Cytogenetic analyses of eight species in the genus Leptodactylus Fitzinger, 1843 (Amphibia, Anura, Leptodactylidae), including a new diploid number and a karyotype with multiple translocations

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Cytogenetics of a new species of Paratelmatobius cardosoi group (Anura : Leptodactylidae), with the description of an apparent case of pericentric inversion

The karyotype of a new species of Paratelmatobius from the P cardosoi group is described. As with other Paratelmatobius and Scythrophrys karyotypes, Paratelmatobius sp. (aff. cardosoi) shows a diploid number of 24 chromosomes, in addition to other similarities with the former karyotypes. The Paratelmatobius sp. (aff. cardosoi) karyotype differs from that of P. cardosoi in the morphology of pair 4, the NOR location and the C-bands in pairs 3 and 8 (exclusive to Paratelmatobius sp.) and those of pairs 7 and 9 (exclusive to P. cardosoi). Both karyotypes also differ in the amount of heterochromatin in pair 1. The presence of interstitial heterochromatin in the long arm of pair 1 and the interstitial C-bands in both arms of chromosome 5 are apparently synapomorphic characters of P. cardosoi and Paratelmatobius sp. (aff. cardosoi), since they are absent in the other Paratelmatobius and Scythrophrys karyotypes. In Paratelmatobius sp. (aff. cardosoi), the nucleolus organizer region is on the short arm of a small metacentric chromosome (pair 9), an arrangement similar to the NOR-bearing chromosome pair in the karyotype of P. poecilogaster and in karyotype 11 of Scythrophrys. A conspicuous heteromorphism unrelated to the sex determining mechanism was also observed and probably arose from a pericentric inversion.

Chromosome banding in three species of Hypsiboas (Hylidae, Hylinae), with special reference to a new case of B-chromosome in anuran frogs and to the reduction of the diploid number of 2n=24 to 2n=22 in the genus

The chromosomes of hylids Hypsiboas albopunctatus, H. raniceps, and H. crepitans from Brazil were analyzed with standard and differential staining techniques. The former species presented 2n = 22 and 2n = 23 karyotypes, the odd diploid number is due to the presence of an extra element interpreted as B chromosome. Although morphologically very similar to the small-sized chromosomes of the A complement, the B was promptly recognized, even under standard staining, on the basis of some characteristics that are usually attributed to this particular class of chromosomes. The two other species have 2n = 24, which is the chromosome number usually found in the species of Hypsiboas karyotyped so far. This means that 2n = 22 is a deviant diploid number, resulted from a structural rearrangement, altering the chromosome number of 2n = 24 to 2n = 22. Based on new chromosome data, some possibilities were evaluated for the origin of B chromosome in Hypsiboas albopunctatus, as well as the karyotypic evolution in the genus, leading to the reduction in the diploid number of 2n = 24 to 2n = 22.

Comparative cytogenetic analysis on four tree frog species (Anura, Hylidae, Hylinae) from Brazil

A comparative cytogenetic analysis was carried out on four Hylinae tree frogs from Brazil (Aparasphenodon brunoi, Corthomantis greeningi, Osteocephalus langsdorffi and Scinax fuscovarius) using Giemsa staining, BrdU replication banding, Ag-NOR staining, C-banding, DAPI and CMA(3) fluorochrome staining, and fluorescence in situ hybridization (FISH) with an rDNA probe. All the species share closely similar 2n = 24 karyotypes, almost indistinguishable by standard staining. The technique of BrdU incorporation allowed the identification of each pair of homologs and the establishment of extensive homeology for the great majority of the chromosomes, mainly of A. brunoi, C greeningi, and O. langsdorffii. Despite highly conserved replication banding patterns, the use of the other banding techniques disclosed some minor differsences, which reinforces the importance of extensive cytogenctic analyses for the karyotypic characterization of Anuran species. The present cytogenctic data confirm the closer proximity of A. brunoi, C greeningi, and O. langsdorfjii, whereas S. fuscovarius is phylogenetically more distant. Copyright (C) 2003 S. Karger AG, Basel

Multiple nucleolus organizer regions in Leptodactylus mystacinus (Amphibia, Anura) and comments on its systematic position in the L fuscus group based on cytogenetic and molecular analyses

Specimens of Leptodactylus mystacinus from Brazil were karyotyped with conventional and differential staining. The 2n = 22 karyotype is similar to that found for the majority of the Leptodactylus, the karyotypic conservatism also confirmed by the similarity of the replication banding patterns with those previously described. L. mystacinus has a small amount of C-banded heterochromatin, located mainly at the centromeres, although telomeric or interstitial bands have also been noticed. With DA/CMA(3) some chromosome regions showed slightly bright fluorescence, and with DA/DAPI, no particular AT-rich repetitive region was observed. Silver staining showed an extensive inter- and intraindividual variation in the number and position of Ag-positive regions, in 1p, 4p, 8p, 8q, and 11p. Nevertheless, FISH using rDNA probes confirmed only the signals on the short arms of chromosomes 4 and 8 as true NORs. The remaining silver stained regions are probably due to the heterochromatin with some affinity to the Ag-staining. Phylogenetic analysis based on partial cytochrome b sequence revealed that L. mystacinus forms a basal branch, so that the presence of multiple NORs in pairs 4 and 8 in this species indicates an autapomorphy.

Conserved karyotypes in the Hyla pukhella species group (Anura, Hylidae)

Cytogenetic analyses were done on specimens of Hyla marginata and on three populations of H. semiguttata differing in morphology and in the physical parameters of their advertisement call, as well as in individuals of Hyla sp. (aff. semiguttata). All specimens had 2n = 24 chromosomes with a morphology very similar to that of other 24-chromosome Hyla species. Hyla semiguttata and H. marginata showed the same C-banding pattern but were distinguished by the location of the NOR on pair 1 in H. semiguttata (in the three populations) and Hyla sp. (aff. semiguttata), and on pair 10 in H. marginata. The H. semiguttata populations did not differ cytogenetically, despite variations in their morphology and advertisement calls. Similarly, H. semiguttata and H. p. joaquini studied previously had identical C-banding patterns and NOR locations, suggesting that they are very closely related.

Chromosomal differentiation of populations of Lysapsus limellus limellus, L. l. bolivianus, and of Lysapsus caraya (Hylinae, Hylidae)

Cytogenetic analysis were done on specimens from two populations of Lysapsus limellus limellus. three of L. l. bolivianus and of one of Lysapsus caraya. All animals showed a diploid chromosomal number of 2n=24. The karyotypes of the two L. limellus subspecies were very similar, differing only by the larger amount of telomeric heterochromatin and a small pericentromeric C-band on the short arms of pair 2 in L. l. limellus specimens. The karyotype of L. caraya differed from those of the two L. limellus subspecies in terms of chromosomal morphology, C-banding pattern and location of the main NOR on chromosomes 7 and 6. respectively. The karyotype of the L. l. bolivianus population from Guajara-Mirim/RO differed from those of the other populations of the same subspecies in morphology and heterochromatin pattern of chromosomes 7 and 8. Additional NORs were detected by silver staining and confirmed by FISH in one of the homologues of pairs 1 and 8 in L. l. bolivianus and in pair 7 in L. caraya. These results suggest that a reassessment of the taxonomic status of L. limellus subspecies, especially of the L. l. bolivianus populations, may be necessary. (c) 2005 Elsevier Ltd. All rights reserved.

Molecular phylogeny and karyotype differentiation in Paratelmatobius and Scythrophrys (Anura, Leptodactylidae)

Paratelmatobius and Scythrophrys are leptodactylid frogs endemic to the Brazilian Atlantic forest and their close phylogenetic relationship was recently inferred in an analysis that included Paratelmatobius sp. and S. sawayae. To investigate the interspecific relationships among Paratelmatobius and Scythrophrys species, we analyzed a mitochondrial region (approximately 2.4 kb) that included the ribosomal genes 12S and 16S and the tRNAval in representatives of all known localities of these genera and in 54 other species. Maximum parsimony inferences were done using PAUP* and support for the clades was evaluated by bootstrapping. A cytogenetic analysis using Giemsa staining, C-banding and silver staining was also done for those populations of Paratelmatobius not included in previous cytogenetic studies of this genus in order to assess their karyotype differentiation. Our results suggested Paratelmatobius and Scythrophrys formed a clade strongly supported by bootstrapping, which corroborated their very close phylogenetic relationship. Among the Paratelmatobius species, two clades were identified and corroborated the groups P. mantiqueira and P. cardosoi previously proposed based on morphological characters. The karyotypes of Paratelmatobius sp. 2 and Paratelmatobius sp. 3 described here had diploid chromosome number 2n = 24 and showed many similarities with karyotypes of other Paratelmatobius representatives. The cytogenetic data and the phylogenetic analysis allowed the proposal/corroboration of several hypotheses for the karyotype differentiation within Paratelmatobius and Scythrophrys. Namely the telocentric pair No. 4 represented a synapomorphy of P. cardosoi and Paratelmatobius sp. 2, while chromosome pair No. 5 with interstitial C-bands could be interpreted as a synapomorphy of the P. cardosoi group. The NOR-bearing chromosome No. 10 in the karyotype of P. poecilogaster was considered homeologous to chromosome No. 10 in the karyotype of Scythrophrys sp., chromosome No. 9 in the karyotype of Paratelmatobius sp. 1, chromosome No. 8 in the karyotypes of Paratelmatobius sp. 2 and of Paratelmatobius sp. 3, and chromosome No. 7 in the karyotype of P. cardosoi. A hypothesis for the evolutionary divergence of these NOR-bearing chromosomes, which probably involved events like gain in heteochromatin, was proposed.

Conserved Karyotypes in Cophomantini: Cytogenetic Analysis of 12 Species from 3 Species Groups of Bokermannohyla (Amphibia: Anura: Hylidae)

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

A radiation hybrid map of river buffalo (Bubalus bubalis) chromosome 7 and comparative mapping to the cattle and human genomes

A preliminary radiation hybrid (RH) map containing 50 loci on chromosome 7 of the domestic river buffalo Bubalus bubalis (BBU; 2n = 50) was constructed based on a comparative mapping approach. The RH map of BBU7 includes thirty-seven gene markers and thirteen microsatellites. All loci have been previously assigned to Bos taurus (BTA) chromosome BTA6, which is known for its association with several economically important milk production traits in cattle. The map consists of two linkage groups spanning a total length of 627.9 cR(5,000). Comparative analysis of the BBU7 RH 5,000 map with BTA6 in cattle gave new evidence for strong similarity between the two chromosomes over their entire length and exposed minor differences in locus order. Comparison of the BBU7 RH 5,000 map with the Homo sapiens (HSA) genome revealed similarity with a large chromosome segment of HSA4. Comparative analysis of loci in both species revealed more variability than previously known in gene order and several chromosome rearrangements including centromere relocation. The data obtained in our study define the evolutionarily conserved segment on BBU7 and HSA4 to be between 3.5 megabases (Mb) and 115.8 Mb in the HSA4 (genome build 36) DNA sequence. Copyright (c) 2008 S. Karger AG, Basel.

Patterns of nucleolar activity during spermatogenesis of two triatomines, Panstrongylus megistus and P-herreri

The present work analyzed spermatogenesis in two species of triatomines (genus Panstrongylus) using silver-ion impregnation. The sex chromosomes of P. megistus and P. herreri had nucleolar organizing activity and became strongly impregnated during the phases of meiotic prophase I. Fragmentation of the nucleolus occurred in both species during the meiotic cycle. The nucleolar region could be observed up to diakinesis in meiotic prophase after which only nucleolar bodies and fragments were seen. Postmeiotic reactivation of rRNA synthesis occurred in these two species and was probably related to cell differentiation.

Meiosis aspects and nucleolar activity in Triatoma vitticeps (Triatominae, Heteroptera)

Some aspects of both the nucleolar organizer activity and meiosis were studied in the testes of Triatoma vitticeps (Heteroptera, Triatominae). The techniques used included squashing followed by lacto-acetic orcein staining, silver-ion impregnation, fluorescent banding (CMA(3), Quinacrine mustard and DAPI) and fluorescent in situ hybridization (FISH). A close relationship between heterochromatin and nucleolus in testicular cells was observed. During meiosis, the silver-ion impregnation pattern varied. At metaphase plate, a small body appeared apart from the chromosomes. In the spermatids this small body was seen in preparations stained with orcein and silver- ion impregnation but not with fluorochromes or FISH. These characteristics combined suggest that these corpuscles represent a source of ribonucleoproteins (RNP) - RNA and specific nucleolar proteins. Silver-ion impregnation and (FISH) revealed nucleolar organizer activity in two metaphase sex chromosomes (X). These results indicate that, in these species, nucleolar organizer regions (NORs) are located in the sex chromosomes, X chromosomes were were CMA(3)(+) and Y chromosome was DAPI(+).

Deletion (1)(q12) and double minutes in a metastatic adenocarcinoma of the prostate

Cytogenetic analysis after short-term cultures were performed on an invasive and moderately-differentiated prostatic adenocarcinoma. The results showed a normal male chromosomal complement in most metaphases examined. Furthermore, several abnormalities were found, including del(1)(q12), double minute and ring chromosomes, acentric fragments, triradial figures, and near-tetraploid cells. (C) Elsevier B.V., 1999. All rights reserved.

Detection of numerical chromosome anomalies in interphase cells of benign and malignant thyroid lesions using fluorescence in situ hybridization

Benign and malignant thyroid tumors constitute a wide range of neoplasias showing recurrent chromosome abnormalities. In an attempt to characterize specific numerical chromosome abnormalities in thyroid tissues, We present here the findings from a study of archival samples depicted by 10 malignant tumors, 30 benign lesions, and 10 normal thyroid tissues. Fluorescence in situ hybridization was performed on noncultured samples using biotinylated centromere-specific probes for chromosomes 7, 10, and 17. Trisomy or tetrasomy 7 were present in 19 benign and in 7 malignant tumors. Trisomy 10 or 17 were observed in 18 adenomas or goiters and in 9 carcinomas, and monosomy 17 was seen in 2 carcinomas. Our findings suggest that such abnormalities are an in vivo phenomenon and may be important in the neoplastic proliferation of thyroid gland. (C) Elsevier B.V., 2000. All rights reserved.

What influences Hb fetal production in adulthood?

Human hemoglobin genes are located in α and β globin gene clusters in chromosomes 16 and 11, respectively. Different types of hemoglobin are synthesized according to the stage of development with fetal hemoglobin (α2γ2) (Hb F) being the main hemoglobin in the fetal period. After birth, there is a reduction (to about 1%) in Hb F levels and adult hemoglobin, Hb A (2α2β2), increases to more than 96% of total hemoglobin. However, some genetic conditions whether linked to the β-globin gene cluster or not are associated with high Hb F levels in adults. Among those linked to β-globin are hereditary persistence of fetal hemoglobin, delta-beta thalassemia (δβ-Thalassemia) and the XmnI polymorphism (-158 C > T). Other polymorphisms not related to β-globin gene cluster are known to influence the γ-globin gene expression in adulthood. The most relevant polymorphisms that increase concentrations of Hb F are the HMIP locus on chromosome 6, the BCL11A locus on chromosome 2, the Xp22.2 region of the X chromosome and the 8q region on chromosome 8. Findings from our research group studying genetic factors involved in γ-globin gene regulation in adults without anemia in the northwestern region of São Paulo State showed that high Hb F levels are influenced by the presence of hereditary persistence of fetal hemoglobin mutations and the XmnI polymorphism, suggesting that both genetic alterations characterize the molecular basis of the evaluated population.