At birth pioneer bacterial colonize the mouth and are succeeded by other populations to form the normal flora which plays an important role in host defense by excluding exogenous pathogens. However, dental caries and periodontal disease, are caused by bacteria that are often isolated from the normal flora. Furthermore, some commensal bacteria are opportunistic pathogens that cause severe systemic disease when the immune system is compromised. Secretory immunoglobulin A (SigA) antibodies reactive with commensal bacteria are present in saliva but their impact ont he normal flora is not known. the objective of this competing continuation is to define the salivary immune response to commensal oral bacteria and studies will focus on the pioneer bacterium S. mitis biovar 1, and the late colonizer S. mutans, for the following reasons: (1) pioneer bacteria are the first to colonize the mucosal surfaces and to be exposed to the secretory immune system; (2) pioneer bacteria are exclusively streptococci of limited phenotype; (3) S. mitis biovar 1 is the principal pioneer Streptococcus; (4) the immune response to the """"""""late"""""""" colonizing streptococcus S. mutans, which is exposed to a more developed secretory immune system, can be compared to the early response to the pioneer S. mitis biovar 1. Saliva, breast milk, cord blood, serum and isolates of S. mitis biovar 1 and S. mutans obtained from the infant-mother pairs during an ongoing longitudinal study and its extension to the third year of the infants' life will be analyzed to test the following hypothesis: Commensal bacteria colonize and persist in the mouth because they subvert SigA antibodies and/or because secretory antibodies are of low avidity, low level or the wrong specificities.
The Specific Aims are as follows:
Aim 1 : Determine the incidence of pioneer viridans streptococci producing IgA1 protease. Isolates of pioneer streptococci will be incubated with IgA1 and IgA1 protease activity will be detected by SDS-PAGE to test the hypothesis that IgA1 protease is an ecological determinant in colonization by destroying the biological activity of SigA1 antibodies.
Aim 2 : Analyze the clonal diversity of S. mitis biovar 1 and S. mutans. Isolates of these streptococci obtained from the infant-mother pairs during the ongoing longitudinal study will be examined by DNA fingerprinting and ribotyping to test the hypotheses that: (a) there is limited genetic heterogeneity within the strains of these species in each infants mouth; (b) different infants harbor different genotypes of these species; (c) clonotypes of these species are stable in the mouth over time; (d) an infant acquires these bacteria from the mother.
Aim 3 : Analyze the diversity of secretory and systemic antibodies reactive with S. mitis biovar 1 and S. mutans. Saliva, breast milk, cord blood and serum obtained from the infant-mother pairs will be analyzed by Western blotting and by ELISA to test the hypotheses that: (a) antibodies reactive with S. mitis biovar 1 and S. mutans are composed of (1) cross-reactive antibodies induced by related oral and/or gut bacteria and (2) specific antibodies induced by colonization of the homologous strain; (b) antibody specificities differ in the secretory and systemic compartments of the humoral immune system.
Aim 4 : Analyze the avidity of SigA, SigA1, and SigA2 antibodies reactive with S. mitis biovar 1 and S. mutans. Chaotrope dissociation ELISA and immunoblots will be used to test the hypothesis that commensal bacteria induce low avidity SigA antibodies that do not exhibit affinity maturation and are ineffective in immune elimination of these bacteria.
