Evolutionary chromosomal differentiation among four species of Conoderus Eschscholtz, 1829 (Coleoptera, Elateridae, Agrypninae, Conoderini) detected by standard staining, C-banding, silver nitrate impregnation, and CMA(3)/DA/DAPI staining

The speciose Brazilian Elateridae fauna is characterized by high karyotypic diversity, including one species (Chalcolepidius zonatus Eschscholtz, 1829) with the lowest diploid number within any Coleoptera order. Cytogenetic analysis of Conoderus dimidiatus Germar, 1839, C. scalaris (Germar, 1824,) C. ternarius Germar, 1839, and C. stigmosus Germar, 1839 by standard and differential staining was performed with the aim of establishing mechanisms of karyotypic differentiation in these species. Conoderus dimidiatus, C. scalaris, and C. ternarius have diploid numbers of 2n(male) = 17 and 2n(female) = 18, and a X0/XX sex determination system, similar to that encountered in the majority of Conoderini species. The karyotype of C. stigmosus was characterized by a diploid number of 2n=16 and a neoXY/neoXX sex determination system that was highly differentiated from other species of the genus. Some features of the mitotic and meiotic chromosomes suggest an autosome/ancestral X chromosome fusion as the cause of the neoXY system origin in C. stigmosus. C-banding and silver impregnation techniques showed that the four Conoderus species possess similar chromosomal characteristics to those registered in most Polyphaga species, including pericentromeric C band and autosomal NORs. Triple staining techniques including CMA(3)/DA/DAPI also provided useful information for differentiating these Conoderus species. These techniques revealed unique GC-rich heterochromatin associated with NORs in C. scalaris and C. stigmosus and CMA(3)-heteromorphism in C. scalaris and C. ternarius.

Predisposing genes and increased chromosome aberrations in lung cancer cigarette smokers

Genotoxic effects linking cigarette smoking with lung cancer have not been consistently demonstrated, therefore claims for the cause-effect relationships are vigorously contested. Using matched populations of 22 lung cancer patients who have been cigarette smokers (LCP), 22 non-cancerous cigarette smokers (SC) and 13 non-smokers (NSC), we have applied the fluorescence in situ hybridization (FISH) tandem probe assay to elucidate the frequency of chromosome breakage among the participants. Two probes were used, a classical satellite probe which hybridizes to the large heterochromatin region of chromosome 1, and an alpha-satellite probe which targets a small region adjacent to the heterochromatin probe. The highest frequency of structural aberrations was observed in LCP (1.4 +/- 0.1) followed by SC (1.25 +/- 0.1) and NSC (0.4 +/- 0.1). Aberration frequencies were not significantly different between LCP and SC (p > 0.05), however, a statistically significant difference was detected between the smoker populations combined (LCP and SC) and the NSC (p < 0.001). The breakage frequencies showed a positive correlation with duration of smoking for LCP (r = 0.5; p < 0.01), but not for SC (P > 0.05). In addition, the aberration frequencies were influenced by the inheritance of polymorphic glutathione S-transferase (GST) genes. LCPs missing one or the other GST (GSTM1 or GSTT1) genes were found to have significantly higher chromosome breaks compared to LCPs with both genes present (p < 0.05), Our data indicate that genetic predisposition and chromosome aberrations may be mechanistically related to the initiation of lung carcinogenesis; therefore, they may be useful biomarkers for lung cancer among cigarette smokers. (C) 1997 Elsevier B.V. B.V.

EFFECT OF PRETREATMENT WITH VENOM OF APIS-MELLIFERA BEES ON THE YIELD OF GAMMA-RAY INDUCED CHROMOSOME-ABERRATIONS IN HUMAN BLOOD-LYMPHOCYTES

Venom of the honey bee Apis mellifera induced a protective effect against the induction of dicentric chromosomes by gamma radiation (2.0 Gy) in human peripheral blood lymphocytes which the cultures were treated with 0.00015 mul venom/1 ml medium 6 h before irradiation. In cultures to which the venom was added immediately before irradiation with 0.25, 1.0 and 2.0 Gy, no significant differences in number of dicentric chromosomes induced was observed when compared to cultures submitted to irradiation only. The venom did not induce clastogenic effects nor did it increase the frequency of sister chromatid exchanges.

Differential susceptibility to chromatid breaks induced by bleomycin in sub-fertile and fertile bovines

The rate of chromatid breaks was studied in cows with a history of sub-fertility by means of a test based on measurement of the average of breaks induced in lymphocytes of peripheral blood cultures. Fourteen female specimens were divided into two groups: fertile and sub-fertile. Peripheral blood lymphocytes were cultured and prepared for cytogenetic analysis. Two types of culture were established for each animal to evaluate the response of peripheral blood lymphocyte cultures to the genotoxic effects of bleomycin. The first culture did not receive bleomycin treatment (spontaneous chromosome aberrations). Our results showed that median breaks per cell (b/c) (+/-semirange) for spontaneous culture of the fertile and sub-fertile animals and bleomycin sensitivity assay for fertile and sub-fertile animals were 0.00 +/- 0.06, 0.02 +/- 0.03, 0.08 +/- 0.05 and 0.22 +/- 0.09, respectively. There was no significant difference (P > 0.05) in the chromosomal breakage in lymphocytes not exposed to bleomycin; however, in comparing the number of chromatid breaks per cell in cultures treated with bleomycin, the sub-fertile group showed a significantly higher (P < 0.05) level than the fertile group. These findings have implications both for identifying cattle with less than optimum fertility as well as for providing potential avenues to study the origins of sub-fertility. (C) 2004 Elsevier B.V. All rights reserved.

Chromosomal studies on five species of the genus Leptodactylus Fitzinger, 1826 (Amphibia, Anura) using differential staining

Cytogenetic studies were carried out on five species of Leptodactylus, namely L. fuscus, L, notoaktites, L. labyrinthicus, L. ocellatus, and L. podicipinus, after standard staining, Ag-NOR and C-banding as well as BrdU incorporation for three of them. The species had 2n = 22 chromosomes and two basic karyotype patterns. Chromosome 8 was a marker bearing a secondary constriction. In all species, this secondary constriction corresponded to the Ag-NOR site. The species had centromeric C-bands in all chromosomes of the complement, but some interstitial or telomeric bands seemed to differentiate some karyotypes, either at the species or the population level. In L. ocellatus, the C-banding pattern confirmed the occurrence of a heteromorphic pericentric inversion in chromosome 8 in specimens from one of the populations. The BrdU incorporation technique showed no detectable difference in the replication patterns of the major bands in the chromosomes of L. noroaktites, L. labyrinthicus, and L. ocellatus.

Chromosomal analysis of the leptodactylids Pleurodema diplolistris and Physalaemus nattereri (Amphibia, Anura)

Detailed characterizations of the karyotypes of the Brazilian leptodactylid frogs Pleurodema diplolistris, the only species of Pleurodema not studied cytogenetically so far, and Physalaemus nattereri, a species in the Ph. biligonigerus group, are presented. Both karyotypes had 2n = 22 and their chromosomes had a very similar morphology, except for pair 11, which was metacentric in Pl. diplolistris and telocentric in Ph. nattereri. The localization of nucleolar organizer regions (NORs) and heterochromatic bands allowed the differentiation of chromosomes that were morphologically indistinguishable between these species, such as pairs 1, 3 and 10, which showed interstitial C-bands in Ph. nattereri, and pair 8, that had an NOR and an adjacent C-band in Pl. diplolistris. Pair 8 also has NOR-bearing chromosomes in many other Pleurodema species. However, in these species, the NOR is located proximal to the centromere on the short arm, while in Pl. diplolistris it occurred distally on the long arm, a condition that may be considered a derived state. In Ph. nattereri, the NOR occurred on chromosome I 1 and differed from the other species of the Ph. biligonigerus group. In contrast, C-banding revealed a heterochromatic block near the centromere on the short arm of pair 3, a characteristic common to all members of this group of Physalaemus.

CHROMOSOMAL CONSTITUTION IN ROB (5/15) GOAT PROGENY

A total of 116 goats originating from three heterozygous bucks for rob (5/15) were cytogenetically studied at Botucatu, Brazil. In the mating 59,XY,t(5/15) x 60,XX, the offspring karyotype proportion did not differ significantly from the expected ratio of 1 normal:1 heterozygous. In the mating 59,XY,t(5/15) x 59,XX,t(5/15) the observed proportion was 1 normal:2 heterozygous:1 homozygous, and in the mating 59,XY,t(5/15) X 58,XX,tt(5/15) the observed proportion was 1 heterozygous:1 homozygous. No animals had unbalanced karyotypes and no differences in the weight at birth of normal, heterozygous and homozygous kids were observed (P > 0.10).

A LINE2 repetitive DNA sequence from the cichlid fish, Oreochromis niloticus: sequence analysis and chromosomal distribution

We report the cloning and characterization of a long interspersed nucleotide element (LINE) fi-om a cichlid fish, Oreochromis niloticus, and show the distribution of this element, called CiLINE2 for cichlid LINE2, in the chromosomes of this species. The identification of an open reading frame in CiLINE2 with amino acid sequence similarity to reverse transcriptases encoded by LINE-like elements in Caenorhabditis elegans, Platemys spixii, Schistosoma mansoni, Gallus gallus (CRI), Drosophila melanogaster (I factor), and Homo sapiens (LINE2), as well as the structure of the element, suggest it is a member of this family of non-long terminal repeat-containing retrotransposons. Search of a DNA sequence database identified sequences similar to CiLINE2 in four other fish species (Haplotaxodon microlepis, Oreochromis mossambicus, Pseudotropheus zebra, and Fugu rubripes). Southern blot hybridization experiments revealed the presence of sequences similar to CiLINE2 in all Tilapiini species analyzed from the genera Oreochromis, Tilapia, and Sarotherodon, and gave an estimated copy number of about 5500 for the haploid genome of O. niloticus. Fluorescent in situ hybridization showed that CiLINE2 sequences were organized in small clusters dispersed over all chromosomes of O. niloticus, with a higher concentration near chromosome ends. Furthermore the long arm of chromosome 1 was strikingly enriched with this sequence. The distribution of LINE2-related elements might underlie the difference in chromosome banding patterns observed between cold-blooded vertebrates and mammals.

Electrochemical measurements of oligonucleotides in the presence of chromosomal DNA using membrane-covered carbon electrodes

Substantial improvements in the selectivity of electrochemical measurements of trace nucleic adds are obtained by using membrane-covered carbon disk electrodes. Access to the electrode surface can be manipulated via a judicious choice of the membrane molecular weight cutoff (MWCO). The resulting separation step, performed in situ at the electrode surface, adds a new dimension of selectivity based on molecular size to electroanalysis of nucleic acids, Transport properties are evaluated with respect to the oligonucleotide length and membrane MWCO. A highly selective response is observed for synthetic oligonucleotides in the presence of otherwise interfering chromosomal DNAs. Discrimination among oligonucleotides of different lengths is also possible, Short accumulation periods (1-5 min) are sufficient for convenient measurements of low milligram per liter concentrations.

Chromosome aberrations as a predictor of clinical outcome for smoking associated lung cancer

The ability to identify individuals at greatest risk of developing lung cancer can significantly enhance the efficacy of intervention modalities. One strategy for identifying these individuals is through biomarkers that reflect the severity of their cancer. In the present study, we evaluated 22 lung cancer patients and 35 controls to determine whether the frequency of chromosome aberrations was significantly associated with specific clinical variables such as the histological type, grade and stage of the turners. Chromosome aberrations (expressed as total breaks) were investigated on chromosome 1 in interphase nuclei obtained from blood Lymphocytes of the study participants using the fluorescence in situ hybridization (FISH) chromosome aberration assay. Our results indicate a significant linear increase (P = 0.01) in the level of breaks with respect to the grade of the carcinoma. The poorly differentiated tumors had a significantly higher level of chromosome breaks mean +/- SD (1.7 +/- 0.46) as compared to the well differentiated tumors (0.98 +/- 0,23, P < 0,05). These results indicate that chromosome aberrations, as determined by the FISH assay, can be used as a biomarker for identifying individuals with aggressive types of lung cancer and potentially, as a predictor for prognostic outcome of the disease. (C) 2000 Elsevier B.V. Ireland Ltd. All rights reserved.

Chromosomal differentiation of Hyla nana and Hyla sanborni (Anura, Hylidae) with a description of NOR polymorphism in H-nana

Specimens of Hyla nana and Hyla sanborni from a syntopic population were studied cytogenetically. These species are morphologically very similar and are frequently misidentified, confused with each other. Both species had a diploid chromosome number, 2n = 30. However, the karyotypes of H. nana and H. sanborni differed considerably from each other in the number of submetacentric and telocentric chromosomes. The two species also differed in their primary NOR-bearing chromosomes (metacentric pair 13 in H. nana and telocentric pair 12 in H. sanborni). Additional nucleolus organizer regions (NORs) were detected by Ag-NOR staining and FISH in chromosome pairs 1, 5, 6, 12, and 14 in seven specimens of H. nana. Thus, a total of six patterns of NOR were identified. These differences in karyotype and in NOR location allowed the unambiguous identification of syntopic individuals of the two species. However, the chromosomal morphology of both species differed from that reported for populations from other geographic regions, suggesting that a systematic reevaluation of this group of Hyla may be necessary.