Project description:Bacterial persister cells are phenotypic variants of regular cells that are tolerant to antibiotics. Analysis of clinical isolates of M. tuberculosis showed that strains vary substantially in their tolerance to antibiotics. The level of persisters was very high is some isolates, suggesting that these are hip mutants. We investigated gene expression differences in eight clinical isolates, four of which we characterized as high-persister strains and four as low-persister, or regular, strains. Comparison of gene expression patterns may provide clues as to the genetic mechanisms underlying persister formation.
Project description:While several groups, including our own, have examined variability of M. tuberculosis at the DNA level, this is the first systematic survey of variability in mRNA expression among clinical isolates of M. tuberculosis. Genes whose expression varies among isolates when assayed under a single growth condition may make poor drug targets and vaccine antigens and may affect molecular diagnostics, so they can be used to narrow down lists of candidate molecules. Because the measurement of gene expression is extremely sensitive to environmental conditions, comparison of gene expression is labour intensive. In this study, we surveyed 12 strains. These strains are a subset of those for which we have already published genomic deletion information (Kato-Maeda et al., 2001). In order to ensure maximum reproducibility of the experiments and avoid complications caused by differences in growth conditions, we measured gene expression under well-controlled in vitro conditions. Our aims were to provide an overview of gene expression variability among clinical isolates under a single growth condition and to test whether gene functional classes are related to variability in expression. Set of arrays that are part of repeated experiments Keywords: Biological Replicate
Project description:The accessory gene regulator (agr) of Staphylococcus aureus is a global regulator of the staphylococcal virulon, which includes secreted virulence factors and surface proteins. The agr locus is important for virulence in a variety of animal models of infection, and has been assumed by inference to have a major role in human infection. Although most human clinical S. aureus isolates are agr(+), there have been several reports of agr-defective mutants isolated from infected patients. Since it is well known that the agr locus is genetically labile in vitro, we have addressed the question of whether the reported agr-defective mutants were involved in the infection or could have arisen during post-isolation handling. We obtained a series of new staphylococcal isolates from local clinical infections and handled these with special care to avoid post-isolation mutations. Among these isolates, we found a number of strains with non-haemolytic phenotypes owing to mutations in the agr locus, and others with mutations elsewhere. We have also obtained isolates in which the population was continuously heterogeneous with respect to agr functionality, with agr(+) and agr(-) variants having otherwise indistinguishable chromosomal backgrounds. This finding suggested that the agr(-) variants arose by mutation during the course of the infection. Our results indicate that while most clinical isolates are haemolytic and agr(+), non-haemolytic and agr(-) strains are found in S. aureus infections, and that agr(+) and agr(-) variants may have a cooperative interaction in certain types of infections.
Project description:Monocytes play a critical role during infection with Mycobacterium tuberculosis (Mtb). They are recruited to the lung where they participate in the contention of infection. Alternatively, inflammatory monocytes may help in prolonging inflammation or serve as niches for Mtb infection. Also, monocyte response to infection may vary depending on the particularities of the clinical isolate of Mtb from which they are infected. In this pilot study, using microarrays we have examined the global mRNA profiles of circulating human monocytes from healthy individuals and patients with active tuberculosis (TB).
Project description:This study uses microarray analyses to examine transcriptional responses of Mycobacterium tuberculosis complex clinical isolates to phagosomal cues encountered inside resting murine bone marrow-derived macrophages 24hr post-infection. Transcript levels of intracellular mycobacteria were compared to extracellular bacteria of the same strain (An aliquot of the inoculum used to infect macrophages was incubated in the absence of macrophages for 24hr in an identical flask). Set of arrays that are part of repeated experiments Keywords: Biological Replicate
Project description:This study uses microarray analyses to examine transcriptional responses of Mycobacterium tuberculosis complex clinical isolates to phagosomal cues encountered inside activated (IFN-gamma+LPS) murine bone marrow-derived macrophages 24hr post-infection. Transcript levels of intracellular mycobacteria were compared to extracellular bacteria of the same strain (An aliquot of the inoculum used to infect macrophages was incubated in the absence of macrophages for 24hr in an identical flask). Set of arrays that are part of repeated experiments Keywords: Biological Replicate
Project description:While several groups, including our own, have examined variability of M. tuberculosis at the DNA level, this is the first systematic survey of variability in mRNA expression among clinical isolates of M. tuberculosis. Genes whose expression varies among isolates when assayed under a single growth condition may make poor drug targets and vaccine antigens and may affect molecular diagnostics, so they can be used to narrow down lists of candidate molecules. Because the measurement of gene expression is extremely sensitive to environmental conditions, comparison of gene expression is labour intensive. In this study, we surveyed 12 strains. These strains are a subset of those for which we have already published genomic deletion information (Kato-Maeda et al., 2001). In order to ensure maximum reproducibility of the experiments and avoid complications caused by differences in growth conditions, we measured gene expression under well-controlled in vitro conditions. Our aims were to provide an overview of gene expression variability among clinical isolates under a single growth condition and to test whether gene functional classes are related to variability in expression. Set of arrays that are part of repeated experiments Keywords: Biological Replicate Biological Replicate Computed
Project description:We have identified erm(Y), a novel gene class that was originally designated ermGM, from a Staphylococcus aureus strain that has a plasmid that also harbors msr(A) and mph(C), genes that encode an efflux mechanism and a putative phosphorylase, respectively. The nucleotide and deduced amino acid sequences of erm(Y) were 81 and 76% identical to those of erm(T), respectively.