The molecular basis of E. coli colonization of the intestinal tract will be investigated using a mouse model. The proposed research focuses on the nature of the molecular interactions which occur at the mucosal surface and the role of such interactions in colonization. The proposal consists of three parts. In part one, the role of bacterial motility, chemotaxis, and adhesion, in colonization of the large intestine will be investigated using mutants of normal human fecal E. coli and avirulent S. typhimurium. Two general categories of mutants will be employed. First, mutants of E. coli and S. typhimurium known to possess an impaired ability to colonize the large intestines of streptomycin-treated mice, relative to the parent strain, will be assayed for chemotaxis toward mucus and the ability to penetrate mucus in vitro. Second, mutants of E. coli and s. typhimurium will be selected for specific defects in motility, chemotaxis, adhesion to mucosal components, and ability to penetrate mucus in vitro, and the relative colonizing abilities of the mutants will be assessed in vivo. In part two, the receptors and bacterial adhesins responsible for in vitro adhesion to mucus and epithelial cell brush borders will be identified, purified and characterized. These studies include further characterization of the outer membrane adhesins of E. coli F-18, E. coli F-17, and S. typhimurium, and the respective cecal and colonic mucus and brush border receptors for these strains. In addition, the adhesive properties of a wide variety of nonpathogenic E. coli will be assessed for their ability to adhere to mucus and brush border receptors immobilized on polystyrene, and the receptors involve in adhesion will be separated and identified by means of gel filtration, SDS-PAGE, and Western blot analysis. In part three, the mucus and brush border receptors responsible for the adhesion of selected enterotoxigenic E. coli to the mucosal surface of the small intestine will be identified, characterized and compared. Additional experiments designed to examine the relationship between in vitro adhesion to specific receptors and in vivo colonization will be conducted. The overall objectives of the proposed research are to further characterize the interactions which occur at the mucosal surface, further investigate and characterize the components involved in these interactions, and clarify the role of such interactions in the bacterial colonization of the intestinal tract.
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