A natural triploid in Trichomycterus davisi (Siluriformes, Trichomycteridae): mitotic and meiotic characterization by chromosome banding and synaptonemal complex analyses

Within a total of 50 analyzed specimens a male individual of Trichomycterus davisi has been recorded with 81 chromosomes including 60 metacentric, 18 submetacentric and three subtelocentric chromosomes. When compared with diploid individuals (2n = 54) and the morphological standard of chromosomes, this male is a triploid with 3 = 81 chromosomes. Since staining with silver nitrate indicates three active nucleolar organizer regions (NORs), the three NOR- bearing chromosomes in this individual are genetically active. Analysis of the synaptonemal complex (SC) by electronic microscopy shows that there is an incomplete pairing of the third set of chromosomes in the triploid individual.

DIFFERENTIATION OF DROSOPHILA-SERIDO (ISOFEMALE LINE A95F3) AND D-KOEPFERAE (ISOFEMALE LINE B20D2) - REPRODUCTIVE ISOLATION, DEVELOPMENT TIME AND POLYTENE CHROMOSOME-BANDING PATTERNS

Drosophila serido is considered to be a superspecies consisting of two species: D. serido, from Brazil and D. koepferae from Argentina and Bolivia. However this probably does not express the entire evolutionary complexity of its populations. Isofemale lines A95F3 (from Brazil) and B20D2 (from Argentina), at present representing, respectively, the first and second species, were analyzed for fertility and fecundity in pair-mating intracrosses and intercrosses, as well as for development time, banding patterns and asynapsis of polytene chromosomes in the isofemale lines and their hybrids.Although variations in experimental conditions resulted in some variability in the results, in general A95F3 fertility and fecundity were lower than in B20D2. Intercrosses of A95F3 females and B20D2 males showed lower fertility and fecundity than the reciprocal crosses, following more closely characteristics of the mother strains. This is in contrast to the results obtained by Fontdevilla et al. (An. Entomol. Soc. Amer. 81: 380-385, 1988) and may be due to the different geographic origin of D. serido strains they used in crosses to B20D2. This difference and others cited in the literature relative to aedeagus morphology, karyotype characteristics, inversion polymorphisms and reproductive isolation strongly indicate that A95F3 and D. serido from the State of Bahia, Brazil are not a single evolutionary entity, reinforcing the idea of greater complexity of the superspecies D. serido than is known today.The reproductive isolation mechanisms found operating between A95F3 and B20D2 were prezygotic and postzygotic, the latter included mortality at the larvae stage in both directions of crosses and sterility of male hybrids in intercrosses involving B20D2 females and A95F3 males. The two isofemale lines differed in egg-adult development time, which was also differently affected by culture medium composition.A95F3 and B20D2 also showed differences in the banding patterns of proximal regions of polytene chromosomes 2, 3 and X, a fixed inversion in chromosome 3 (here named 3t), apparently not described previously, and a high degree of asynapsis in hybrids.These observations, especially those related to reproductive isolation and chromosomal differentiation (including the karyotype, previously described, and the differentiation of banding patterns, described in this paper), as well as the extensive asynapsis observed in hybrids reinforces the distinct species status of A95F3 and B20D2 isofemale lines.

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.

Cytogenetics of three breeds of river buffalo (Bubalus bubalis L.), with evidence of a fragile site on the X chromosome

The cytogenetic study of 182 river buffalo (Bubalus bubalis L., 2n=50) of Murrah, Mediterranean and Jaffarabadi breeds, from the State of São Paulo, was carried out to characterize their chromosomes and to detect possible chromosomal abnormalities. The karyotypes were indistinguishable with conventional staining as well as with C and replication R banding techniques. In about 44% of the sample (8 males and 72 females), an X marker chromosome due to a fragile site was shown. The frequency of metaphases expressing the fragility site on the X was highly variable, from 2.86 to 41.03%. In females, the fragile site, rarely appeared on both X chromosomes. Most of the metaphases showed only 1 marker chromosome. In R-banded metaphases using 5-bromodeoxyuridine (BrdU) treatment, it corresponded in general to the late replicating X chromosome. No correlation between the X fragile site and altered phenotype was found. Structural and numerical chromosome rearrangements were ruled out in the present sample of buffalo. (C) 1998 by Elsevier B.V.

Heterochromatic banding pattern in two Brazilian populations of Aedes aegypti

We analysed samples of Aedes aegypti from Sao Jose do Rio Preto and Franca (Brazil) by C-banding and Ag-banding staining techniques. C-banding pattern of Ae. aegypti from Sao Jose do Rio Preto examined in metaphase cells differed from Franca. The chromosomes 2, 3 and X showed centromeric C-bands in both populations, but a slightly stained centromeric band in the Y chromosome was observed only in Sao Jose do Rio Preto. In addition, the X chromosome in both populations and the Y chromosome of all individuals from Sao Jose do Rio Preto showed an intercalary band on one of the arms that was absent in Franca. An intercalary, new band, lying on the secondary constriction of chromosome 3 was also present in mosquitoes of both populations. The comparison of the present data with data in the literature for Ae. aegypti from other regions of the world showed that they differ as to the banding pattern of sex chromosomes and the now described intercalary band in chromosome 3. The observations suggested that the heterochromatic regions of all chromosomes are associated to constitute a single C-banded body in interphase cells. Ag-banding technique stained the centromeric regions of all chromosomes (including the Y) and the intercalary C-band region of the X chromosome in both populations. As Ae. aegypti populations are widespread in a great part of the world, the banding pattern variations indicate environmental interactions and may reveal both the chromosome evolutionary patterns in this species and the variations that may interfere with its vector activity.

Restriction endonuclease banding and digestion resistant heterochromatin in triatomines, genus Panstrongylus

The present work realized a comparative study in meiosis of two triatomines, Panstrongylus herreri and P. megistus, by cytogenetic techniques involving the restriction endonucleases Hae III and Alu I and C-banding. The system of sex chromosomes in Panstrongylus is of the X1X2Y type, and experiments corroborated the common origin hypothesis of the X chromosomes by fragmentation of single X. In both species the restriction endonucleases (RE) presented banding patterns in part similar to C-banding. However, in some early meiotic phases it was possible to verify differentiation of the heterochromatic pattern. This work suggests that other elements besides presence of recognition sites, such as chromatin packing degree and DNA-protein interaction, act in RE results, since digestion patterns occur in early spermatogenesis. However, metaphase chromosomes were practically inaccessible to the endonucleases.

Application of C-banding in two Arachis wild species, Arachis Pintoi Krapov. and W.C. Gregory and A. villosulicarpa Hoehne to mitotic chromosome analyses

Two wild diploid (2n = 20 chromosomes) and self-pollinating Arachis species, Arachis Pintoi Krapov and W.C. Gregory and A. villosulicarpa Hoehne were submmited to C-band technique to karyotype analyses. Root tips were employed in the analyses. Morphometric data chose that chromosome lengths varied from 3.12 in A. villosulicarpa to 1.45 in A. Pintoi. Karyotype formula obtained was 10sm to A. Pintoi and 9sm + 1m to A. villosulicarpa. There was a predominance of pericentromeric C-band in all mitotic metaphasic chromosomes in both species. Besides C-band values, both species still did not differ in respect to chromosome absolute and relative lengths, centromeric index, symmetry index and total karyotype haploid length. C-band and morphometric data did not show strong or significant differences which could separate these two species of peanut which belong to evolutive different sections.

Pathological and molecular analysis of Turkey coronavirus replication in captive wild Turkey embryos (broad-breasted bronze breeder)

The degree of genetic and pathologic variation exhibited by a turkey Coronavirus (TCoV) strain was investigated after nine serial passages in 25-day-old turkey embryos obtained from wild broad-breasted bronze breeders. In spite of spleen, liver, kidneys, cloacal bursa and thymus have been collected and analysed, the main histopathological changes were only documented in the intestine sections. Microscopic lesions were characterized as mild enteritis, low degree of enterocyte vacuolization and detachment of the intestinal villous after five consecutive passages and were considered absent in the last passages. Genealogic analysis based on S1 and S2 DNA sequences suggested that Brazilian isolate might be considered as originated from TCoV strains circulating in the United States, as 100% identity with TCoV-Gl strain. Although S1 S2 sequences from each passage revealed no significant point mutations, and no correlation could be speculate between S2 nucleotide changes and pathologic features in infected embryos. This is the first demonstration of wild turkey embryos as a model for TCoV isolation and propagation.

The eukaryotic translation initiation factor 3 subunit L protein interacts with Flavivirus NS5 and may modulate yellow fever virus replication

Background: Yellow fever virus (YFV) belongs to the Flavivirus genus and causes an important disease. An alarming resurgence of viral circulation and the expansion of YFV-endemic zones have been detected in Africa and South America in recent years. NS5 is a viral protein that contains methyltransferase and RNA-dependent RNA polymerase (RdRp) domains, which are essential for viral replication, and the interactions between NS5 and cellular proteins have been studied to better understand viral replication. The aim of this study was to characterize the interaction of the NS5 protein with eukaryotic translation initiation factor 3 subunit L (eIF3L) and to evaluate the role of eIF3L in yellow fever replication. Methods. To identify interactions of YFV NS5 with cellular proteins, we performed a two-hybrid screen using the YFV NS5 RdRp domain as bait with a human cDNA library, and RNApol deletion mutants were generated and analyzed using the two-hybrid system for mapping the interactions. The RNApol region involved was segmented into three fragments and analyzed using an eIF3L-expressing yeast strain. To map the NS5 residues that are critical for the interactions, we performed site-direct mutagenesis in segment 3 of the interaction domain (ID) and confirmed the interaction using in vitro assays and in vivo coimmunoprecipitation. The significance of eIF3L for YFV replication was investigated using eIF3L overexpression and RNA interference. Results: In this work, we describe and characterize the interaction of NS5 with the translation factor eIF3L. The interaction between NS5 and eIF3L was confirmed using in vitro binding and in vivo coimmunoprecipitation assays. This interaction occurs at a region (the interaction domain of the RNApol domain) that is conserved in several flaviviruses and that is, therefore, likely to be relevant to the genus. eIF3L overexpression and plaque reduction assays showed a slight effect on YFV replication, indicating that the interaction of eIF3L with YFV NS5 may play a role in YFV replication. Conclusions: Although the precise function of eIF3L on interactions with viral proteins is not entirely understood, these results indicate an interaction of eIF3L with YF NS5 and that eIF3L overexpression facilitates translation, which has potential implications for virus replication. © 2013 Morais et al.; licensee BioMed Central Ltd.