988 resultados para Sao Tome and Principe


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Aims: To evaluate the sensitivity and specificity of polyclonal and monoclonal antibodies (Mabs) against intimin in the detection of enteropathogenic and enterohaemorrhagic Escherichia coli isolates using immunoblotting. Methods and Results: Polyclonal and Mabs against the intimin-conserved region were raised, and their reactivities were compared in enteropathogenic E. coli (EPEC) and enterohaemorrhagic E. coli (EHEC) isolates using immunoblotting analysis. In comparison with rat antiserum, rabbit anti-intimin IgG-enriched fraction had a stronger recognition pattern to a wide spectrum of intimin types in different EPEC and EHEC serotypes. On the other hand, murine monoclonal IgG2b specific to intimin, with dissociation constant of 1 center dot 3 x 10-8 mol l-1, failed in the detection of some of these isolates. Conclusion: All employed antibodies showed 100% specificity, not reacting with any of the eae-negative isolates. The sensitivity range was according to the employed antisera, and 97% for rabbit anti-intimin IgG-enriched fraction, followed by 92% and 78% sensitivity with rat antisera and Mab. Significance and Impact of the Study: The rabbit anti-intimin IgG-enriched fraction in immunoblotting analysis is a useful tool for EPEC and EHEC diagnoses.

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Parvimonas micra are gram positive anaerobic cocci isolated from the oral cavity and frequently related to polymicrobial infections in humans. Despite reports about phenotypic differences, the genotypic variation of P. micra and its role in virulence are still not elucidated. The aim of this study was to determine the genotypic diversity of P. micra isolates obtained from the subgingival biofilm of subjects with different periodontal conditions and to correlate these findings with phenotypic traits. Three reference strains and 35 isolates of P. micro were genotyped by 16S rRNA PCR-RFLP and phenotypic traits such as collagenase production, elastolytic and hemolytic activities were evaluated. 16S rRNA PCR-RFLP showed that P. micra could be grouped into two main clusters: C1 and C2; cluster C1 harbored three genotypes (HG1259-like, HG1467-like and ICBM0583-like) while cluster C2 harbored two genotypes (ATC03270-like and ICBM036). A wide variability in collagenolytic activity intensities was observed among all isolates, while elastolytic activity was detected in only two isolates. There was an association between hemolytic activity in rabbit erythrocytes and cluster C2. There was an association between hemolytic activity in rabbit erythrocytes and cluster C1. Although these data suggest a possible association between P. micra genetic diversity and their pathogenic potential, further investigations are needed to confirm this hypothesis. (C) 2009 Elsevier Ltd. All rights reserved.

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Glutathione S-transferase (GST) is a family of enzymes involved in the detoxification of electrophilic compounds. Different classes of GST are expressed in various organs, such as liver, lungs, stomach and others. Expression of GST can be modulated by diet components and plant-derived compounds. The importance of controlling GST expression is twofold: increasing levels of GST are beneficial to prevent deleterious effects of toxic and carcinogenic compounds, while inhibition of GST in tumor cells may help overcoming tumor resistance to chemotherapy. A screening of 16 plants used in the Brazilian pharmacopoeia tested their effects on GST expression in hepatocytes and Jurkat (leukemia) T-cells. The methanol extracts of five plants inhibited GST expression in hepatocytes. Three plants significantly inhibited and four others induced GST expression in Jurkat cells. Among these, the extracts of Bauhinia forficata Link. (Leguminosae) and Cecropia pachystachya Trec. (Urticaceae) inhibited GST expression at relatively low concentrations. With the exception of B. forficata, all plants were cytotoxic when administered to Jurkat cells at high doses (1 mg/mL) and some extracts were considerably cytotoxic even at lower concentrations.

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The objective of the present study was to evaluate the effects of different gamma radiation doses on the growth of Alternaria alternata and on the production of toxins alternariol (AOH), and alternariol monomethyl ether (AME) in sunflower seed samples. After irradiation with 2, 5 and 7 kGy, the spore mass was resuspended in sterile distilled water and the suspension was inoculated into sunflower seeds. The number of colony-forming units per gram (CFU/g) was determined after culture on Dichloran Rose Bengal Chloramphenicol and Dichloran Chloramphenicol Malt Extract Agar. The presence of AOH and AME was investigated by liquid chromatography coupled to mass spectrometry. The radiation doses used resulted in a reduction of the number of A. alternata CFU/g and of AOH and AME levels when compared to the nonirradiated control group. Maximum reduction of the fungus (98.5%) and toxins (99.9%) was observed at a dose of 7 and 5 kGy, respectively. Under the present conditions, gamma radiation was found to be an alternative for the control of A. alternata and, consequently, of AOH and AME production in sunflower seeds. (C) 2009 Elsevier Ltd. All rights reserved.

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Background and Objective: Inflammatory cytokines such as tumor necrosis factor-alpha are involved in the pathogenesis of periodontal diseases. A high between-subject variation in the level of tumor necrosis factor-alpha mRNA has been verified, which may be a result of genetic polymorphisms and/or the presence of periodontopathogens such as Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola (called the red complex) and Aggregatibacter actinomycetemcomitans. In this study, we investigated the effect of the tumor necrosis factor-alpha (TNFA) -308G/A gene polymorphism and of periodontopathogens on the tumor necrosis factor-alpha levels in the periodontal tissues of nonsmoking patients with chronic periodontitis (n = 127) and in control subjects (n = 177). Material and Methods: The TNFA-308G/A single nucleotide polymorphism was investigated using polymerase chain reaction-restriction fragment length polymorphism analysis, whereas the tumor necrosis factor-alpha levels and the periodontopathogen load were determined using real-time polymerase chain reaction. Results: No statistically significant differences were found in the frequency of the TNFA-308 single nucleotide polymorphism in control and chronic periodontitis groups, in spite of the higher frequency of the A allele in the chronic periodontitis group. The concomitant analyses of genotypes and periodontopathogens demonstrated that TNFA-308 GA/AA genotypes and the red-complex periodontopathogens were independently associated with increased levels of tumor necrosis factor-alpha in periodontal tissues, and no additive effect was seen when both factors were present. P. gingivalis, T. forsythia and T. denticola counts were positively correlated with the level of tumor necrosis factor-alpha. TNFA-308 genotypes were not associated with the periodontopathogen detection odds or with the bacterial load. Conclusion: Our results demonstrate that the TNFA-308 A allele and red-complex periodontopathogens are independently associated with increased levels of tumor necrosis factor-alpha in diseased tissues of nonsmoking chronic periodontitis patients and consequently are potentially involved in determining the disease outcome.

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Libraries are very propitious environments for the growth of fungi. The great concentration of organic material available for these microorganisms, and often with the lack of adequate ventilation or climate control, would favour this situation. This study was conducted in 2003 to determine the predominant genera of fungi in public libraries by a survey of fungi contaminating the upper surface of books, with and without air conditioning in the city of Sao Paulo, Brazil, in the winter and summer, during the respective periods with high and low levels of airborne fungi in that city. Six libraries were chosen, located on the campus of the University of Sao Paulo, three of them with air conditioning and the other three with natural ventilation. In these six libraries, 31 genera of fungi were identified in total. The genera and frequency of contaminant fungi recovered differed significantly between the libraries with and without air conditioning and in the samples collected in the summer as opposed to the winter. Cladosporium was the most frequent in the libraries with and without air conditioning, and in the winter. Aspergillus was isolated more often in the summer.

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A different organization for the xyl operon was found in different genomes of Burkholderia and Pseudomomas species. Degenerated primers were designed based on Burkholderia genomes and used to amplify the xylose isomerase gene (xylA) from Burkholderia sacchari IPT101 The gene encoded a protein of 329 amino acids, which showed the highest similarity (90%) to the homologous gene of Burkholderia dolosa. It was cloned in the broad host range plasmid pBBR1MCS-2, which partially restored growth and polyhydroxybutyrate production capability in xylose to a B. sacchari xyl(-) mutant. When xylA was overexpressed in the wild-type strain, it was not able to increase growth and polyhydroxybutyrate production, suggesting that XylA activity is not limiting for xylose utilization in B. sacchari.

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Aims: To determine the prevalence and expression of metallo-beta-lactamases (MBL)-encoding genes in Aeromonas species recovered from natural water reservoirs in southeastern Brazil. Methods and Results: Eighty-seven Aeromonas isolates belonging to Aeromonas hydrophila (n = 41) and Aer. jandaei (n = 46) species were tested for MBL production by the combined disk test using imipenem and meropenem disks as substrates and EDTA or thioglycolic acid as inhibitors. The presence of MBL genes was investigated by PCR and sequencing using new consensus primer pairs designed in this study. The cphA gene was found in 97.6% and 100% of Aer. hydrophila and Aer. jandaei isolates, respectively, whereas the acquired MBL genes bla(IMP), bla(VIM) and bla(SPM-1) were not detected. On the other hand, production of MBL activity was detectable in 87.8% and 10.9% of the cphA-positive Aer. hydrophila and Aer. jandaei isolates respectively. Conclusions: Our results indicate that cphA seems to be intrinsic in the environmental isolates of Aer. hydrophila and Aer. jandaei in southeastern Brazil, although, based on the combined disk test, not all of them are apparently able to express the enzymatic activity. Significance and Impact of the Study: These data confirm the presence of MBL-producing Aeromonas species in natural water reservoirs. Risk of water-borne diseases owing to domestic and industrial uses of freshwater should be re-examined from the increase of bacterial resistance point of view.

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The type I and type II heat-labile enterotoxins (LT-I and LT-II) are strong mucosal adjuvants when they are coadministered with soluble antigens. Nonetheless, data on the parenteral adjuvant activities of LT-II are still limited. Particularly, no previous study has evaluated the adjuvant effects and induced inflammatory reactions of LT-II holotoxins or their B pentameric subunits after delivery via the intradermal (i.d.) route to mice. In the present report, the adjuvant and local skin inflammatory effects of LT-IIa and its B subunit pentamer (LT-IIaB(5)) were determined. When coadministered with ovalbumin (OVA), LT-IIa and, to a lesser extent, LT-IIaB(5) exhibited serum IgG adjuvant effects. In addition, LT-IIa but not LT-IIaB(5) induced T cell-specific anti-OVA responses, particularly in respect to induction of antigen-specific cytotoxic CD8(+) T cell responses. LT-IIa and LT-IIaB(5) induced differential tissue permeability and local inflammatory reactions after i.d. injection. Of particular interest was the reduced or complete lack of local reactions, such as edema and tissue induration, in mice i.d. inoculated with LT-IIa and LT-IIaB(5), respectively, compared with mice immunized with LT-I. In conclusion, the present results show that LT-IIa and, to a lesser extent, LT-IIaB(5) exert adjuvant effects when they are delivered via the i.d. route. In addition, the low inflammatory effects of LT-IIa and LT-IIaB(5) in comparison to those of LT-I support the usefulness of LT-IIa and LT-IIaB(5) as parenterally delivered vaccine adjuvants.

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Epidemiological and molecular characteristics of human metapneumovirus (hMPV) were compared with human respiratory syncytial virus (hRSV) in infants and young children admitted for acute lower respiratory tract infections in a prospective study during four consecutive years in subtropical Brazil. GeneScan polymerase chain assays (GeneScan RT-PCR) were used to detect hMPV and hRSV in nasopharyngeal aspirates of 1,670 children during January 2003 to December 2006. hMPV and hRSV were detected, respectively, in 191 (11.4%) and in 702 (42%) of the children admitted with acute lower respiratory tract infections at the Sao Paulo University Hospital. Sequencing data of the hMPV F gene revealed that two groups of the virus, each divided into two subgroups, co-circulated during three consecutive years. It was also shown that a clear dominance of genotype B1 occurred during the years 2004 and 2005, followed by genotype A2 during 2006. J. Med. Virol. 81:915-921,2009. (C) 2009 Wiley-Liss, Inc.

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The biosynthesis of quinolinate, the de novo precursor of nicotinamide adenine dinucleotide (NAD), may be performed by two distinct pathways, namely, the bacterial aspartate (aspartate-to-quinolinate) and the eukaryotic kynurenine (tryptophan-to-quinolinate). Even though the separation into eukaryotic and bacterial routes is long established, recent genomic surveys have challenged this view, because certain bacterial species also carry the genes for the kynurenine pathway. In this work, both quinolinate biosynthetic pathways were investigated in the Bacteria clade and with special attention to Xanthomonadales and Bacteroidetes, from an evolutionary viewpoint. Genomic screening has revealed that a small number of bacterial species possess some of the genes for the kynurenine pathway, which is complete in the genus Xanthomonas and in the order Flavobacteriales, where the aspartate pathway is absent. The opposite pattern (presence of the aspartate pathway and absence of the kynurenine pathway) in close relatives (Xylella ssp. and the order Bacteroidales, respectively) points to the idea of a recent acquisition of the kynurenine pathway through lateral gene transfer in these bacterial groups. In fact, sequence similarity comparison and phylogenetic reconstruction both suggest that at least part of the genes of the kynurenine pathway in Xanthomonas and Flavobacteriales is shared by eukaryotes. These results reinforce the idea of the role that lateral gene transfer plays in the configuration of bacterial genomes, thereby providing alternative metabolic pathways, even with the replacement of primary and essential cell functions, as exemplified by NAD biosynthesis.

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Heat-labile toxins (LT) encompass at least 16 natural polymorphic toxin variants expressed by wild-type enterotoxigenic Escherichia coli (ETEC) strains isolated from human beings, but only one specific form, produced by the reference ETEC H10407 strain (LT1), has been intensively studied either as a virulence-associated factor or as a mucosal/transcutaneous adjuvant. In the present study, we carried out a biological/immunological characterization of a natural LT variant (LT2) with four polymorphic sites at the A subunit (S190L, G196D, K213E, and S224T) and one at the B subunit (T75A). The results indicated that purified LT2, in comparison with LT1, displayed similar in vitro toxic activities (adenosine 3`,5`-cyclic monophosphate accumulation) on mammalian cells and in vivo immunogenicity following delivery via the oral route. Nonetheless, the LT2 variant showed increased adjuvant action to ovalbumin when delivered to mice via the transcutaneous route while antibodies raised in mice immunized with LT2 displayed enhanced affinity and neutralization activity to LT1 and LT2. Taken together, the results indicate that the two most frequent LT polymorphic forms expressed by wild ETEC strains share similar biological features, but differ with regard to their immunological properties.

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The correlation between the microdilution (MD), Etest (R) (ET), and disk diffusion (DD) methods was determined for amphotericin B, itraconazole and fluconazole. The minimal inhibitory concentration (MIC) of those antifungal agents was established for a total of 70 Candida spp. isolates from colonization and infection. The species distribution was: Candida albicans (n = 27), C. tropicalis (n = 17), C. glabrata (n = 16), C. parapsilosis (n = 8), and C. lusitaniae (n = 2). Non-Candida albicans Candida species showed higher MICs for the three antifungal agents when compared with C. albicans isolates. The overall concordance (based on the MIC value obtained within two dilutions) between the ET and the MD method was 83% for amphotericin B, 63% for itraconazole, and 64% for fluconazole. Considering the breakpoint, the agreement between the DD and MD methods was 71% for itraconazole and 67% for fluconazole. The DD zone diameters are highly reproducible and correlate well with the MD method, making agar-based methods a viable alternative to MD for susceptibility testing. However, data on agar-based tests for itraconazole and amphotericin B are yet scarce. Thus, further research must still be carded out to ensure the standardization to other antifungal agents. J. Clin. Lab. Anal. 23:324-330, 2009. (C) 2009 Wiley-Liss, Inc.

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Cultures of cosmomycin D-producing Streptomyces olindensis ICB20 that were propagated for many generations underwent mutations that resulted in production of a range of related anthracyclines by the bacteria. The anthracyclines that retained the two trisaccharide chains of the parent compound were separated by HPLC. Exact mass determination of these compounds revealed that they differed from cosmomycin D (CosD) in that they contained one to three fewer oxygen atoms (loss of hydroxyl groups). Some of the anthracyclines that were separated by HPLC had the same mass. The location from which the hydroxyl groups had been lost relative to CosD (on the aglycone and/or on the sugar residues) was probed by collisionally-activated dissociation using an electrospray ionisation linear quadrupole ion trap mass spectrometer. The presence of anthracyclines with the same mass, but different structure, was confirmed using an electrospray ionisation travelling wave ion mobility mass spectrometer.

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The META cluster of Leishmania amazonensis contains both META1 and META2 genes, which are upregulated in metacyclic promastigotes and encode proteins containing the META domain. Previous studies defined META2 as a 48.0-kDa protein, which is conserved in other Leishmania species and in Trypanosoma brucei. In this work, we demonstrate that META2 protein expression is regulated during the Leishmania life cycle but constitutive in T. brucei. META2 protein is present in the cytoplasm and flagellum of L amazonensis promastigotes. Leishmania META2-null replacement mutants are more sensitive to oxidative stress and, upon heat shock, assume rounded morphology with shortened flagella. The increased susceptibility of null parasites to heat shock is reversed by extra-chromosomal expression of the META2 gene. Defective Leishmania promastigotes exhibit decreased ability to survive in macrophages. By contrast, META2 expression is decreased by 80% in RNAi-induced T. brucei bloodstream forms with no measurable effect on survival or resistance to heat shock. (C) 2010 Elsevier Inc. All rights reserved.