Isolation of viable Toxoplasma gondii from naturally infected aborted bovine fetuses

Neospora caninum and Toxoplasma gondii are related parasites. The former is a common cause of abortion in dairy cattle. The latter has not been conclusively demonstrated in bovine fetuses. During the course of attempts to isolate N. caninum from aborted fetuses, T. gondii was isolated from 2 aborted fetuses, 1 from Portugal and 1 from the United States. Both isolates were made by bioassay of fetal brains in mice. The fetus from Portugal was about 5 mo in gestational age, and the fetus from the United States was a full-term stillborn.     

Improvement of DNA transfection with cationic liposomes

The increasing use of cationic liposomes as vectors for DNA transfection of eukaryotic cells is due to its high efficiency and reproducibility. After the interaction of the DNA cationic-liposome complexes (DNA-CLC) with the plasma membrane, the entry into the cells delivers the DNA-CLC to the endosome-lysosome pathway where some of the DNA-CLC are degraded. The non-degraded DNA that escapes to the cytoplasm, still has to transverse the nuclear membrane to be transcribed and then translated. To improve the efficiency of the whole process, we can manipulate the DNA (sequences, promoters, enhancers, nuclear localisation signals, etc), the DNA-CLC (lipids) or the plasmatic, endosomal and/or nuclear cellular membranes (ultrasound, electroporation, Ca++, pH of the endosomes, mitosis, fusogenic peptides, nuclear localisation signals, etc). Most of these methods have been generally used individually but in combination, may greatly improve the efficiency and reproducibility of in vitro transfection. While much of this work remains yet to be done and present results further explored, the application of these efforts is essential to the future development of new gene therapy strategies.     

Total synthesis and evaluation of lamellarin alpha 20-Sulfate analogues

In order to explore the influence of sulfate groups on the bioactivity profiles of marine alkaloids of the lamellarin class, three such alkaloids, lamellarin alpha, lamellarin alpha 13,20-disulfate and lamellarin H, were synthesized and their activities against HIV-1 integrase and cancer cell lines were compared with those of lamellarin alpha 20-sulfate, which is a selective inhibitor of HIV-1 integrase. Lamellarin alpha does not inhibit HIV-1 integrase but shows moderate cytotoxicity with good cell line selectivity. Lamellarin alpha 13,20-disulfate is a moderate inhibitor of both HIV-1 integrase and cancer cell lines. Lamellarin H is a more potent inhibitor of HIV-1 integrase but lacked the specificity required to be medicinally useful.     

A new family of highly variable proteins in the Chlamydophila pneumoniae genome

Chlamydiaceae are obligate intracellular bacterial pathogens characterized by a wide range of vertebrate host, tissue tropism and spectrum of diseases. To get insights into the biological mechanisms involved in these differences, we have put forward a computational and experimental procedure to identify the genome recombination hotspots, as frequent sequence variation allows rapid adaptation to environmental changes. We find a larger potential for recombination in Chlamydophila pneumoniae genomes as compared with Chlamydia trachomatis or Chlamydia muridarum. Such potential is mostly concentrated in a family of seven previously uncharacterized species-specific elements that we named ppp for C.pneumoniae polymorphic protein genes, which have the potential to vary by homologous recombination and slipped-mispair. Experimentally, we show that these sequences are indeed highly polymorphic among a collection of nine C.pneumoniae strains of very diverse geographical and pathological origins, mainly by slippage of a poly(C) tract. We also show that most elements are transcribed during infection. In silico analyses suggest that Ppps correspond to outer membrane proteins. Given their species specificity, their putative location in the outer membrane and their extreme polymorphism, Ppps are most likely to be important in the pathogenesis of C.pneumoniae and could represent targets for future vaccine development.     

Origin and fate of repeats in bacteria

We investigated 53 complete bacterial chromosomes for intrachromosomal repeats. In previous studies on eukaryote chromosomes, we proposed a model for the dynamics of repeats based on the continuous genesis of tandem repeats, followed by an active process of high deletion rate, counteracted by rearrangement events that may prevent the repeats from being deleted. The present study of long repeats in the genomes of Bacteria and Archaea suggests that our model of interspersed repeats dynamics may apply to them. Thus the duplication process might be a consequence of very ancient mechanisms shared by all three domains. Moreover, we show that there is a strong negative correlation between nucleotide composition bias and the repeat density of genomes. We hypothesise that in highly biased genomes, non-duplicated small repeats arise more frequently by random effects and are used as primers for duplication mechanisms, leading to a higher density of large repeats.     

Variation in fruit and seed size from 38 wild populations of Phaseolus lunatus (Fabaceae) from Central Valley, Costa Rica

We studied the morfological diversity in fruits and seeds in 38 wild populations of Phaseolus lunatus var. lunatus (lima beans) in the central valley of Costa Rica. In order to do so, measured the length and width of the fruits and the length, width and thickness of seeds. We also calculated the ratio between these traits and determined the weight of 100 seeds. In general, we found significant variation between populations for all variables. When we grouped the 38 populations into eight geographical regions within the study area, we found significant differences between regions. However, the levels of variation between populations within geographical regions was larger than that found between geographical regions. These findings suggested that there is no clear relationship between these variables and the geographical grouping established in this study. The implications of these findings for the establishment of strategies for in situ conservation of wild populations of lima beans are discussed.     

A stereological study of medium antral follicles during the bovine estrous cycle

Stereological methods were applied to bovine cumulus-oocyte complexes (COCs) in order to characterize them quantitatively during the estrous cycle. COCs from medium (4-8mm) antral follicles with a compact and complete cumulus mass, and with an uniform or a non-visible ooplasm were aspirated from ovaries of Holstein-Friesian cows, fixed in glutaraldehyde, randomly embedded in glycol-methacrylate, and sectioned at 20 microm. The unbiased nucleator principle was used for estimating the mean volumes of complexes, oocytes, cumulus cells, and nuclei of oocytes and cumulus cells. The thickness of the zona pellucida and the relative numerical percentages of the several morphological types (C1-C3) of cumulus cells were also evaluated. The optical disector procedure was used for cumulus cell sampling. Quantitative data show that COCs appear heterogeneous for all studied parameters. From metestrus to proestrus the volumes of COCs and oocytes remained constant, the volumes of oocytes and oocyte nuclei were correlated, the thickness of the outer zona pellucida decreased, and the relative numerical frequency of follicular type C3 cells increased. Results suggest that COCs from distinct estrus stages are structurally different, with type C3 follicular cells gradually differentiating from cell types C1 and C2.     

Host-ectoparasite specificity in a small mammal community in an area of Atlantic Rain Forest (Ilha Grande, State of Rio de Janeiro), Southeastern Brazil

The analyses of the ectoparasite species associated with a small mammal community on Ilha Grande, a coastal island in southern of the state of Rio de Janeiro, Brazil, evaluated the level of host-ectoparasite specificity. Was used the Jaccard index for qualitative data to analyse the similarity. The lowest value of similarity occurred between Proechimys iheringi and Marmosops incanus and between Sciurus aestuans and Nectomys squamipes (Cj=0.08) and the highest between P. iheringi and Oxymycterus sp. (Cj=0.33). This index showed a low value of similarity across the ectoparasite community. The only exception from this pattern of high host specificity occurred with P. iheringi and Oxymycterus sp., which shared five species of ectoparasites. The similarity values, for most of the cases, is smaller than 0.2.