In the present study, specimens of Bryconamericus ecai collected from the Forquetinha River/RS, were cytogenetically analyzed, disclosing a wide karyotypic diversity in this species. All individuals had 2n = 50, with different karyotypic formulae, resulting in four cytotypes and one B macrochromosome observed in cytotype III. Heterochromatin was distributed in the pericentromeric region of most chromosomes on the four cytotypes and also on a chromosome pair with interstitial markings in cytotype IV. Staining with CMA(3) and DAPI fluorochromes revealed a C-band region rich in AT base pairs in cytotypes I, II and III, and a pair with GC-rich heterochromatin in cytotypes II and III. Cytotype IV presented CMA(3) and DAPI positive heterochromatin. Silver nitrate impregnation, in situ hybridization, and fluorochrome staining showed a multiple system of AgNORs, 18S rDNA and CMA(3) sites in cytotypes I, III and IV, with both inter-and intraindividual variability in the number and location of these sites. Cytotype II had only one pair of NORs coincident with the 18S rDNA and CMA(3) sites, indicating a simple system. The chromosomal polymorphism observed among the specimens of B. ecai added to the literature data show that chromosomal rearrangements, especially pericentric inversions, play an important role in the karyotypic evolution of this group of fish. It can also be implied that more than one species of Bryconamericus is probably occurring, living in sympatry in the Forquetinha River/RS.
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.
SummaryIn the present study, we analyzed 20 specimens of Bryconamericus aff. iheringii (12 males and 8 females), collected from the Três Bocas Stream, a tributary of the Tibagi River/PR. All individuals analyzed presented 2n=52 and different karyotypic formulae resulting in 4 cytotypes: cytotype I, with 12m+16sm+10st+14a and FN=90; cytotype II, with 14m+18sm+10st+10a and FN=94; cytotype III, with 10m+24sm+6st+12a and FN=92; and cytotype IV with 10m+14sm+8st+20a and NF=84. Such structural karyotypic variations among Bryconamericus aff. iheringii specimens are probably due to the occurrence of chromosomal rearrangements, such as pericentric inversions. In some individuals, nucleolus organizer regions (AgNORs) were observed on the short arm of a single submetacentric chromosome pair. In other individuals of this same population, an interindividual variation from 2-5 NOR-bearing chromosomes with markings located on chromosomes of different types and size was detected, along with AgNOR size heteromorphism between the homologous chromosomes in a metacentric pair. Thence, this population of Bryconamericus aff. iheringii from the Três Bocas Stream seems to show the occurrence of both multiple and single NOR systems, and also the presence of different karyotypes (cytotypes), which might suggest the occurrence of 2 different species of Bryconamericus living in sympatry in that location. The data discussed here and those presented in the literature show the need for a taxonomic revision in this group of fish.
Astyanax has been the subject of extensive cytogenetic studies due to its wide karyotypic diversity. This genus comprises species complexes, namely groups of fish of difficult morphological differentiation, such as the bimaculatus complex, which includes the characids with a rounded humeral spot. Thence, the present study proposed to accomplish a cytogenetic characterization of two species of this complex: A. asuncionensis and A. altiparanae, aiming to find chromosomal markers that differentiate these species, as well as achieve a better understanding of the karyotype evolution in the genus. For this we used different techniques of chromosome banding as C-banding, impregnation by silver nitrate, fluorochrome staining and FISH with 18S rDNA probe. This is the first cytogenetic study in A. asuncionensis, from Miranda river, which presented 2n = 50 and 18 m + 22sm + 6st + 4a (FN = 96) and single NORs. The populations of A. altiparanae also presented 2n = 50, but with different karyotypic formulae: the population of the Quexada river presented 16 m + 24sm + 4st + 6a (FN = 94) and the Esperança stream and Jacutinga river showed 16 m + 20sm + 4st + 10a (FN = 90). All analyzed populations showed an interindividual variation in the number and location of the nucleolar organizer regions (NORs). Single and multiple NORs were detected either by impregnation with silver nitrate or by FISH with 18S rDNA probe. After C-banding, the two species differed in relation to the composition and heterochromatin distribution. The meiotic cells of A. altiparanae male individuals were also analyzed, showing that, despite the high karyotype variability, chromosome pairing occurs normally. The data show that A. altiparanae and A. asuncionensis share some characteristics with other species of the bimaculatus complex, suggesting a close phylogenetic relationship among those species. However, some features can be used as differentiation chromosomal markers in altiparanae/asuncionensis morphotypes, which could result from a natural speciation process.
Análises de sequências de DNAs repetitivos proporcionam uma caracterização cromossômica mais apurada, auxiliando na compreensão de problemas taxonômicos e filogenéticos. Rhamdia quelen é considerada um complexo de espécies e, apesar de ser a espécie mais estudada dentro de Heptapteridae, pouco se sabe sobre sequências de DNA repetitivo no grupo. Sendo assim, o objetivo deste trabalho foi realizar uma análise citogenética comparativa entre seis populações de R. quelen. Foi confirmado o número diplóide igual 58, com variação nas fórmulas cariotípicas sendo: 40m+10sm+4st+4a e número fundamental (NF) igual 112, para a população do rio Quexada/PR; 40m+12sm+6st (NF=116) para exemplares do Ribeirão do Penacho/PR; 34m+16sm+8st (NF=116) para rio Miranda/MS e 32m+8sm+18st (NF=116) para a população do rio Cambé. A região organizadora de nucléolos (AgRONs) foi evidenciada na região terminal de um cromossomo submetacêntrico, nas quatro populações de R. quelen, sendo coincidente com a coloração pelo fluorocromo CMA3. A população do rio Quexada apresentou um heteromorfismo de tamanho das AgRONs entre os cromossomos homólogos. O mapeamento cromossômico de genes ribossomais 18S e 5S, e de U2 DNAsn foi realizado nas quatro populações citadas de R. quelen, mais as do rio Taquari e do ribeirão Lindóia, ambas do estado do Paraná. A distribuição dos genes de DNAr 18S confirmou o padrão simples da RON, em todas as populações, estando co-localizados com alguns sítios de U2 DNAsn. Os resultados mostraram uma variação das sequências de DNAs repetitivos na população do rio Miranda em comparação com as demais populações. Apenas os indivíduos desta localidade apresentaram um único par portador da sequência de U2 DNAsn, assim como foi a única população a mostrar um padrão múltiplo de distribuição do DNAr 5S. Esta heterogeneidade da população do rio Miranda em relação às demais populações de R. quelen, foi confirmada pela análise da sequência que codifica a subunidade I da enzima citocromo oxidase C, mostrando uma diferenciação em relação à população do rio Quexada. Os dados revelam uma variação interpopulacional na microestrutura cromossômica da espécie nunca antes evidenciada, caracterizando essas famílias de DNA repetitivos como importantes ferramentas em estudos filogenéticos e taxonômicos.
A male miniature Schnauzer with cryptorchidism and a mass growth in the right inguinal canal was admitted to the clinical center of a veterinary hospital. During surgical resection of the mass, tubular formations were found, connecting the mass to the left testicle. Histopathology revealed that the tubular formations were uterine tubes and the mass was a seminoma associated with a sertolioma of the right testicle. Further analysis also showed atrophy of the left testicle. Cytogenetic evaluation revealed normal chromosomes of male gender, 78, XY, which led to the conclusion that this was a male pseudohermaphrodite. Key words: Canine. Cryptorchid. Cytogenetic. Persistent Müllerian duct syndrome. Tumor. ResumoFoi atendido na rotina clínica de um hospital veterinário, um cão, macho, da raça Schnauzer, criptorquida com massa em canal inguinal direito. Durante o procedimento cirúrgico para a retirada da massa, foi encontrada uma formação tubular a qual fazia a ligação entre essa massa e o testículo esquerdo. A análise histopatológica concluiu que a formação tubular tratava-se de tubas uterinas, a massa era o testículo direito com seminoma e sertolioma e o testículo esquerdo apresentava atrofia. A avaliação citogenética revelou cromossomos normais, do sexo masculino, 78, XY concluindo que se tratava de um pseudo-hermafrodita masculino. Palavras-chave: Canino. Citogenética. Criptorquidismo. Síndrome da persistência dos ductos de Müller. Tumor.
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