Listeria monocytogenes (L. m.) causes serious invasive diseases in humans and animals, with a human case mortality rate of approximately 20%. One goal of the US Dept. of Health and Human Services Healthy People 2010 Initiative is to reduce human listeriosis cases by 50%. The long-term objective of our research program is to contribute to that end through identification of factors that influence L. m. pathogenesis, which ultimately will enable development of novel and effective intervention strategies for preventing listerial infections. The work proposed in this application is designed to test the specific hypotheses that (i) the sigma/B general stress response system in gram-positive bacterial pathogens (and specifically in L. monocytogenes) provides a key transcriptional regulatory mechanism that facilitates environmental survival and virulence through induction of stress response genes; and that (ii) bacterial stress response systems contribute to pathogenesis by responding to specific environments, including those encountered in the host, through initiation of stress response and virulence gene expression (e.g., prfA).
The specific aims of these studies are to: (1) Define the L. m. sigmaB regulon through proteomic and genetic approaches. (2) Determine sigmaB regulon expression patterns under environmental stress conditions, sigma/B -dependent gene expression patterns will be evaluated using microarrays and reporter (-3) Measure sigmaB-dependent gene expression during host cell infection. Reporter fusions to selected sigmaB-dependent genes (e.g., prfA) in wildtype L. m. and selected null mutant strains (e.g., AsigB) will be used to identify gene expression patterns during cellular infection in tissue culture models. (4) Characterize deltasigmaB mutant virulence in tissue culture and animal models. At the conclusion of these studies, we will have developed an understanding of the contribution of cyB and the sigmaB-dependent stress response system to L. monocytogenes environmental survival and infection. More broadly, L. monocytogenes will serve as a model system for examining the role of alternative sigma factor-directed general stress response systems in survival and pathogenesis of gram-positive food-borne pathogens.
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