The human colon contains a dense and complex population of bacteria that persists in an individual throughout life. A major carbon source for these bacteria is polysaccharide in the host's diet, but polysaccharide produced by the host itself may also be utilized by colonic bacteria. Bacteroides, a genus of gram negative obligate anaerobes, is one of the numerically predominant genera of colonic bacteria and appear to be major contributors to the colonic fermentation. Bacteroides species have a novel system for utilization of polysaccharide, a polysaccharide import system. They first bind the polysaccharide to their outer membrane, then translocate it into the periplasm where breakdown begins. This strategy not only helps the bacteria to particulate matter in the host's diet. A major long term goal of the proposed work is to used genetic and biochemical techniques to determine the characteristics and function of proteins involved in the binding and internalization of polysaccharides by Bacteroides species. The starch utilization system of Bacteroides thetaiotaomicron will be used as a model system. Some genes in this system have already been cloned and the function of proteins encoded by these genes will be determined. Other essential genes will be cloned and characterized. The hypothesis to be tested is that outer membrane and periplasmic proteins identified in previous studies form a polysaccharide binding/translocation complex that extends through the outer membrane and into the periplasm. A second goal of the proposed research is to understand what contribution host-derived polysaccharide make to the nutrition of colonic bacteria. Further characterization of mutants unable to utilize host polysaccharide will be done, with particular focus on a mutation that decreases the ability of B. thetaiotaomicron to compete for colonization of the intestinal tracts of germfree mice. The activities of the major groups of colonic mucosa and probably have major long term effects on the mucosa. Although there is no satisfactory assay systems for assessing hr effect of bacterial products on colonic cells. A final goal of this proposal is to begin to develop in vitro assay systems for determining the effects of various bacterial products on colonic cells. Such assay systems are needed not only for studies of the possible contribution of bacterial activities to diseases such as colon cancer, but also for assessing the effects of new food additives such as artificial fats and bulking agents that are known to be fermented extensively by colonic bacteria.
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