Cytogenetic aspects of a canine breast carcinosarcoma - a case report.

This study searched a rare and aggressive type of cancer in dogs and humans, the breast carcinosarcoma. Both clinical and pathological traits of mammary carcinosarcomas in dogs are similar to humans, such as infrequent occurrence, fast tumor growth, and unfavorable prognosis when compared to carcinomas. Other possible alterations include chromosomal abnormalities that can be useful for the identification of tumoral cells and diagnosis. The aim of this study was to compare the chromosomal features of peripheral lymphocytes and tumor cells in a mammary carcinosarcoma of a 14-year-old female Poodle. Chromosomes were analyzed from 210 metaphases by conventional Giemsa staining, C-banding, and base-specific fluorochrome staining with chromomycin A3 (CMA3+) and DAPI. Of the 105 blood cells, 56.3% followed the standard karyotype of dogs (2n = 78). In contrast, the carcinosarcoma cells showed high chromosomal numbers (104 to 153), divided into 80% hypertriploid (118 to 136 chromosomes), 10.5% hypotetraploid (137 to 153 chromosomes), 5.7% hypotriploid (104 to 116 chromosomes), and 3.8% triploid cells (117 chromosomes). Among the aneuploid cells identified, we highlighted the trisomy of pair 1 and X chromosome once these elements were easily recognized in karyotype because of their size (pair 1) or differential morphology. Heterochromatin in normal cells was restricted to the pericentromeric region of all chromosomes while few C-bands were observed in tumor cells. This apparent loss of heterochromatin in neoplastic cells was supposed to favor centric fusion among formerly acrocentric chromosomes. Fluorochrome staining reinforced this hypothesis once GC-rich segments (CMA3+) were identified on 10 chromosomes from normal cells (2n = 78) whereas carcinosarcoma metaphases had up to 11 chromosomes bearing CMA3 signals in spite of their remarkable high chromosomal numbers. We concluded that, like in humans, the carcinosarcoma in dogs caused genome instability that eventually led to structural and numerical chromosomal aberrations. Besides, this study reinforced the importance of cytogenetic studies in dogs as a reference material for human cancer studies, especially in rare cases, since it is possible to increase knowledge about the characteristics of breast neoplasms in which there is a little availability of similar cases for comparative studies.

Direct preparation protocol to obtain mitotic chromosomes from canine mammary tumors.

Currently, mammary neoplasms in female canines are a serious problem in veterinary clinics. In addition, the canine species is an excellent disease model for human oncology because of the biological and genetic similarities between the species. Cytogenetics has allowed further study of the characterization of neoplasms in canines. We hypothesized that the use of a direct preparation protocol for mitotic chromosome analysis would provide a simple and low cost protocol for use in all laboratories. The objective of this method is to display in a few hours of dividing cells just like the time of collection since cell division in tissue can be obtained. Ten female canines with the spontaneous occurrence of mammary neoplasia were used to test a pioneering direct preparation protocol to obtain mitotic chromosomes. The excised breast tumor tissue fragments were subjected to the protocol consisting of treatment with colchicine, treatment with hypotonic solution, and fixation. Mitotic chromosomes were absent in cell suspensions of only two samples among the 10 materials analyzed, based on the analysis of five blades for each preparation obtained. So, the cell suspension obtained allowed for the observation of eight tissue samples viable for cytogenetic analysis, five of which had excellent numbers of mitotic chromosomes. However, the technique was unsuccessful in producing high-quality cell suspensions because of inadequate condensation and scattering of chromosomes. While adjustments to methodological procedures are needed, this protocol represents a low cost and simplified method to study the cytogenetics of canine tumors.

Identification of naturalized goat breeds under conservation from northeastern Brazil using chromosomal markers.

In order to provide the first cytogenetic data of naturalized and threatened goat breeds from northeastern Brazil, cytogenetic analyses were carried out in individuals of Repartida and Moxotó breeds raised in Bahia and Ceará States. Males and females of both breeds had 2n = 60, with 29 autosomal acrocentric pairs plus the sex chromosome pair. The number of nucleolar organizer region (NOR)-bearing chromosomes ranged from 6 to 8 per metaphase in Moxotó and Repartida goats, respectively. The active NORs in Repartida individuals were located exclusively at the terminal regions of the long arms, as usually detected in Bovidae. Otherwise, Moxotó specimens presented a large autosomal pair with NORs on short arms. GC-rich heterochromatin was detected at the centromeres in both breeds, although polymorphic terminal C-bands were visualized on pair 25 in Moxotó. In addition, GC-rich regions were detected at the terminal regions of the long arms of a single pair in Repartida and of 20 chromosomes in Moxotó goats. The differences in both the number and/or position of Ag-NORs and GC-rich sites between Repartida and Moxotó breeds represent efficient cytogenetic markers that can be used in the identification and conservation of the genetic integrity of each lineage. In spite of the small effective population size of these breeds, chromosomal abnormalities related to drift or inbreeding effects were absent in the samples analyzed.

Remarkable karyotypic homogeneity in a widespread tropical fish species: Hoplosternum littorale (Siluriformes, Callichthyidae).

The first chromosomal data in Hoplosternum littorale from an isolated South American drainage in north-eastern Brazil are presented. All specimens were characterized by a diploid number (2n) of 60 chromosomes divided into three metacentric, one submetacentric and 26 acrocentric pairs; single nucleolar organizer regions (NOR) on the sixth pair; centromeric and interstitial heterochromatin; GC-rich sites on four large acrocentric chromosomes, including the NOR-bearing pair, and 5S ribosomal genes at terminal region on short arms of two acrocentric pairs. These data are invariably similar to previous reports in H. littorale from distant localities throughout South America, which contrasts with the chromosomal diversity of Callichthyidae and reinforces the role of human activities on the dispersal and colonization of this fish.

Population divergence and peculiar karyoevolutionary trends in the loricariid fish Hypostomus aff. unae from northeastern Brazil.

Loricariidae (Siluriformes, Hypostominae) is one of the most diverse catfish families. In spite of the wide distribution of loricariids in South America, cytogenetic reports are available for only a few species, mostly from southern and southeastern Brazil. We made the first chromosomal analysis of Hypostomus aff. unae from the Contas River basin in northeastern Brazil. Four populations isolated by short distances but from distinct landscapes were studied based on conventional staining, C-banding, argyrophilic nucleolar organizer regions (Ag-NOR), CMA(3)/DAPI fluorochrome staining, and fluorescent in situ hybridization with 18S rDNA probes. Although sharing the same diploid number (2n = 76) and NOR locations, each population presented exclusive karyotype formulae and specific patterns of heterochromatic and AT-rich regions. The derived karyotypes of H. aff. unae (2n >54; high number of acrocentrics bearing AT-rich interstitial heterochromatin) indicated a divergent karyoevolution, mostly driven by centric fissions, pericentric inversions and particular heterochromatin dispersion models. This finding of distinct evolutionary units in H. aff. unae will be useful for understanding the natural history of loricariids from relatively unexplored coastal basins in South America.

Heterochromatin heterogeneity in Hypostomus prope unae (Steindachner, 1878) (Siluriformes, Loricariidae)from Northeastern Brazil.

Cytogenetic analyses using C-banding and chromosomal digestion by several restriction enzymes were carried out in four populations (named A, B, C and D) of Hypostomus prope unae (Loricariidae, Hypostominae) from Contas river basin, northeastern Brazil. These populations share 2n=76 and single NORs on the second metacentric pair but exclusive karyotype forms for each locality. Populations A and B presented conspicuous terminal and interstitial heterochromatic blocks on most of acrocentric chromosomes and equivalent to NORs with differences in both position and bearing pair. Population D showed evident marks at interstitial regions and interspersed with nucleolar region while population C presented interstitial and terminal heterochromatin segments, non-coincident with NORs. The banding pattern after digestion with the endonucleases Alu I, Bam HI, Hae III and Dde I revealed a remarkable heterogeneity within heterochromatin, allowing the identification of distinctive clusters of repeated DNA in the studied populations, besides specific patterns along euchromatic regions. The analysis using restriction enzymes has proved to be highly informative, characterizing population differences and peculiarities in the genome organization of Hypostomus prope unae.

Chromosomal diversity and phylogenetic inferences concerning thrips (Insecta, Thysanoptera) in a semi-arid region of Brazil.

The order Thysanoptera is composed of cosmopolitan phytophagous and predaceous insects with diverse life histories, behaviors and habits. This order is currently thought to form a trichotomy with Hemiptera and Psocodea; Hemiptera and Thysanoptera are considered to be sister groups. The interrelationships within Thysanoptera remain unclear and cytotaxonomic studies are scarce in thrips. We report, for the first time, chromosomal data on seven species of thrips collected from a semi-arid region in the States of Bahia and Pernambuco (Northeast Brazil). A distinctive chromosomal pattern was observed in Thysanoptera when compared to other members within the infraclass Paraneoptera. Considerable karyotypic differences were also found within genera and species of Thysanoptera. Based on these data, we suggest that Paraneoptera forms a polyphyletic group and that Terebrantia and Tubulifera should be regarded as sister groups. The high chromosomal variability observed in Thysanoptera indicates that chromosomal rearrangements have played a key role in their speciation pathways.

Comparação de metodologias de seleção sob oscilação genética / Comparison of selection methodologies under genetic drift

Determinou-se o número adequado de repetições na comparação de métodos de seleção tradicionais e associados a marcadores moleculares, com diferentes tamanhos efetivos e sob diferentes sistemas de acasalamento dos reprodutores selecionados, usando simulação com o programa GENESYS. Para comparar os diferentes métodos de seleção utilizaram-se populações com tamanhos efetivos de 18,18 (TE1) e de 66,66 (TE2) e uma, 10 e 30 repetições por geração, avaliando-se os valores fenotípicos médios. Para as situações com apenas uma repetição, os resultados apresentaram incoerências, independentemente do tamanho efetivo (TE1 ou TE2) ou do sistema de acasalamento (RAA - reprodutores acasalados aleatoriamente, EIC - exclusão de irmãos completos ou EICMI - exclusão de irmãos completos e meio-irmãos). Observou-se que a oscilação genética influencia o ganho genético, principalmente, em populações com pequeno tamanho efetivo e que um valor mínimo de 10 repetições por geração é necessário para assegurar a consistência dos resultados obtidos pelos métodos de seleção.

This work was carried out to determine the required number of replicates for comparison of conventional and molecular marker-associated selection methods in distinct effective population sizes and different mating systems, by simulations using the software GENESYS. Effective populational size of 18.18 (ES1) and 66.66 (ES2), and one, 10, and 30 replicates per generation were used to compare the different selection methods, based on mean phenotypic values. Incongruences results were observed when a single replicate was considered independently of effective size (ES1or ES2) and the mating system (random mating; exclusion of full-sibs or exclusion of both full and half-sibs). Genetic oscillation influenced the genetic gain, mainly in populations of small effective size. Furthermore, at least 10 replicates per generation were required to obtain sound consistent results for both selection methods.

Genetic diversity of three ornamental reef fishes (Families Pomacanthidae and Chaetodontidae) from the Brazilian coast

Reef fishes of the families Pomacanthidae (angelfish) and Chaetodontidae (butterflyfish) are popular ornamental species, intensively harvested for the aquarium trade. The impacts of such activity on intra-specific diversity and reef ecosystems are still poorly understood in the south Atlantic. In the present work, a fine-scale genetic analysis using RAPD markers was performed in distinct samples of the queen angelfish (Holacanthus ciliaris), French angelfish (Pomacanthus paru), and banded butterflyfish (Chaetodon striatus) along the Brazilian coast. Most of the genetic variation in the three species was related to intra-population diversity. However, AMOVA results demonstrated that H. ciliaris presents a subtle population structure (sigmast = 0.132, P = 0.003), while P. paru and C. striatus present low genetic differentiation, especially remarkable in the latter (sigmast = 0.090, P = 0.001 and sigmast = 0.041, P = 0.028, respectively). Gene flow (Nm) was also higher in C. striatus than in the angelfish species. The reported patterns of genetic differentiation contrast with the similar pelagic stage of the selected species, suggesting that larval dispersal per se is a poor predictor of population structure in these reef fishes. Ecological features coupled with biogeographic history and distinct local selective pressures might play a major role on the genetic composition of each species. Although preliminary, the present results provide a baseline for monitoring the genetic variability in these reef species. These differences in the genetic structure among co-occurring species should be taken into consideration for the conservation of eventual evolutionary units along the Brazilian Province.

Os peixes recifais das famílias Pomacanthidae (peixes-anjo) e Chaetodontidae (peixes-borboleta) são espécies ornamentais populares, intensivamente coletadas para o comércio aquariófilo. Os impactos dessa atividade na diversidade intra-específica e no ecossistema recifal ainda são pouco conhecidos no Atlântico Sul. No presente trabalho, uma análise genética em fina escala usando marcadores RAPD foi realizada em diferentes amostras de Holacanthus ciliaris (anjo ciliaris), Pomacanthus paru (peixe frade) e Chaetodon striatus (borboleta striatus) ao longo da costa brasileira. A maior parte da variação genética nas três espécies relacionou-se à diversidade intrapopulacional. Contudo, resultados da AMOVA demonstraram que H. ciliaris apresenta uma estruturação populacional sutil (sigmast = 0,132, P = 0,003), enquanto P. paru e C. striatus apresentam uma baixa diferenciação genética, particularmente reduzida na última (sigmast = 0,090, P = 0,001 e sigmast = 0,041, P = 0,028, respectivamente). O fluxo gênico (Nm) também foi mais alto em C. striatus do que nas espécies de Pomacanthidae. Os padrões de diferenciação genética registrados contrastam com o estágio pelágico inicial similar nas espécies selecionadas, sugerindo que a dispersão larval per se não é um bom indicador de estrutura populacional nesses peixes recifais. Fatores ecológicos associados à história biogeográfica e distintas pressões seletivas locais podem desempenhar um papel fundamental na composição genética de cada espécie. Embora preliminares, os resultados apresentados fornecem subsídios para o monitoramento da variabilidade genética dessas espécies recifais. Tais diferenças na estrutura genética entre espécies co-existentes devem ser consideradas para a conservação de eventuais unidades evolutivas dentro da Província Brasileira.

Genetic diversity of three ornamental reef fishes (Families Pomacanthidae and Chaetodontidae) from the Brazilian coast.

Reef fishes of the families Pomacanthidae (angelfish) and Chaetodontidae (butterflyfish) are popular ornamental species, intensively harvested for the aquarium trade. The impacts of such activity on intra-specific diversity and reef ecosystems are still poorly understood in the south Atlantic. In the present work, a fine-scale genetic analysis using RAPD markers was performed in distinct samples of the queen angelfish (Holacanthus ciliaris), French angelfish (Pomacanthus paru), and banded butterflyfish (Chaetodon striatus) along the Brazilian coast. Most of the genetic variation in the three species was related to intra-population diversity. However, AMOVA results demonstrated that H. ciliaris presents a subtle population structure (phi(st)=0.132, P=0.003), while P. paru and C. striatus present low genetic differentiation, especially remarkable in the latter (phi(st) = 0.090, P=0.001 and phi(st)=0.041, P=0.028, respectively). Gene flow (Nm) was also higher in C. striatus than in the angelfish species. The reported patterns of genetic differentiation contrast with the similar pelagic stage of the selected species, suggesting that larval dispersal per se is a poor predictor of population structure in these reef fishes. Ecological features coupled with biogeographic history and distinct local selective pressures might play a major role on the genetic composition of each species. Although preliminary, the present results provide a baseline for monitoring the genetic variability in these reef species. These differences in the genetic structure among co-occurring species should be taken into consideration for the conservation of eventual evolutionary units along the Brazilian Province.