Mapping of Repetitive Sequences in Brachyhypopomus brevirostris (Hypopomidae, Gymnotiformes) from the Brazilian Amazon.

Brachyhypopomus (Hypopomidae, Gymnotiformes) is a monophyletic genus consisting of 28 formally described species. Karyotypic data are available for 12 species. The same karyotype is described for two species (B. brevirostris and B. hamiltoni), as well as different karyotypes for the same species from distinct locations (B. brevirostris). In this context, B. brevirostris may constitute a cryptic species complex. Thus, in the present study, we analyzed the karyotype of B. brevirostris, from Santarém, Pará, and Tefé, Amazonas, using classical cytogenetics (conventional staining and C-banding) and molecular techniques (fluorescence in situ hybridization using 18S rDNA, 5S rDNA, U2 snRNA, and telomeric probes). The results show that samples from both locations present 2n = 38, with all chromosomes being acrocentric (FC = 38a). In both populations, 18S rDNA sequences are present on only one pair of homologous chromosomes and telomeric sequences occur only at the ends of the chromosomes. In the Tefé sample, the 5S rDNA occurs in two pairs, and the U2 snRNA in three pairs. These results are the first descriptions of these sequences for B. brevirostris samples from the Tefé locality, as well as the first karyotypic description for the Santarém locality. Future cytotaxonomic studies of this genus can benefit from these results.

Chromosome Painting in Gymnotus carapo "Catalão" (Gymnotiformes, Teleostei): Dynamics of Chromosomal Rearrangements in Cryptic Species.

The genus Gymnotus is a large monophyletic group of freshwater weakly-electric fishes, with wide distribution in Central and South America. It has 46 valid species divided into six subgenera (Gymnotus, Tijax, Tigre, Lamontianus, Tigrinus and Pantherus) with large chromosome plasticity and diploid numbers (2n) ranging from 34 to 54. Within this rich diversity, there is controversy about whether Gymnotus (Gymnotus) carapo species is a single widespread species or a complex of cryptic species. Cytogenetic studies show different diploid numbers for G. carapo species, ranging from 40 to 54 chromosomes with varied karyotypes found even between populations sharing the same 2n. Whole chromosome painting has been used in studies on fish species and recently has been used for tracking the chromosomal evolution of Gymnotus and assisting in its cytotaxonomy. Comparative genomic mapping using chromosome painting has shown more complex rearrangements in Gymnotus carapo than shown in previous studies by classical cytogenetics. These studies demonstrate that multiple chromosome pairs are involved in its chromosomal reorganization, suggesting the presence of a complex of cryptic species due to a post zygotic barrier. In the present study, metaphase chromosomes of G. carapo occidentalis "catalão" (GCC, 2n = 40, 30m/sm+10st/a) from the Catalão Lake, Amazonas, Brazil, were hybridized with whole chromosome probes derived from the chromosomes of G. carapo (GCA, 2n = 42, 30m/sm+12st/a). The results reveal chromosome rearrangements and a high number of repetitive DNA sites. Of the 12 pairs of G. carapo chromosomes that could be individually identified (GCA 1-3, 6, 7, 9, 14, 16 and 18-21), 8 pairs (GCA 1, 2, 6, 7, 9, 14, 20, 21) had homeology conserved in GCC. Of the GCA pairs that are grouped (GCA [4, 8], [5, 17], [10, 11] and [12, 13, 15]), most kept the number of signals in GCC (GCA [5, 17], [10, 11] and [12, 13, 15]). The remaining chromosomes are rearranged in the GCC karyotype. Analysis of both populations of the G. carapo cytotypes shows extensive karyotype reorganization. Along with previous studies, this suggests that the different cytotypes analyzed here may represent different species and supports the hypothesis that G. carapo is not a single widespread species, but a group of cryptic species.

Archolaemus janeae (Gymnotiformes, Teleostei): First insights into karyotype and repetitive DNA distribution in two populations of the Amazon.

Archolaemus, one of the five genera of Neotropical freshwater fish of the family Sternopygidae (Gymnotiformes), was long considered a monotypic genus represented by Archolaemus blax. Currently, it consists of six species, most of them occurring in the Amazon region. There are no cytogenetic data for species of this genus. In the present study, we used classical cytogenetics (conventional staining and C-banding) and molecular cytogenetics (probes of telomeric sequences and multigenic families 18S rDNA, 5S rDNA, and U2 snDNA) to study the karyotype of Archolaemus janeae from Xingu and Tapajós rivers in the state of Pará (Brazil). The results showed that the two populations have identical karyotypes with 46 chromosomes: four submetacentric and 42 acrocentric (2n = 46; 4m/sm + 42a). Constitutive heterochromatin occurs in the centromeric region of all chromosomes, in addition to small bands in the interstitial and distal regions of some pairs. The 18S rDNA occurs in the distal region of the short arm of pair 2; the 5S rDNA occurs in five chromosome pairs; and the U2 snDNA sequence occurs in chromosome pairs 3, 6, and 13. No interstitial telomeric sequence was observed. These results show karyotypic similarity between the studied populations suggesting the existence of a single species and are of great importance as a reference for future cytotaxonomic studies of the genus.

Particular Chromosomal Distribution of Microsatellites in Five Species of the Genus Gymnotus (Teleostei, Gymnotiformes).

Microsatellites show great abundance in eukaryotic genomes, although distinct chromosomal distribution patterns might be observed, from small dispersed signals to strong clustered motifs. In Neotropical fishes, the chromosome mapping of distinct microsatellites was employed several times to uncover the origin and evolution of sex and supernumerary chromosomes, whereas a detailed comparative analysis considering different motifs at the chromosomal level is scarce. Here, we report the chromosomal location of several simple sequence repeats (SSRs) in distinct electric knife fishes showing variable diploid chromosome numbers to unveil the structural organization of several microsatellite motifs in distinct Gymnotus species. Our results showed that some SSRs are scattered throughout the genomes, whereas others are particularly clustered displaying intense genomic compartmentalization. Interestingly, the motifs CA, GA, and GAG exhibited a band-like pattern of hybridization, useful for the identification of homologous chromosomes. Finally, the colocalization of SSRs with multigene families is probably related to the association of microsatellites with gene spacers in this case.

Extensive Karyotype Reorganization in the Fish Gymnotus arapaima (Gymnotiformes, Gymnotidae) Highlighted by Zoo-FISH Analysis.

The genus Gymnotus (Gymnotiformes) contains over 40 species of freshwater electric fishes exhibiting a wide distribution throughout Central and South America, and being particularly prevalent in the Amazon basin. Cytogenetics has been an important tool in the cytotaxonomy and elucidation of evolutionary processes in this genus, including the unraveling the variety of diploid chromosome number (2n = from 34 to 54), the high karyotype diversity among species with a shared diploid number, different sex chromosome systems, and variation in the distribution of several Repetitive DNAs and colocation and association between those sequences. Recently whole chromosome painting (WCP) has been used for tracking the chromosomal evolution of the genus, showing highly reorganized karyotypes and the conserved synteny of the NOR bearing par within the clade G. carapo. In this study, painting probes derived from the chromosomes of G. carapo (GCA, 2n = 42, 30 m/sm + 12 st/a) were hybridized to the mitotic metaphases of G. arapaima (GAR, 2n = 44, 24 m/sm + 20 st/a). Our results uncovered chromosomal rearrangements and a high number of repetitive DNA regions. From the 12 chromosome pairs of G. carapo that can be individually differentiated (GCA1-3, 6, 7, 9, 14, 16, and 18-21), six pairs (GCA 1, 9, 14, 18, 20, 21) show conserved homology with GAR, five pairs (GCA 1, 9, 14, 20, 21) are also shared with cryptic species G. carapo 2n = 40 (34 m/sm + 6 st/a) and only the NOR bearing pair (GCA 20) is shared with G. capanema (GCP 2n = 34, 20 m/sm + 14 st/a). The remaining chromosomes are reorganized in the karyotype of GAR. Despite the close phylogenetic relationships of these species, our chromosome painting studies demonstrate an extensive reorganization of their karyotypes.

Gymnotus coatesi (Gymnotiformes): A Case of Colocation of Multiple Sites of 18S rDNA with Telomeric Sequences.

Gymnotus coatesi is a small and rare species of banded knife fish that was originally described by LaMonte in 1935, found along the main stretch of the Amazon River. There is no described cytogenetic data on this species. We analyzed the karyotype of five specimens of G. coatesi collected from Cururutuia Stream in Bragança, Pará, Brazil. The obtained diploid number is 50 and the karyotypic formula is 24 m/sm +26 st/a. The constitutive heterochromatin is DAPI positive and distributed mainly in the centromeric and pericentromeric regions of the chromosomes. Ag-nucleolus organizer regions staining showed nine active sites. The 5S rDNA probe hybridized chromosome pair 17 in the interstitial part of the long arm. Fluorescence in situ hybridization (FISH) with telomeric probes revealed signals only at terminal regions of the chromosomes. The 18S rDNA probe hybridized to 21 sites, and these signals colocalized with the telomeric sequences. This relatively high number of 18S rDNA sites may reflect gene duplication mediated by transposable elements. These results indicate that although the diploid number of G. coatesi is within the range previously observed for other members of the genus, various karyotypic characteristics distinguish G. coatesi from the other species of the genus and members of the Gymnotiform order.