This research proposal seeks to determine the role of the Staphylococcus aureus capsular polysaccharide in pathogenesis. Specifically, the importance of the bacterial capsul in S. aureus-induced abscesses will be investigated, as well as the nature of the protective host immune response elicited against abscess formation. From three clinical strains of S. aureus, isogenic encapsulated variants have been recovered, which differ from the parental strains in their ability to synthesize large quantities of capsular polysaccharide. The capsular polysaccharide will be purified from cultures of the highly encapsulated (mucoid) variants, and its components will be biochemically characterized. To determine if they synthesize any capsular material, cultures of the parental (non-mucoid) strains will also be extracted biochemically by methods used to recover capsular material from the mucoid strains. Transmission electron microscopy will be used to visualize the capsule on both parental and variant strains. The most common manifestation of staphylococcal disease in humans is the localized abscess, from which S. aureus can multiply and disseminate. Therefore, a mouse model of infection has been developed to assess the role of the staphylococcal capsular polysaccharide in abscess formation. Encapsulated and unencapsulated isogenic strains of S. aureus will be injected IP into mice, and gross abscess formation within the abdominal cavity will be assessed visually and by culture. Dissemination and organ involvement, particularly in the kidney, will be closely monitored by quantitative cultures. Mice will be immunized with either encapsulated or unencapsulated S. aureus strains. Both groups will be challenged with encapsulated S. aureus strains, and protection will be defined as a significant increase in the AD50 dose in immunized mice compared with unimmunized controls. Purified capsular polysaccharide will be used to immunize additional groups of mice which will be challenged in a similar fashion. To determine the host defense mechanisms effective in immunity, the protective role of immune serum or immune spleen cells will be evaluated in passive immunization experiments. These studies will provide a solid base of knowledge concerning the importance of the capsule in staphylococcal virulence and provide insight into the future accine potential of the purified capsular polysaccharide.
