Revealing the latent mobilization capability of the staphylococcal bacteriocinogenic plasmid pRJ9

Plasmid pRJ9 is a non-self-mobilizable bacteriocinogenic plasmid from Staphylococcus aureus. Despite this feature, DNA sequencing and RT-PCR experiments showed that it presents a Mob region with three genes (mobCAB), transcribed as an operon. In silico analysis of the Mob proteins encoded by pRJ9 showed that they present all the conserved functional features reported until present as being essential for plasmid mobilization. Moreover, they showed a high identity to Mob proteins encoded by mobilizable plasmids from Staphylococcus spp., especially to those encoded by plasmid pRJ6, which presents four mob genes (mobCDAB). A putative oriT region was also found upstream of the pRJ9 mob operon. pRJ9 could only be successfully mobilized by pGO1 when pRJ6 was present in the same strain. Further experiments showed that the pRJ9 oriT can be recognized by the pRJ6 Mob proteins, confirming its functionality. As pRJ9 does not possess a mobD gene while pRJ6 does, the absence of this gene was believed to be responsible for its lack of mobilization. However, conjugation experiments with a donor strain carrying also mobD cloned into an S. aureus vector showed that pRJ9 does not become mobilized even in the presence of the protein MobD encoded by pRJ6. Therefore, the reasons for pRJ9 failure to be mobilized are presently unknown.     

Modulatory effects of Duguetia furfuracea (A. St. Hil) Benth. and Hook. f. in Drosophila melanogaster somatic and germinative cells

Mutagenic and antimutagenic activities of the medicinal plant Duguetia furfuracea were assessed using SMART/wing and ring-X-loss tests. For the ring-X-loss test, 2- to 3-day-old Drosophila melanogaster ring-X-lineage males and virgin ywsn3 females received D. furfuracea infusion at doses of 0.085, 0.042, or 0.014 g/mL for 24 h. We found that D. furfuracea did not produce any mutagenic effects in D. melanogaster germinative cells. The somatic cells of D. melanogaster were analyzed using the SMART/wing test involving three lineages - mwh, flr3, and ORR - and the same doses of D. furfuracea infusion employed in the ring-X-loss test, as well as 20 mM urethane. The results of both standard (ST) and high bioactivation (HB) crosses showed absence of mutagenic activity of D. furfuracea. In contrast, in both ST and HB crosses, we observed a modulatory effect of D. furfuracea against the genotoxic activity of urethane.     

Microsatellite variation and evolution of human lactase persistence

The levels of haplotype diversity within the lineages defined by two single-nucleotide polymorphisms (SNPs) (–13910 C/T and –22018 G/A) associated with human lactase persistence were assessed with four fast-evolving microsatellite loci in 794 chromosomes from Portugal, Italy, Fulbe from Cameroon, São Tomé and Mozambique. Age estimates based on the intraallelic microsatellite variation indicate that the –13910*T allele, which is more tightly associated with lactase persistence, originated in Eurasia before the Neolithic and after the emergence of modern humans outside Africa. We detected significant departures from neutrality for the –13910*T variant in geographically and evolutionary distant populations from southern Europe (Portuguese and Italians) and Africa (Fulbe) by using a neutrality test based on the congruence between the frequency of the allele and the levels of intraallelic variability measured by the number of mutations in adjacent microsatellites. This result supports the role of selection in the evolution of lactase persistence, ruling out possible confounding effects from recombination suppression and population history. Reevaluation of the available evidence on variation of the –13910 and –22018 loci indicates that lactase persistence probably originated from different mutations in Europe and most of Africa, even if 13910*T is not the causal allele, suggesting that selective pressure could have promoted the convergent evolution of the trait. Our study shows that a limited number of microsatellite loci may provide sufficient resolution to reconstruct key aspects of the evolutionary history of lactase persistence, providing an alternative to approaches based on large numbers of SNPs.Electronic supplementary material Supplementary material is available for this article at     

Effect of Different Carbon Sources on Endochitinase Production by Colletotrichum gloeosporioides

The present work analyzes the production of endochitinase by Colletotrichum gloeosporioides, a phytopathogenic fungus, using six different carbon sources and two pH values. For quantitative assay of endochitinase activity in solution, the synthetic substrate 4-methylumbelliferyl-β-D-N,N’,N”-triacetylchitotrioside was used. The major productions were obtained at pH 7.0 and 9.0, when colloidal chitin and glucose were used, whereas xylose and lactose were not good carbon sources. When testing different concentrations of colloidal chitin, glucose and glucosamine, colloidal chitin 0.5% was the best substrate, giving values of 2.4 U at the fifth day. When using glucose, best production occurred at 0.3% concentration, after 5 days growth, with values of 1.31 U. Endochitinase production was markedly decreased in high levels of glucose and in all glucosamine concentrations tested. SDS-PAGE co-polymerized with glycol-chitin analysis showed three major activity bands of 200, 100, and 95 kDa, when incubated at 50°C.     

Eicosanoid-mediated proinflammatory activity of Pseudomonas aeruginosa ExoU

As Pseudomonas aeruginosa ExoU possesses two functional blocks of homology to calcium-independent (iPLA(2)) and cytosolic phospholipase A(2) (cPLA(2)), we addressed the question whether it would exhibit a proinflammatory activity by enhancing the synthesis of eicosanoids by host organisms. Endothelial cells from the HMEC-1 line infected with the ExoU-producing PA103 strain exhibited a potent release of arachidonic acid (AA) that could be significantly inhibited by methyl arachidonyl fluorophosphonate (MAFP), a specific PLA(2) inhibitor, as well as significant amounts of the cyclooxygenase (COX)-derived prostaglandins PGE(2) and PGI(2). Cells infected with an isogenic mutant defective in ExoU synthesis did not differ from non-infected cells in the AA release and produced prostanoids in significantly lower concentrations. Infection by PA103 induced a marked inflammatory response in two different in vivo experimental models. Inoculation of the parental bacteria into mice footpads led to an early increase in the infected limb volume that could be significantly reduced by inhibitors of both COX and lipoxygenase (ibuprofen and NDGA respectively). In an experimental respiratory infection model, bronchoalveolar lavage (BAL) from mice instilled with 10(4) cfu of PA103 exhibited a marked influx of inflammatory cells and PGE(2) release that could be significantly reduced by indomethacin, a non-selective COX inhibitor. Our results suggest that ExoU may contribute to P. aeruginosa pathogenesis by inducing an eicosanoid-mediated inflammatory response of host organisms.     

Biomphalaria tenagophila: dynamics of populations of resistant and susceptible strains to Schistosoma mansoni, with or without pressure of the parasite

Resistant (Taim, RS) and susceptible albino (Joinville, SC) Biomphalaria tenagophila populations were kept together, at different proportions, throughout a 18-month-period. Some of the snail groups were submitted to Schistosoma mansoni infection. The targets of this study were (a) to analyze the populational dynamics among resistant and susceptible individuals to S. mansoni; (b) to study the resistance phenotype in descendants of cross-breeding; (c) to observe whether the parasite could exert any kind of selection in those snail populations. Throughout the experiment it could be observed that the susceptible B. tenagophila strain (Joinville) underwent a selective pressure of the parasite that was negative, since the individuals showed a high mortality rate. Although B. tenagophila (Taim) population presented a higher mortality rate without pressure of the parasite, this event was compensated by a reproductive capacity. B. tenagophila Taim was more fecund than B. tenagophila Joinville and was able to transmit the resistance character to their descendants. F1 generation obtained by cross-breeding between resistant and susceptible lineages was completely resistant to S. mansoni infection, irrespective of the Taim proportion. Moreover, less than 5% of F2 progeny were susceptible to S. mansoni infection.     

Resource availability and population density in primates: A socio-bioenergetic analysis of the energy budgets of Guatemalan howler and spider monkeys

This paper presents data on the food and energy utilization budgets of two sympatric species of nonhuman primates living in the quasi-rain forest of Tikal, Guatemala. Seven researchers working in teams obtained approximately 1200 hours of animal-observer contact time, representing approximately 2400 hours in the field. The following information is herein presented: (1) data on the estimated energy expenditure of the cebidae population living in the study site; (2) the nutritional composition of some foods eaten by Tikal's howler and spider population; (3) an estimate of the nutritional intake of howlers and spiders; and (4) an estimate of the carrying capacity of the habitat. The above types of data were combined in an attempt to answer the following question: To what extent are population density and size an indication of the carrying capacity of the habitat either in terms of food availability or quality? Data presented in this paper suggest that low population density and small group size are not necessarily the result of a nutritionally poor environment. The Tikal study site is capable of supporting 8,500 animal/km². In the absence of other comparable quantitative socio-bioenergetic surveys it is herein suggested that the concept of food resource limitation is largely a myth among primatologists.     

Characterization of PbPga1, an Antigenic GPI-Protein in the Pathogenic Fungus Paracoccidioides brasiliensis

Paracoccidioides brasiliensis is the etiologic agent of paracoccidioidomycosis (PCM), one of the most prevalent mycosis in Latin America. P. brasiliensis cell wall components interact with host cells and influence the pathogenesis of PCM. Cell wall components, such as glycosylphosphatidylinositol (GPI)-proteins play a critical role in cell adhesion and host tissue invasion. Although the importance of GPI-proteins in the pathogenesis of other medically important fungi is recognized, little is known about their function in P. brasiliensis cells and PCM pathogenesis. We cloned the PbPga1 gene that codifies for a predicted GPI-anchored glycoprotein from the dimorphic pathogenic fungus P. brasiliensis. PbPga1 is conserved in Eurotiomycetes fungi and encodes for a protein with potential glycosylation sites in a serine/threonine-rich region, a signal peptide and a putative glycosylphosphatidylinositol attachment signal sequence. Specific chicken anti-rPbPga1 antibody localized PbPga1 on the yeast cell surface at the septum between the mother cell and the bud with stronger staining of the bud. The exposure of murine peritoneal macrophages to rPbPga1 induces TNF-α release and nitric oxide (NO) production by macrophages. Furthermore, the presence of O-glycosylation sites was demonstrated by β-elimination under ammonium hydroxide treatment of rPbPga1. Finally, sera from PCM patients recognized rPbPga1 by Western blotting indicating the presence of specific antibodies against rPbPga1. In conclusion, our findings suggest that the PbPga1gene codifies for a cell surface glycoprotein, probably attached to a GPI-anchor, which may play a role in P. brasiliensis cell wall morphogenesis and infection. The induction of inflammatory mediators released by rPbPga1 and the reactivity of PCM patient sera toward rPbPga1 imply that the protein favors the innate mechanisms of defense and induces humoral immunity during P. brasiliensis infection.