest in number of of Neotropical freshwater fishes. The chromosome numbers reported thus far for Loricariidae are restricted to the subfamilies Hypostominae and and are highly variable, ranging from 2n=44 to 2n=74 (Oliveira et al. 1988). Two cases sugges ting the occurrence of a XX/XY sex chromosome system were described for species of family Loricariidae (Hypostomus ancistroides and H. macrops), both cited as belonging to the Plecostomus by Michele et al. (1977). These determinations were done using the Giemsa staining technique and were not confirmed by other techniques. Pseudotocinclus is a monotypic genus of Hypoptopomatinae and its only species, P. tie tensis, is endemic to the headwaters of the Tiete river (Britski and Garavello 1984). The present study reports the karyotype, C-banding pattern and the location of nucleous organizer regions (NORs) of P. tietensis which presents heteromorphic sex chromosomes.

Materials and methods

The analyses were done on preparations obtained from 13 specimens (9 males and 4 females) of Pseudotocinclus tietensis collected in the headwaters of the Tiete river at Para napiacaba, Sao Paulo, Brazil. The specimens were identified and deposited in the Museu de Zoologia da Universidade de Sao Paulo (MZUSP), Sao Paulo, Brazil. Mitotic chromosome preparations were obtained from kidney and gill cells using the air drying technique (Oliveira et al. 1988b); 371 cells were analysed. Chromosome morphology was determined on the basis of arm ratio, as proposed by Levan et al. (1964), and chromosomes were classified as metacentrics (M), submetacentrics (SM) and subtelocentrics (ST). C-band ing was performed by the method of Sumner (1972) and the NOR silver staining by the method of Howell and Black (1980).

Results

P. tietensis has 2n=54 chromosomes for both sexes. The basic karyotypic constitution is 26M+20SM+6ST, plus one heteromorphic pair (SM/M) in males (Fig. la) and one ho momorphic pair (SM/SM) in females (Fig. lb). Ag-NORs are located interstitially on the long arm of the largest metacentric pair (Fig. 1a).

1 Correspondence to Lurdes Foresti de Almeida-Toledo. 370 Artur Antonio Andreata, et aL. Cytologia 57

A small amount of constitutive heterochromatin is present in the karyotypes of males and females (pairs 1 and 10, Fig. 2); the region corresponding to the NOR in pair 1 is C-band negative. The occurrence of a heteromorphic pair in the males is demonstrable by the presence of one extra C-banded block in the Y chromosome (Fig. 2a).

Fig. 1. Giemsa stained karyotypes of Pseudotocinclus tietensis: a) male; b) female. In the inset, the NOR-bearing chromosome pair stained by the Ag-NOR technique. Bar= 10ƒÊm.

Fig. 2. C-band stained karyotypes of Pseudotocinclus tietensis: a) male; b) female.

Discussion

The description of the karyotypic constitution of Pseudotocinclus tietensis is the first for 1992 XX/XY Sex Chromosomes in Pseudotocinclus 371 the subfamily Hypoptopomatinae. The results show that this species has 2n=54 chromo somes (Fig. 1), one chromosome pair with NOR (Fig . 1a) and small amount of constitutive heterochromatin (Fig. 2). The occurrence of one chromosome pair with NOR is very common among fishes (Al meida-Toledo and Foresti 1985) but only few species have interstitial NORs . The karyotypes of males and females of Pseudotocinclus tietensis differ by the presence of a C-band positive block present only in males (Fig. 2). The presence of constitutive hetero chromatin in sex chromosomes is a widespread phenomenon among vertebrates. In general , the Y chromosome of mammals and W chromosome of birds and snakes are almost entirely heterochromatic and rich in satellite DNA (Hsu and Arrighi 1971, Singh et al. 1976). Cases of cytologically recognizable sex chromosomes in fishes have been identified thus far in about 60 species (Chourrout 1986), but in a very few cases the differential presence of heterochromatin blocks in one of the sexes was reported to involve male or female heterogamety and simple or multiple systems (Haaf and Schmid 1984, Phillips and Ihssen 1985, Galleti Jr. and Foresti 1986, Almeida-Toledo et al. 1988). Nanda et al. (1990) demonstrated by hy bridizaton of restriction enzyme-digested genomic DNA with the oligonucleotide probe (GACA)4 that simple repetitive sequences are associated with differentiation of sex chro mosomes in the guppy fish. According to extensive studies on the sex chromosomes of snakes, by Singh et al. (1976, 1980), the primary step in the differentiation between the Z and W chromosomes in this group was the development of highly repetitive DNA sequences in the W chromosome. The presence of a heteromorphic region results in restriction in the process of meiotic recombina tion between the originally homologous chromosomes, isolating a region that includes genes involved in sex determination. Thus, constitutive heterochromatin may represent an effective form of crossing-over restriction in the sex chromosomes. The presence of one heteromorphic chromosome pair in P. tietensis males and of a homomorphic pair in the females due to the presence of a heteromorphic block may be interpreted as an initial process of differentiantion of an XX/XY sex chromosome system in this species. Further studies on other species of Hypoptopomatinae are needed to reveal if the sex chromosome system is limited to P. tietensis or commonly occurs in this group, and the use of other techniques will permit a better characterization of these sex chromosomes.

Summary

The cytogenetic analysis of 13 specimens (9 males and 4 females) of Pseudotocinclus tietensis showed that this species has 2n=54 chromosomes (26M+20SM+6ST) plus one heteromorphic pair (SM/M) in the males and one homomorphic pair (SM/SM) in the females. Ag-NORs are located interstitially on the long arm of the largest metacentric pair and a small amount of constitutive heterochromatin is present in the karyotypes. The occurrence of a heteromorphic pair in the males is demonstrable by the presence of one extra C-banded block in the Y chromosome. The presence of one heterochromatin block distinguishing the sexual chromosomes suggests that this species presents a very primitive mechanism of sex chromo some differentiation.

Key words: Karyotype, NOR, C-band, XX/XY sex chromosomes, Pseudotocinclus

Acknowledgments

The authors are grateful to Dr. Heraldo A. Britski for taxonomic identification of the 372 Artur Antonio Andreata, et al. Cytologia 57 specimens, and to Mrs. Cristina Barnabe and Mirian Romeo for technical assistance. Funds supporting this study were provided by the FAPESP, CNPq, CAPES and FINER

References

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