Previous studies have demonstrated an adjuvant effect for the C3d fragment of complement C3 when coupled to T-dependent protein antigens. In this study, we examined the antibody response to covalent conjugates of C3d and a T-independent antigen, the capsular polysaccharide of serotype 14 Streptococcus pneumoniae (PPS14). We prepared a conjugate of mouse C3d and PPS14 and compared its immunogenicity with that of a conjugate of PPS14 and ovalbumin (OVA). When BALB/c mice were immunized with PPS14-C3d, there was a significant increase in serum anti-PPS14 concentrations compared with either native PPS14 or control PPS14-glycine conjugates. This was accompanied by a switch in anti-PPS14 from predominantly immunoglobulin M (IgM) to IgG1 by day 25 following primary immunization. Following secondary immunization with PPS14-C3d, there was a marked booster response and a further increase in the ratio of IgG1 to IgM anti-PPS14. Although the primary antibody response to the PPS14-OVA conjugate exceeded that induced by immunization with PPS14-C3d, serum anti-PPS14 concentrations after a second injection of PPS14-C3d were nearly identical to those induced by secondary immunization with PPS14-OVA. Experiments with athymic nude mice suggested that T cells were not required for the adjuvant effect of C3d on the primary immune response to PPS14 but were necessary for enhancement of the memory response after a second injection of PPS14-C3d. These studies show that the adjuvant effects of C3d extend to T-independent antigens as well as T-dependent antigens. As a means of harnessing the adjuvant potential of the innate immune system, C3d conjugates may prove useful as a component of vaccines against encapsulated bacteria.

Serum antibodies (Abs) specific for the capsular polysaccharides of Streptococcus pneumoniae provide protection against invasive pneumococcal disease. Previous studies indicate that Abs to pneumococcal polysaccharide (PPS) serotypes 1 and 6B have limited clonal diversity. To determine if restricted diversity was a feature common to other PPS specificities, we examined the light (L)-chain expression and isoelectric heterogeneity of type 6B, 14, and 23F Abs elicited in 15 adults following PPS vaccination. At the population level, both PPS-6B and PPS-14 Abs expressed and chains, although 6B Abs more frequently expressed chains lambda and 14 Abs more frequently expressed chains. In individual sera, Abs were generally skewed towards either or expression. 23F-specific Abs had predominantly chains. Isoelectric focusing analyses showed that sera contained one or at most a few immunoglobulin G Ab spectrotypes to all three respective capsular serotypes, a result indicative of oligoclonality. A sequence analysis of a purified PPS-14-specific Ab having a single spectrotype gave uniform amino-terminal sequences for both the heavy chain (V H III subgroup) and the L chain (III-A27 V region). From these results we conclude that within individual adults, serum Ab responses to PPS serotypes 6B, 14, and 23F derive from a small number of dominant B-cell clones, and consequently variable-region expression is probably individually limited as well. Oligoclonality appears to be a general characteristic of human PPS-specific Ab repertoires, and we suggest that this property could lead to individual differences in Ab fine specificity and/or functional activity against encapsulated pneumococci.

No abstract