Studies aimed at inducing immunity against infectious diseases, including Dental caries, have provided valuable information on microbial antigens important in inducing protective responses, the role of the mucosal immune system and IgA antibodies in defense against infections involving surfaces bathed by external secretions, and mechanisms involved in the induction of immune responses. The overall goal of this project is to define mechanisms by which mucosal vaccines consisting of recombinant, avirulent Salmonella strains expressing cloned genes of mutans streptococci, with and without adjuvant induce specific immune responses to the cloned antigen, which provide long-term protection. Specifically these studies will: 1) Determine the effect of persistence of the Salmonella vaccine strain and the amount of the expressed cloned antigens of mutans streptococci on the induction, nature and memory of immune response. Levels and isotype of antibodies to cloned antigens in serum and external secretions of animals immunized by the oral or intranasal (IN) route with Salmonella vaccines which persist for short or long times in the host, and which produce various amounts of cloned antigen will be measured by ELISA to determine the effect of these characteristics on the induction of mucosal immune responses. Protection will be assessed in an experimental model. The effect of Salmonella on the immune response to the cloned proteins will be characterized by measuring antigen-specific proliferation, cytokine production by ELISA and ELISPOT assay, and expression of co-stimulatory molecules by FACS in cell preparations from systemic and mucosal tissues. 2) Determine the effect of mucosal adjuvants on modulating host responses to recombinant antigens of mutans streptococci. Levels and isotype of antibodies to cloned antigens of mutans streptococci in serum and secretions of animals immunized by the oral or IN route with chimeric protein consisting of cloned antigens genetically linked to the B subunit of cholera toxin (CTB) or Salmonella vector vaccine expressing various amounts of chimeric proteins +/- free CTB will be measured by ELISA. The effect of the Salmonella on the adjuvant properties of CTB will be assessed by evaluating cells from systemic and mucosal tissues for the expression of co-stimulatory molecules and the profile of cytokines induced. 3) Determine if chimeric proteins consisting of cloned antigens of mutans streptococci are more effective than each antigen alone in inducing protective immune responses. Levels and isotype of antibodies to the cloned antigens in saliva and serum will be measured in animals immunized by the oral or IN route to determine if chimeric proteins of mutans streptococci antigens induce higher salivary IgA antibody responses and greater protection against infection by mutans streptococci than each cloned protein alone. The results will be relevant to establish the practicability of Salmonella vaccine delivery systems and the usefulness of genetically derived chimeric proteins from virulence factors of a pathogen and adjuvants for the induction of protective immune responses against mucosal pathogens including those associated with the oral cavity.
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