Evolution of ancient satellite DNAs in extant alligators and caimans (Crocodylia, Reptilia).

BACKGROUND: Crocodilians are one of the oldest extant vertebrate lineages, exhibiting a combination of evolutionary success and morphological resilience that has persisted throughout the history of life on Earth. This ability to endure over such a long geological time span is of great evolutionary importance. Here, we have utilized the combination of genomic and chromosomal data to identify and compare the full catalogs of satellite DNA families (satDNAs, i.e., the satellitomes) of 5 out of the 8 extant Alligatoridae species. As crocodilian genomes reveal ancestral patterns of evolution, by employing this multispecies data collection, we can investigate and assess how satDNA families evolve over time. RESULTS: Alligators and caimans displayed a small number of satDNA families, ranging from 3 to 13 satDNAs in A. sinensis and C. latirostris, respectively. Together with little variation both within and between species it highlighted long-term conservation of satDNA elements throughout evolution. Furthermore, we traced the origin of the ancestral forms of all satDNAs belonging to the common ancestor of Caimaninae and Alligatorinae. Fluorescence in situ experiments showed distinct hybridization patterns for identical orthologous satDNAs, indicating their dynamic genomic placement. CONCLUSIONS: Alligators and caimans possess one of the smallest satDNA libraries ever reported, comprising only four sets of satDNAs that are shared by all species. Besides, our findings indicated limited intraspecific variation in satellite DNA, suggesting that the majority of new satellite sequences likely evolved from pre-existing ones.

Multiple karyotype differences between populations of the Hoplias malabaricus (Teleostei; Characiformes), a species complex in the gray area of the speciation process.

Neotropical fishes exhibit remarkable karyotype diversity, whose evolution is poorly understood. Here, we studied genetic differences in 60 individuals, from 11 localities of one species, the wolf fish Hoplias malabaricus, from populations that include six different "karyomorphs". These differ in Y-X chromosome differentiation, and, in several cases, by fusions with autosomes that have resulted in multiple sex chromosomes. Other differences are also observed in diploid chromosome numbers and morphologies. In an attempt to start understanding how this diversity was generated, we analyzed within- and between-population differences in a genome-wide sequence data set. We detect clear genotype differences between karyomorphs. Even in sympatry, samples with different karyomorphs differ more in sequence than samples from allopatric populations of the same karyomorph, suggesting that they represent populations that are to some degree reproductively isolated. However, sequence divergence between populations with different karyomorphs is remarkably low, suggesting that chromosome rearrangements may have evolved during a brief evolutionary time. We suggest that the karyotypic differences probably evolved in allopatry, in small populations that would have allowed rapid fixation of rearrangements, and that they became sympatric after their differentiation. Further studies are needed to test whether the karyotype differences contribute to reproductive isolation detected between some H. malabaricus karyomorphs.

Karyotypic changes and diversification time in Epinephelidae groupers (Perciformes). Implications on reproductive isolation.

Groupers (Epinephelidae and Serranidae) have attracted special attention to fish farming, and their species offer good opportunities for successful hybridizations. Cytogenetic data allow a better understanding of the role of karyotypic diversification in the acquisition of post-zygotic reproductive isolation (RI). Thus, chromosomal analyses were performed on E. striatus (Caribbean Sea), E. coioides and E. tauvina (Indo-Pacific Region), using standard procedures and mapping of six repetitive DNA classes by the in situ hybridization. The three species have 2n=48 chromosomes. The karyotypes of E. coioides and E. striatus are composed only of acrocentric chromosomes (FN=48), while E. tauvina has 8 submetacentric chromosomes (FN=56). Heterochromatin has a preferential centromeric distribution, and the microsatellite repeats are dispersed throughout the chromosomes of all species. The 18S and 5S rDNA sites are unique but show a colocalization arrangement in E. tauvina and E. striatus. The chromosomal organization suggests that the three species still maintain a significant amount of syntenic regions. The range of the karyotype divergence and the RI levels showed low, but goes turn proportionally greater in relation to the divergence time between the parental species. The slow acquisition of postzygotic RI is consistent with the high karyotype homogeneity presented by Epinephelidae family.

Cytogenetics Meets Genomics: Cytotaxonomy and Genomic Relationships among Color Variants of the Asian Arowana Scleropages formosus.

Scleropages formosus (Osteoglossiformes, Teleostei) represents one of the most valued ornamental fishes, yet it is critically endangered due to overexploitation and habitat destruction. This species encompasses three major color groups that naturally occur in allopatric populations, but the evolutionary and taxonomic relationships of S. formosus color varieties remain uncertain. Here, we utilized a range of molecular cytogenetic techniques to characterize the karyotypes of five S. formosus color phenotypes, which correspond to naturally occurring variants: the red ones (Super Red); the golden ones (Golden Crossback and Highback Golden); the green ones (Asian Green and Yellow Tail Silver). Additionally, we describe the satellitome of S. formosus (Highback Golden) by applying a high-throughput sequencing technology. All color phenotypes possessed the same karyotype structure 2n = 50 (8m/sm + 42st/a) and distribution of SatDNAs, but different chromosomal locations of rDNAs, which were involved in a chromosome size polymorphism. Our results show indications of population genetic structure and microstructure differences in karyotypes of the color phenotypes. However, the findings do not clearly back up the hypothesis that there are discrete lineages or evolutionary units among the color phenotypes of S. formosus, but another case of interspecific chromosome stasis cannot be excluded.

Chromosomal Analysis of Ctenolucius hujeta Valenciennes, 1850 (Characiformes): A New Piece in the Chromosomal Evolution of the Ctenoluciidae.

Ctenoluciidae (Characiformes), a family of freshwater fishes, comprises 2 genera, Ctenolucius and Boulengerella, with 7 recognized species. Up to now, only species of the genus Boulengerella have been subjected to cytogenetic studies. Here, we investigated the karyotype and other cytogenetic features of pike characin, Ctenolucius hujeta, using conventional (Giemsa staining, C-banding, Ag-NOR staining) and molecular (rDNA, telomeric sequences, and fiber-FISH mapping) procedures. This species has a diploid chromosome number of 2n = 36, and a karyotype composed of 12m + 20sm + 4a and FN = 68, similar to that found in Boulengerella species. However, differences regarding the number and distribution of several chromosomal markers support a distinct generic status. Colocalization of the 18S and 5S rDNA genes is an exclusive characteristic of the C. hujeta genome, with an interspersed distribution in the chromosomal fiber, an unusual phenomenon among eukaryotes. Additionally, our results support the view that Ctenoluciidae and Lebiasinidae families are closely related.

Cytogenetic Analysis of Panaqolus tankei Cramer & Sousa, 2016 (Siluriformes, Loricariidae), an Ornamental Fish Endemic to Xingu River, Brazil.

Despite conservation of the diploid number, a huge diversity in karyotype formulae is found in the Ancistrini tribe (Loricariidae, Hypostominae). However, the lack of cytogenetic data for many groups impairs a comprehensive understanding of the chromosomal relationships and the impact of chromosomal changes on their evolutionary history. Here, we present for the first time the karyotype of Panaqolus tankei Cramer & Sousa, 2016. We focused on the chromosomal characterization, using conventional and molecular cytogenetic techniques to unravel the evolutionary trends of this tribe. P. tankei, as most species of its sister group Pterygoplichthini, also possessess a conserved diploid number of 52 chromosomes. We observed heterochromatin regions in the centromeres of many chromosomes; pairs 5 and 6 presented interstitial heterochromatin regions, whereas pairs 23 and 24 showed extensive heterochromatin regions in their q arms. In situ localization of 18S rDNA showed hybridization signals correlating with the nucleolus organizer regions, which are located in the q arms of pair 5. However, the 5S rDNA was detected in the centromeric and terminal regions of the q arms of pair 8. (TTAGGG)n hybridized only in the terminal regions of all chromosomes. Microsatellite in situ localization showed divergent patterns, (GA)15 repeated sequences were restricted to the terminal regions of some chromosomes, whereas (AC)15 and (GT)15 showed a scattered hybridization pattern throughout the genome. Intraspecific comparative genomic hybridization was performed on the chromosomes of P. tankei to verify the existence of sex-specific regions. The results revealed only a limited number of overlapping hybridization signals, coinciding with the heterochromatin in centromeric regions without any sex-specific signals in both males and females. Our study provides a karyotype description of P. tankei, highlighting extensive differences in the karyotype formula, the heterochromatin regions, and sites of 5S and 18S rDNA, as compared with data available for the genus.

Comparative Cytogenetics in Four Leptodactylus Species (Amphibia, Anura, Leptodactylidae): Evidence of Inner Chromosomal Diversification in Highly Conserved Karyotypes.

With 82 species currently described, the genus Leptodactylus is the most diverse and representative one in the family Leptodactylidae. Concerning chromosomal organization, this genus represents an interesting and underexplored group since data from molecular cytogenetics are incipient, and little is known about the organization and distribution of repetitive DNA elements in the karyotypes. In this sense, this study aimed at providing a comparative analysis in 4 Leptodactylus species (L. macrosternum, L. pentadactylus, L. fuscus, and Leptodactylus cf. podicipinus), combining conventional cytogenetics (Giemsa staining, C-banding, and AgNOR staining) and mapping of molecular markers (18S rDNA, telomeric and microsatellite probes), to investigate mechanisms underlying their karyotype differentiation process. The results showed that all species had karyotypes with 2n = 22 and FN = 44, except for Leptodactylus cf. podicipinus which presented FN = 36. The 18S rDNA was observed in pair 8 of all analyzed species (corresponding to pair 4 in L. pentadactylus), coinciding with the secondary constrictions and AgNOR staining. FISH with microsatellite DNA probes demonstrated species-specific patterns, as well as an association of these repetitive sequences with constitutive heterochromatin blocks and ribosomal DNA clusters, revealing the dynamics of microsatellites in the genome of the analyzed species. In summary, our data demonstrate an ongoing process of genomic divergence inside species with almost similar karyotype, driven most likely by a series of pericentric inversions, followed by differential accumulation of repetitive sequences.

Chromosomal Evolution in Aspredinidae (Teleostei, Siluriformes): Insights on Intra- and Interspecific Relationships with Related Groups.

The family Aspredinidae comprises a clade of complex systematic relationships, both from molecular and morphological approaches. In this study, conventional and molecular cytogenetic studies coupled with nucleotide sequencing were performed in 6 Aspredininae species (Amaralia hypsiura, Bunocephalus cf. aloikae, Bunocephalus amaurus, Bunocephalus aff. coracoideus, Bunocephalus verrucosus, and Platystacus cotylephorus) from different locations of the Amazon hydrographic basin. Our results showed highly divergent diploid numbers (2n) among the species, ranging from 49 to 74, including the occurrence of an XX/X0 sex chromosome system. A neighbor-joining phylogram based on the cytochrome c oxidase I (COI) showed that Bunocephalus coracoideus is not a monophyletic clade, but closely related to B. verrucosus. The karyotypic data associated with COI suggest an ancestral karyotype for Aspredinidae with a reduced 2n, composed of bi-armed chromosomes and a trend toward chromosomal fissions resulting in higher diploid number karyotypes, mainly composed of acrocentric chromosomes. Evolutionary relationships were discussed under a phylogenetic context with related species from different Siluriformes families. The karyotype features and chromosomal diversity of Aspredinidae show an amazing differentiation, making this family a remarkable model for investigating the evolutionary dynamics in siluriforms as well as in fish as a whole.

Molecular Cytogenetic Analysis in Freshwater Prawns of the Genus Macrobrachium (Crustacea: Decapoda: Palaemonidae).

Freshwater prawns of the genus Macrobrachium are one of the important components of circumtropical marine, estuarine, and freshwater environments. They have been extensively exploited for human consumption for many years. More than 250 species reflect the evolutionary success of this highly diversified group, with a complex and challenging taxonomy due to morphological variations and vast geographical distribution. Although genetic approaches have been used to clarify phylogenetic and taxonomic aspects of Macrobrachium species, cytogenetic information is still very scarce and mostly focused on chromosome number and morphology. Here, we present chromosome data for three species from the Neotropical region, M. carcinus, M. acanthurus, and M. amazonicum, and one species from the Oriental region, M. rosenbergii. Using conventional cytogenetic approaches and chromosome mapping of repetitive DNAs by fluorescence in situ hybridization (FISH), we identified numerical diversification of the diploid set, within and between both zoogeographic regions. These included M. acanthurus and M. amazonicum sharing diploid chromosomes of 98, while M. carcinus has 94, and M. rosenbergii has 118 chromosomes. Argentophilic sites are also variable in number, but they occur in a much higher number than 18S rDNA, representing two to 10 sites within the study species. Microsatellites repeat motifs are also abundant in the chromosomes, with a co-localization and uniform distribution along the chromosome arms, but completely absent in the AT-rich centromeric regions. As a whole, our study suggests that the 2n divergence was followed by a considerable rDNA diversification. The abundance of the exceptional amount of microsatellite sequences in the chromosomes also suggests that they are essential components of the Macrobrachium genome and, therefore, maintained as a shared feature by the species, the reason for which is yet unknown.

Paracentric Inversions Differentiate the Conservative Karyotypes in Two Centropomus Species (Teleostei: Centropomidae).

Centropomus is the sole genus of the Centropomidae family (Teleostei), comprising 12 species widely distributed throughout the Western Atlantic and Eastern Pacific, with 6 of them occurring in the Western Atlantic in extensive sympatry. Their life history and phylogenetic relationships are well characterized; however, aspects of chromosomal evolution are still unknown. Here, cytogenetic analyses of 2 Centropomus species of great economic value (C. undecimalis and C. mexicanus) were performed using conventional (Giemsa, Ag-NOR, and fluorochrome staining, C- and replication banding) and molecular (chromosomal mapping of 18S and 5S rDNA, H2A-H2B and H3 hisDNA, and (TTAGGG)n repeats) approaches. The karyotypes of both species were composed of 48 solely acrocentric chromosomes (2n = 48; FN = 48), but the single ribosomal site was located in varying positions in the long arms of the second largest chromosome pair. Replication bands were generally similar, although conspicuous differences were observed in some chromosome regions. In both species, the histone H3 genes were located on 3 apparently homeologous chromosome pairs, but the exact position of these clusters differed slightly. Interspecific hisDNA and rDNA site displacements can indicate the occurrence of multiple paracentric inversions during the evolutionary diversification of the Centropomus genomes. Although the karyotypes remained similar in both species, our data demonstrate an unsuspected microstructural reorganization between them, driven most likely by a series of paracentric inversions.

Evolution of Bird Sex Chromosomes Narrated by Repetitive Sequences: Unusual W Chromosome Enlargement in Gallinula melanops (Aves: Gruiformes: Rallidae).

Among birds, species with the ZZ/ZW sex determination system generally show significant differences in morphology and size between the Z and W chromosomes (with the W usually being smaller than the Z). In the present study, we report for the first time the karyotype of the spot-flanked gallinule (Gallinula melanops) by means of classical and molecular cytogenetics. The spot-flanked gallinule has 2n = 80 (11 pairs of macrochromosomes and 29 pairs of microchromosomes) with an unusual W chromosome that is larger than the Z. Besides being totally heterochromatic, it has a secondary constriction in its long arm corresponding to the nucleolar organizer region, as confirmed by both silver staining and mapping of 18S rDNA probes. This is an unprecedented fact among birds. Additionally, 18S rDNA sites were also observed in 6 microchromosomes, while 5S rDNA was found in just 1 microchromosomal pair. Seven out of the 11 used microsatellite sequences were found to be accumulated in microchromosomes, and 6 microsatellite sequences were found in the W chromosome. In addition to the involvement of heterochromatin and repetitive DNAs in the differentiation of the large W chromosome, the results also show an alternative scenario that highlights the plasticity that shapes the evolutionary history of bird sex chromosomes.

Karyotype Evolution and Distinct Evolutionary History of the W Chromosomes in Swallows (Aves, Passeriformes).

As in many other bird groups, data on karyotype organization and distribution of repetitive sequences are also lacking in species belonging to the family Hirundinidae. Thus, in the present study, we analyzed the karyotypes of 3 swallow species (Progne tapera, Progne chalybea, and Pygochelidon cyanoleuca) by Giemsa and AgNOR staining, C-banding, and FISH with 11 microsatellite sequences. The diploid chromosome number was 2n = 76 in all 3 species, and NORs were observed in 2 chromosome pairs each. The microsatellite distribution pattern was similar in both Progne species, whereas P. cyanoleuca presented a distinct organization. These repetitive DNA sequences were found in the centromeric, pericentromeric, and telomeric regions of the macrochromosomes, as well as in 2 interstitial blocks in the W chromosome. Most microchromosomes had mainly telomeric signals. The Z chromosome displayed 1 hybridization signal in P. tapera but none in the other species. In contrast, the W chromosome showed an accumulation of different microsatellite sequences. The swallow W chromosome is larger than that of most Passeriformes. The observed enlargement in chromosome size might be explained by these high amounts of repetitive sequences. In sum, our data highlight the significant role that microsatellite sequences may play in sex chromosome differentiation.

Evolutionary Relationships among Boulengerella Species (Ctenoluciidae, Characiformes): Genomic Organization of Repetitive DNAs and Highly Conserved Karyotypes.

Ctenoluciidae is a Neotropical freshwater fish family whose representatives are known as bicudas. The genus Boulengerella contains 5 species, and 4 of them (B. cuvieri, B. lateristriga, B. lucius, and B. maculata) were cytogenetically analyzed in the present study by conventional and molecular procedures. All 4 species have a very similar karyotype, with 2n = 36 chromosomes (14 metacentrics + 16 submetacentrics + 6 subtelocentrics; FN = 72). However, the heterochromatin distribution pattern is species-specific. In all 4 species, the nucleolus organizer region is located in pair 18, as also confirmed by cytogenetic mapping of 18S rDNA. In turn, 5S rRNA genes are present in 2 chromosome pairs: in pair 1 of all 4 species, and in pair 10 of B. lateristriga, B. maculata, and B. cuvieri, but in pair 4 of B. lucius. The telomeric probe highlighted terminal regions in all chromosomes, as well as an interstitial centromeric sequence in pair 3 of the 3 first-mentioned species. Notably, a conspicuous heteromorphic secondary constriction in chromosomes 18 was found only in the males of the 3 species, rendering one of the homologs much larger than the other one. This feature, associated with a large 18S rDNA block and accumulation of telomeric sequences, suggests the presence of an XX/XY sex chromosome system in the analyzed Boulengerella species.

Genomic Organization of Repetitive DNAs and Differentiation of an XX/XY Sex Chromosome System in the Amazonian Puffer Fish, Colomesus asellus (Tetraodontiformes).

The genus Colomesus is the sole representative of the family Tetraodontidae in the Amazon region. Here, Colomesus asellus was analyzed using conventional and molecular cytogenetic protocols. Its diploid chromosome number is 2n = 46 with 12 meta-, 10 submeta-, 16 subtelo-, and 8 acrocentric chromosomes and a fundamental number of FN = 84. An XX/XY sex chromosome system was identified. Mapping of 18S rDNA correlated with the nucleolus organizer regions (Ag-NORs) in the short arms of the 2 X chromosomes in females and in the Y chromosome in males. C-banding revealed heterochromatin in the centromeric regions of all chromosomes, except for pair 3. Prominent sex chromosome-specific heterochromatin amplification was observed, covering the short arms of the Y chromosome almost entirely. FISH with telomeric and tropomyosin (tpm1) sequences, respectively, revealed terminal signals in all chromosomes. The analysis of extended DNA fibers confirmed the colocalization and the interspersed pattern of the telomeric and tpm1 sequences. Thus, this study highlights the remarkable evolutionary dynamism presented by the Amazonian puffer fish regarding the differentiation of a heteromorphic XY sex chromosome system and a particular sex-specific amplification of rDNA sites. This is the first record of such an association in the Tetraodontidae family.

Chromosomal Mapping of Repetitive DNAs in Myiopsitta monachus and Amazona aestiva (Psittaciformes, Psittacidae) with Emphasis on the Sex Chromosomes.

Here, for the first time, we describe the karyotype of Myiopsitta monachus (Psittacidae, Arini). We found 2n = 48, corresponding to the lowest diploid number observed in Neotropical Psittaciformes so far, with an uncommonly large W chromosome homomorphic to the Z. In order to better understand the evolution of the sex chromosomes in this species, we applied several molecular cytogenetic approaches, including C-banding, FISH mapping of repetitive DNAs (several microsatellite repeats), and whole-chromosome painting on metaphases of M. monachus. For comparison, another species belonging to the same tribe but with a smaller W chromosome (A. aestiva) was also analyzed. The results show that the constitutive heterochromatin has a very diverse distribution pattern in these species revealing heterochromatic blocks in the centromeric region of all chromosomes and in most of the length of the W chromosome in A. aestiva, while in M. monachus they were found in interstitial and telomeric regions. Concerning the microsatellites, only the sequence (CG)n produced signals on the W chromosome of A. aestiva, in the distal region of both arms. However, in M. monachus, (CAA)n, (CAG)n, and (CG)n probes were accumulated on the W chromosome, and, in addition, the sequence (CAG)n also hybridized to heterochromatic regions in macrochromosomes, as well as in microchromosomes. Based on these results, we suggest that the increase in length of the W chromosome in M. monachus is due to the amplification of repetitive elements, which highlights their significant role in the evolutionary process of sex chromosome differentiation.

Karyotype and Mapping of Repetitive DNAs in the African Butterfly Fish Pantodon buchholzi, the Sole Species of the Family Pantodontidae.

The monophyletic order Osteoglossiformes represents one of the most ancestral groups of teleosts and has at least 1 representative in all continents of the southern hemisphere, with the exception of Antarctica. However, despite its phylogenetic and biogeographical importance, cytogenetic data in Osteoglossiformes are scarce. Here, karyotype and chromosomal characteristics of the lower Niger River population of the African butterfly fish Pantodon buchholzi, the sole species of the family Pantodontidae (Osteoglossiformes), were examined using conventional and molecular cytogenetic approaches. All specimens examined had 2n = 46 chromosomes, with a karyotype composed of 5 pairs of metacentric, 5 pairs of submetacentric, and 13 pairs of acrocentric chromosomes in both sexes. No morphologically differentiated sex chromosomes were identified. C-bands were located in the centromeric/pericentromeric region of all chromosomes and were associated with the single AgNOR site. FISH with ribosomal DNA probes revealed that both 5S and 18S rDNA were present in only 1 pair of chromosomes each, but did not colocalize. CMA3+ bands were observed near the telomeres in several chromosome pairs and also at the 18S rDNA sites. The mapping of di- and trinucleotide repeat motifs, Rex6 transposable element, and U2 snRNA showed a scattered distribution over most of the chromosomes, but for some microsatellites and the U2 snRNA also a preferential accumulation at telomeric regions. This study presents the first detailed cytogenetic analysis in the African butterfly fish by both conventional and molecular cytogenetic protocols. This is the first of a series of further cytogenetic and cytogenomic studies on osteoglossiforms, aiming to comprehensively examine the chromosomal evolution in this phylogenetically important fish order.

Hypermethylated Chromosome Regions in Nine Fish Species with Heteromorphic Sex Chromosomes.

Sites and amounts of 5-methylcytosine (5-MeC)-rich chromosome regions were detected in the karyotypes of 9 Brazilian species of Characiformes fishes by indirect immunofluorescence using a monoclonal anti-5-MeC antibody. These species, belonging to the genera Leporinus, Triportheus and Hoplias, are characterized by highly differentiated and heteromorphic ZW and XY sex chromosomes. In all species, the hypermethylated regions are confined to constitutive heterochromatin. The number and chromosome locations of hypermethylated heterochromatic regions in the karyotypes are constant and species-specific. Generally, heterochromatic regions that are darkly stained by the C-banding technique are distinctly hypermethylated, but several of the brightly fluorescing hypermethylated regions merely exhibit moderate or faint C-banding. The ZW and XY sex chromosomes of all 9 analyzed species also show species-specific heterochromatin hypermethylation patterns. The analysis of 5-MeC-rich chromosome regions contributes valuable data for comparative cytogenetics of closely related species and highlights the dynamic process of differentiation operating in the repetitive DNA fraction of sex chromosomes.

Unusual dispersion of histone repeats on the whole chromosomal complement and their colocalization with ribosomal genes in Rachycentron canadum (Rachycentridae, Perciformes).

Rachycentron canadum, the only representative of the family Rachycentridae, has been the focus of biotechnological interest due to its significant potential in marine fish farming. The chromosome set of this species has been widely investigated with respect to the location of genes and multigene families. A FISH analysis was performed using 4 multigene families as probes, represented by 5S and 18S ribosomal genes and histones H2B-H2A and H3. Earlier data suggested that differential replication of heterochromatin could be partially associated with functional genes. Indeed, our results showed that the DNA contained in heterochromatic regions of R. canadum contains 5S and 18S ribosomal genes as well as the gene sequences of histones H2B-H2A and H3, which were colocalized. The distribution of H3 sequences in all heterochromatic regions, except in 13q, could indicate an important evolutionary role for this class of repetitive sequences. Besides, the presence of chromosome regions bearing multifunctional repetitive sequences formed by H2B-H2A/H3/18S rDNA and H2B-H2A/H3/5S rDNA clusters was demonstrated for the first time in fishes. The implications of differential histone gene extension and its functionality in the karyotype of R. canadum remain unknown.

Chromosomal mapping of repetitive DNAs in Gobionellus oceanicus and G. stomatus (Gobiidae; Perciformes): A shared XX/XY system and an unusual distribution of 5S rDNA sites on the Y chromosome.

With nearly 2,000 species, Gobiidae is the most specious family of the vertebrates. This high level of speciation is accompanied by conspicuous karyotypic modifications, where the role of repetitive sequences remains largely unknown. This study analyzed the karyotype of 2 species of the genus Gobionellus and mapped 18S and 5S ribosomal RNA genes and (CA)15 microsatellite sequences onto their chromosomes. G. oceanicus (2n = 56; ♂ 12 metacentrics (m) + 4 submetacentrics (sm) + 1 subtelocentric (st) + 39 acrocentrics (a); ♀ 12m + 4sm + 2st + 38a) and G. stomatus (2n = 56; ♂ 20m + 14sm + 1st + 21a; ♀ 20m + 14sm + 2st + 20a) possess the highest diploid chromosome number among the Gobiidae and have different karyotypes. Both species share an XX/XY sex chromosome system with a large subtelocentric X and a small acrocentric Y chromosome which is rich in (CA)15 sequences and bears 5S rRNA sites. Although coding and noncoding repetitive DNA sequences may be involved in the genesis or differentiation of the sex chromosomes, the exclusive presence of 5S rDNA sites on the Y, but not on the X chromosome of both species, represents a novelty in fishes. In summary, the karyotypic differences, as well as new data on the sex chromosome systems in these 2 Gobiidae species, confirm the high chromosomal dynamism observed in this family.