The long-term goal of this study is to understand the role in pathogenesis and immunity of an environmentally-regulated, in vivo expressed surface polysaccharide of Staphylococcus aureus chemically characterized as poly-N-succinyl-beta-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 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 5 different of animal models of S. aureus infection. The models encompass nasal colonization, acute lethality, kidney abscess formation, endocarditis and localized skin abscesses. 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 in experimental animals. Rabbits will be immunized with the variants and sera assessed for antibody titer and opsonic killing ability. The PNSG variant structures will also be used to immunize mice to evaluate the ability of these constructs to generate protective immunity in the same models used for the study of the role of PNSG in S. aureus virulence. In addition, passive protective capacity of the rabbit sera raised to the variant PNSG constructs will be evaluated in the 5 animal models. All the above mentioned studies will provide new and useful information regarding pathogenesis and immunity of staphylococcal infections, because it has not previously been appreciated that S. aureus expresses the PNSG antigen as a surface, capsule-like polysaccharide. 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 models, 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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