The mapping of repetitive DNA sites by fluorescence in situ hybridization has been widely used for karyotype studies in different species of fish, especially when dealing with related species or even genera presenting high chromosome variability. This study analyzed three populations of Bryconamericus, with diploid number preserved, but with different karyotype formulae. Bryconamericus ecai, from the Forquetinha river/RS, presented three new cytotypes, increasing the number of karyotype forms to seven in this population. Other two populations of Bryconamericus sp. from the Vermelho stream/PR and Cambuta river/PR exhibited interpopulation variation. The chromosome mapping of rDNA sites revealed unique markings among the three populations, showing inter- and intrapopulation variability located in the terminal region. The molecular analysis using DNA barcoding complementing the cytogenetic analysis also showed differentiation among the three populations. The U2 small nuclear DNA repetitive sequence exhibited conserved features, being located in the interstitial region of a single chromosome pair. This is the first report on its occurrence in the genus Bryconamericus. Data obtained revealed a karyotype variability already assigned to the genus, along with polymorphism of ribosomal sites, demonstrating that this group of fish can be undergoing a divergent evolutionary process, constituting a substantive model for studies of chromosomal evolution.
Rhamdia quelen, a species of Heptapteridae, is considered to be a complex because of taxonomic and phylogenetic inconsistencies. Determining the physical location of repetitive DNA sequences on the chromosomes and the DNA barcode might increase our understanding of these inconsistencies within different groups of fish. To this end, we analyzed R. quelen populations from two river basins in Brazil, Paraguay and Parana, using DNA barcoding and different chromosomal markers, including U2 snDNA, which has never been analyzed for any Rhamdia species. Cytochrome c oxidase I gene sequence analysis revealed a significant differentiation among populations from the Miranda and Quexada rivers, with genetic distances compatible to those found among different species in neotropical fishes. Our results, in general, revealed a conservative chromosomal evolution in R. quelen and a differential distribution of some markers, such as 5S rDNA and U2 snDNA, in different populations. We suggest that R. quelen must undergo a major revision in its morphological, genetic, and cytogenetic molecular and taxonomic structure to elucidate possible operational taxonomic units.
The family Curimatidae is a fish group usually considered chromosomally conserved in their diploid number. However, some studies show small changes in the karyotype microstructure, and the presence of B chromosomes, indicating a chromosomal diversification within the group, even if structural changes in the karyotypes are not visible. Few studies associate this trait with an evolutionary pattern within the family. This study aimed to characterize the karyotype, (NORs)nucleolus organizer regions , and heterochromatin distribution of six species of Curimatidae of the genera Cyphocharax Fowler, 1906 and Steindachnerina Fowler, 1906: Cyphocharax voga (Hensel, 1870), Cyphocharax spilotus (Vari, 1987), Cyphocharax saladensis (Meinken, 1933), Cyphocharax modestus (Fernández-Yépez, 1948), Steindachnerina biornata (Braga et Azpelicueta, 1987) and Steindachnerina insculpta (Fernández-Yépez, 1948) and contribute data to a better understanding of the mechanisms involved in the chromosomal evolution of this group of fish. All specimens had 2n=54, m-sm, and B microchromosomes. Five species exhibited single NORs, except for Steindachnerina biornata, which showed a multiple pattern of ribosomal sites. NORs were chromomycin A3 positive (CMA3+) and 4’-6-diamino-2-phenylindole (DAPI-) negative, exhibiting differences in the pair and chromosomal location of each individual of the species. FISH with 5S rDNA probe revealed sites in the pericentrometic position of a pair of chromosomes of five species. However, another site was detected on a metacentric chromosome of Cyphocharax spilotus. Heterochromatin distributed both in the pericentromeric and some terminal regions was revealed to be CMA3+/DAPI-. These data associated with the previously existing ones confirm that, although Curimatidae have a very conservative karyotype macrostructure, NORs and heterochromatin variability are caused by mechanisms of chromosome alterations, such as translocations and/or inversions, leading to the evolution and diversification of this group of fish.
Crenicichla is the largest genus in the Cichlidae family in South America. The genus includes 100 valid species that are popularly known in Brazil as jacundás or joaninhas and are widely distributed in rivers east of the Andes. Cytogenetic analyses were carried out on seven species in this genus . All species showed a diploid number of 48 with interspecific differences in karyotype formulas and AgNORs located in interstitial position on the short arm of the largest metacentric pair, except for the two populations from C. britskii. Population A showed terminal markings on the long arm of the fifth pair of the complement, and population B showed up to two marked chromosome pairs. FISH with an 18S rDNA probe was coincident with AgNORs and CMA 3 , except for pair 6 from population B of C. britskii that did not presented positive CMA 3 sites. This work presents first cytogenetic data for C. haroldoi , C. maculata, and C. punctata , and the results show karyotypic patterns similar to those in the literature. However, the diversity found in populations of C. britskii represents new information about the evolution of the karyotype of the Cichlidae family, which has been conservative. Furthermore, the data could assist in phylogenetic studies of Crenicichla .
O processo de splicing do RNA mensageiro é realizado pelo spliceossomo em um dos maiores e mais complexos mecanimos moleculares da célula. O spliceossomo consiste em cinco RNAs nucleares (snRNAs) – U1, U2 U4,U5 e U6 que, unidos com vários fatores proteicos, formam uma ribonucleoproteína nuclear (snRNPs). Os ciclídeos são um dos maiores e mais diversificados grupos de peixes, sendo importantes modelos de evolução biológica, além de serem apreciados na aquariofilia e aquicultura. Apesar disso, ainda são poucos os trabalhos citogenéticos, com somente 154 espécies cariotipadas até o momento e este número se restringe ainda mais quando se trata de análises com sequências de DNAs repetitivos, principalmente em ciclídeos neotropicais. Sendo assim, o presente estudo tem como objetivo a localização cromossômica do gene snRNA U2 em diferentes espécies da família Cichlidae. Foi realizada a extração de DNA de Cichlasoma e utilizada como molde para amplificação do gene U2, com os primers U2F (59-ATC GCT TCT CGG CCT TAT G–39) e U2R (59-TCC CGG CGG TAC TGC AAT A-39). Essa PCR foi marcada com Cy3dUTP. O fragmento da PCR obtido no presente trabalho foi de, aproximadamente, 200 pb o que parece ser semelhante ao encontrado para o mesmo gene no genoma humano, que mostra uma região codificante de 188 pb. A hibridação fluorescente in situ (FISH) foi realizada em seis espécies de ciclídeos neotropicais. Cichlasoma portalegrense, Gymnogeophagus gymnogenys, Crenicichla lepdota, Crenicichla maculata e Crenicichla niederleinii apresentaram um par portador do sitio de DNAsn U2, enquanto Crenicichla britski apresentou dois pares. Os estudos com a sequência de DNAsn U2 em peixes ainda são muito restritos, mas em sua maioria evidenciam essa sequência em um único par cromossômico contudo, estudos recentes demonstram uma variabilidade maior em relação ao mapeamento físico desta sequência. Em ciclideos existem análises de DNAsn U1 para 12 espécies, todas com um único sítio portador da sequência entretanto, para DNAsn U2 os dados aqui apresentados são os primeiro neste grupo de peixes e, apesar de preliminares, já indicam que análises desta e outras sequências de DNA repetitivo podem contribuir para um maior entendimento da organização genômica da família Cichlidae.
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