We previously demonstrated that intact, inactivated Streptococcus pneumoniae (unencapsulated strain R36A) inhibits IgG responses to a number of coimmunized soluble antigens (Ags). In this study, we investigated the mechanism of this inhibition and whether other extracellular bacteria exhibited similar effects. No inhibition was observed if R36A was given 24 h before or after immunization with soluble chicken ovalbumin (cOVA), indicating that R36A acts transiently during the initiation of the immune response. Using transgenic cOVA-specific CD4؉ T cells, we observed that R36A had no significant effect on T-cell activation (24 h) or generation of regulatory T cells (day 7) and only a modest effect on T-cell proliferation (48 to 96 h) in response to cOVA. However, R36A mediated a significant reduction in the formation of Ag-specific splenic germinal center T follicular helper (GC Tfh) and GC B cells and antibody-secreting cells in the spleen and bone marrow in response to cOVA or cOVA conjugated to 4-hydroxy-3-nitrophenylacetyl hapten (NP-cOVA). Of note, the inhibitory effect of intact R36A on the IgG anti-cOVA response could be reproduced using R36A-derived cell walls. In contrast to R36A, neither inactivated, unencapsulated, intact Neisseria meningitidis nor Streptococcus agalactiae inhibited the OVA-specific IgG response. These results suggest a novel immunosuppressive property within the cell wall of Streptococcus pneumoniae. During the natural course of bacterial infections, the immune system is exposed to both cell-associated and soluble microbial components (1-3). Distinct differences exist in the immunologic properties between particulate and soluble antigens (Ags) (4-8), suggesting the possibility of cross-regulatory processes occurring upon their simultaneous encounter by the immune system. In addition, the expression of innate stimulating moieties (9, 10), scavenger receptor ligands (11), and virulence factors by intact pathogens may further influence the immune response to coimmunizing soluble antigens. A number of studies in mice have demonstrated inhibitory effects of infectious agents on antibody responses, including germinal center (GC) reactions to soluble protein antigens (12-16). Although the underlying mechanisms of these inhibitory effects were not fully clarified, pathogen-mediated inhibition of dendritic cell (DC) maturation and induction of regulatory T cells (Tregs) were implicated, depending upon the specific pathogen. An understanding of the interplay between intact microbes and immune responsiveness to soluble antigens may have implications on processes involving natural immunity, autoimmunity, and vaccination.We previously demonstrated that intact, heat-killed Streptococcus pneumoniae inhibited the protein-and polysaccharide-specific IgG responses to a number of soluble conjugate vaccines, as well as soluble chicken ovalbumin (cOVA), upon coimmunization of mice intraperitoneally (i.p.) in the presence of alum plus CpG-containing oligodeoxynucleotides (CpG-ODN) as an adjuvant (17). In con...

Intact, inactivated Streptococcus pneumoniae (Pn) [including the unencapsulated strain, R36A], markedly inhibits the humoral immune response to co-immunized heterologous proteins, a property not observed with several other intact Gram-positive or Gram-negative bacteria. In this study, we determined the nature of this immunosuppressive property. Since phosphorylcholine (PC), a major haptenic component of teichoic acid in the Pn cell wall, and lipoteichoic acid in the Pn membrane, was previously reported to be immunosuppresive when derived from filarial parasites, we determined whether R36A lacking PC (R36Apc-) was inhibitory. Indeed, although R36Apc- exhibited a markedly reduced level of inhibition of the IgG response to co-immunized cOVA, no inhibition was observed when using several other distinct PC-expressing bacteria or a soluble, protein-PC conjugate. Further, treatment of R36A with periodate, which selectively destroys PC residues, had no effect on R36A-mediated inhibition. Since R36Apc- also lacks choline-binding proteins (CBPs), that require PC for cell wall attachment, and since treatment of R36A with trypsin eliminated its inhibitory activity, we incubated R36A in choline chloride, which selectively strips CBPs from its surface. R36A lacking CBPs lost most of its inhibitory property, whereas the supernatant of choline chloride-treated R36A, containing CBPs, was markedly inhibitory. Co-immunization studies using cOVA and various Pn mutants, each genetically deficient in one of the CBPs, demonstrated that only Pn lacking the CBP, pneumococcal surface protein A (PspA), lost its ability to inhibit the IgG anti-cOVA response. These results strongly suggest that PspA plays a major role in mediating the immunosuppressive property of Pn.