Cytologia 41: 559-565, 1976 General Considerations on the Karyotypic Evolution of Chelonia from the Amazon Region of Brazil R. M. Barros,1 M. M. Sampaio,1 M. F. Assis,1 M. Ayres1 and O. R. Cunha2 Received February 8, 1975 In comparison with other orders of Reptilia, there have been very few reports on the chromosomes of Chelonia in the recent literature. Van Brink (1959) described the chromosomes of Emys orbiculares and Chrysemys bellii bellii, both with a diploid number of 50 with macro and microchromosomes. Later, Sasaki and Itoh (1967) described identical karyotypes for Clemmys japonica and Geoclemmys reevesii, with a diploid number of 52, including many microchromosomes. Ayres et al. (1968, 1969) described two kinds of the diploid number, 28 for P. cayennensis, P. expansa, P. unifilis and P. sextuberculata, and 26 for P. dumeriliana. These same numbers were described by Huang and Clark (1969) for P. unifilis and P. expansa. These species do not have microchromosomes, their diploid number being very low in comparison to other species already studied. Sampaio et al. (1969, 1971) described the karyotypes of Geochelone carbonaria and G. denticulata, both with 52 chromo somes. The karyotypes are identical except for no. 12 pair which differed in having pericentric inversion. The karyotype of G. carbonaria is identical to that of Clem mys japonica and Geoclemmys reevesii described by Sasaki and Itoh (1967). Barros et al. (1970, 1972) found 56 chromosomes in the subspecies Kinosternon scorpioides scorpioides and K. s. carajasensis. These two subspecies have a karyotype very similar to that of Geoemyda punctularia punctularia, with the same diploid number. The differences between them could well be explained by pericentric inversions (Bar ros et al. 1973). This paper deals with the chromosomes of the following species of Chelonia (family Chelidae) from the Amazon Region of Brazil: Platemys platicephala, Mesoclemmys gibba and Chelus fimbriatus. Material and methods One male of Platemys platicephala, one female of Mesoclemmys gibba, and one male and two females of Chelus fimbriatus were chromosomally studied. The chro mosome slides were prepared following the method of Gorman et al. (1967). The procedure has already been described in detail by Barros et al. (1972). Results The karyotypes are summarized in Table 1. 1 Centro de Ciencias Biolbgicas , Universidade Federal do Para, Belem, Para, Brasil. 2 Museu Paraense "Emilio Goeldi", Belem, Para, Brasil. 560 R. M. Barros et al. Cytologia 41 Table 1. Karyotypic data of the species studied Platemys platicephala Diploid number: 2n=64. Haploid number: n=32. Number of micro chromosomes : 42. Fundamental number: 64. Karyotype: All chromosomes are acrocentric (Fig. 1). Fig. 1. Male karyotype of Platemys platicephala . Meiosis: The majority of cells in the initial stages of prophase I represent positive heteropycnosis. In diakinesis the larger and medium bivalents show 2 to 3 chiasmata, while the smaller ones have only one chiasma (Figs. 4, d, e, f) . Mesoclemmys gibba Diploid number: 2n=60. Number of microchromosomes: 40. Fundamental number: 66. Karyotype: Pairs no. 1 and 2 are metacentric . The no. 3 pair is submetacentric. Pairs 4 to 11 are acrocentric. Pairs 12 to 30 are microchromosomes , in which the morphology is difficult to ascertain (Fig. 2). 1976 General Considerations on the Karyotipic Evolution of Chelonia 561 Chelus fimbriatus Diploid number: 2n=50. Haploid number: n=25. Number of micro chromosomes: 30. Fundamental number: 64. Fig. 2. Female karyotype of Mesoclemmys gibba. Fig. 3. Female karyotype of Chelus fimbriatus. 562 R. M. Barros et al. Cytologia 41 Karyotype: Pairs 1, 2, 6 and 9 are metacentric. Pairs 3, 4 and 5 are subme tacentric. Pairs 7, 8 and 10 are acrocentric. Pairs 11 to 25 are represented by microchromosomes (Fig. 3). Meiosis: The majority of cells in the pachytene stage exhibit positive hetero pycnosis, and one bivalent which showed a region without pairing. Larger bivalents with 3 to 6 chiasmata are observable in diakinesis, those of medium-size have two chiasmata while the smaller ones have one chiasma (Figs. 4, a, b, c). Fig. 4. A, metaphase II of Chelus fimbriatus, male. B, pachytene of Chelus fimbriatus, male. Arrow points to a region where does not occur pairing. C, diakinesis of Chelus fimbriatus, male. D, diaki nesis of Platemys platicephala, male. E, pachytene of Platemys platicephala, male, showing posi tive heteropycnosis. F, metaphase II of Platemys platicephala, male. Discussion 1. Karyotype variation The family Chelidae shows a considerable karyotypic variation. The species, Platemys platicephala, has the largest diploid number (64) with the greatest number of microchromosomes (42); the chromosomes of this species are all acrocentric. 1976 General Considerations on the Karyotipic Evolution of Chelonia 563 Mesoclemmys gibba has 60 chromosomes and medium-sized chromosomes are all acrocentric. Chelus fimbriatus, with 50 chromosomes, has the smallest diploid number among the three species studied having 30 microchromosomes, and meta centric and acrocentric medium-sized chromosomes. Chelus fimbriatus has a funda mental number identical to that of Platemys platicephala. Robertsonian fission fusion mechanism could be responsible to the difference occurring between these two species. There seem to be a relationship between the diploid number, the number of me tacentric and acrocentric chromosomes and microchromosomes of these three species of the Chelidae. Karyotypes with a low diploid number have more metacentric and less acrocen tric chromosomes and microchromosomes, than karyotypes with a high diploid number. The relationship may be the result of centric fusions between the acro centric chromosomes, resulting in an increased number of metacentric and a re duction of the diploid number. Furthermore, if the decreasing number of micro chromosomes is related to an increase of metacentric chromosomes, it could be argued that a process of centric fusions occurred among the larger microchromo somes and a few of the acrocentric chromosomes. On the other hand, the decrease in the number of microchromosomes could be resulted from their gradual elimina tion (Nogusa 1953). Interesting is the fact that the species of Podocnemis with a low diploid number and many metacentric, have no microchromosomes (Ayres et al. 1968, 1969). In P. dumeriliana which has the lowest diploid number over found in Chelonia (26), all the chromosomes are metacentric. These findings suggest that, in general, the species with many metacentric chromosomes are characterized by a low number of micro chromosomes. The cytogenetics data of Chelonia seem to indicate the occurrence of three groups of karyotypes: Group A-Karyotypes with a high diploid number (60-64), having many acrocentric chromosomes, few or no metacentric and many micro chromosomes. The species, Mesoclemmys gibba and Platemys platicephala, belong to this group. Group B-Karyotypes with a slightly lower diploid number (50-56) having many metacentric and few acrocentric chromosomes. In this group there are fewer microchromosomes than group A. In this group fall the following species, Chelus fimbriatus, Kinosternon scorpioi des scorpioides, Kinosternon scorpioides carajasensis, Geoemyda punctularia punctularia, and two species of genus Geochelone. Group C-Karyotypes with a low diploid number (26-28) characterized by few or no acrocentric many metacentric and an absence of micro chromosomes. The species of genus Podocnemis belong to this group. On the above basis it is apparent that centric fusions or elimination of micro chromosomes or both, are a principal mechanism involved in the evolution of chelo nian karyotypes. 564 R. M. Barros et al. Cytologia 41 2. Meiotic aspects and the mechanism of sex differentiation Chelonia presumably do not have heteromorphic sex chromosomes. This has been, however, a point of contention amongst cytologists. Many papers have indicated sex determination of XO type in females: Oguma (1934) in Amyda japonica; Nakamura (1935, 1949) in Caretta caretta olivacea and Clemmys japonica, Susuki (1950) in Amyda maaki, and Makino (1952) in Chelonia japonica. In these species the sex chromosomes would be represented by micro chromosomes. On the other hand, Matthey (1931, 1957) and van Brink (1959) did not find sex chromosome differentiation in Emys orbiculares. Similarly in the genus Podocnemis (Ayres et al. 1968, 1969; Huang and Clark 1969), Geochelone (Sampaio et al. 1969, 1971), Kinosternon (Barros et a1. 1970, 1972) and Geoemyda (Barros et al. 1973), sexual heteromorphism was not shown to occur. More recent data indicate that sexual chromosomic differences do not occur in the Chelonia. In the present study, where both males and females were studied (Chelus fim briatus), there was no difference between the chromosomic complements of the two sexes. However, the meiotic specimens of Platemys platicephala showed heterochro matic regions in the initial stages of prophase I. In Chelus fimbriatus, besides heterochromatin, one bivalent showed a region without pairing in the pachytene stage. Similar autosomic heterochromatic regions are present in two subspecies of the genus Kinosternon (Barros et al. 1970, 1972). In the meiotic cells of these subspecies, there were areas of partial or total condensation which were asynchronous in some bivalents, and in the diplotene there was one bivalent which showed a typical early behavior in relation to the others. Heteropycnosis and bivalents with asynchronous behavior in