The long-term goal of this study is to understand the role in pathogenesis and immunity of an environmentally regulated surface polysaccharide of Staphylococcus aureus chemically characterized as poly-N-succinyl-B-1-6 glucosamine (PNSG). PNSG has previously been determined to be the protective capsular polysaccharide/adhesin (PS/A) antigen of Staphylococcus epidermidis, raising the possibility that PNSG could be used as a """"""""pan-staphylococcal"""""""" vaccine. To define the role of PNSG in pathogenesis of S. aureus infection 5 different PNSG-deficient S. aureus strains representative of major lineages will be constructed by genetic means via interruption of the genes in the intracellular adhesin (ica) locus that encodes proteins needed for synthesis of PNSG. Isogenic parental, mutant and ica complemented strains will be evaluated in vitro to determine the role of PNSG in promoting S. aureus adherence to catheters and in providing resistance of bacterial cells to phagocytic killing by leukocytes and complement. The same strains will also be tested for infectious capability in several animal systems of S. aureus infection, including animals actively and passively immunized with ica-deleted S. aureus and normal human serum to reflect the immunologic status of humans, who have high levels of natural antibody to S. aureus surface antigens. Because PNSG isolated from some strains of staphylococci have up to 30 percent of the succinate substituents on the polyglucosamine backbone replaced by acetate, purified PNSG, with differing ratios of succinate and acetate substituents on the polyglucosamine backbone, will be produced for immunologic studies. Rabbits will be immunized with the variants and sera assessed for antibody titer and opsonic killing ability. The PNSG variant structures will be used to immunize mice to evaluate their ability to generate protective immunity in the same systems used for the study of the role of PNSG in S. aureus virulence. In addition, passive protection by the rabbit sera raised to the variant PNSG constructs will be evaluated in the animal systems. All the above mentioned studies will provide new and useful information regarding pathogenesis and immunity of staphylococcal infections, stressing the use of animal systems that reflect naturally acquired immunity in humans to S. aureus. By the end of these studies we expect to have a clear understanding of the role of PNSG in virulence, as determined in a variety of staphylococcal infection systems, the immunochemical properties of PNSG that can engender protective immunity, and the types of S. aureus infections wherein PNSG-specific immunotherapies show the most potential for success.
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