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Streptococcus pneumoniae is an important pathogen that causes disease in young and elderly individuals. The currently available polysaccharide vaccines have limited efficacy in those age groups most susceptible to pneumococcal infections. This study focuses on mapping the epitopes of a surface protein of S. pneumoniae by biopanning a 15 mer phage display library using 5 different monoclonal antibodies (MAbs) against the Pneumoccal surface adhesin A (PsaA). PsaA is a component of the bacterial cell wall that is highly species specific and is involved in bacterial adherence and virulence. Biopanning of the phage display library reveals three distinct epitopes on the PsaA protein. The sequence homology of these epitopes ranges from two to six amino acids when compared to the native PsaA protein type 2. Two of these epitopes have been evaluated for their immunogeneicity in mice. The peptide selected by the MAbs 8G12, 6F6, and 1B7 is referred to as the consensus peptide and is immunogenic in mice. Optimal anti-PsaA response is observed in mice immunized with 50microg of the consensus peptide complexed to proteosomes in 1:1 ratio. The anti-PsaA response is significantly lower than the response to the PsaA native protein. The peptide selected by monoclonal antibody 4E9 in its lipidated form is significantly protective in mice challenged with S. pneumoniae serotype 2 when compared to mice immunized with the native protein. These results show that the selected epitopes of PsaA protein are immunogenic and protective in mice. These epitopes need to be evaluated further as alternatives to currently available vaccines.

Streptococcus pneumoniae is a human bacterial pathogen responsible for serious infections including pneumonia. The currently licensed polysaccharide vaccine provides 60 to 80% protection in young adults, but in the elderly the vaccine efficacy is drastically reduced despite normal antibody levels. We hypothesized that the reduced vaccine efficacy in the elderly results from altered variable gene family usage. We have analyzed the light chain gene usage in 20 young (20 to 30 years of age) and 20 elderly (65 to 86 years of age) adults in response to pneumococcal polysaccharide 4 (PPS4) and PPS14. We generated a variable light chain library using B cells specific for PPS4 and PPS14 from each vaccinated individual. We determined complete sequences and somatic mutation frequencies in all isolated variable light chain fragments. Six gene families, 1, 2, 3, 4, 1, and 3, were identified in response to PPS4 and PPS14 in both age groups. Comparison of young and elderly adults demonstrated significant differences in 4, 1, and 3 gene usage in response to PPS4 and PPS14. With aging, there was a significant increase in 4 gene usage and a significant decrease in 1 and 3 gene usage in response to both PPS4 and PPS14. Although both V1 and V3 gene products demonstrated extensive mutations, there was no age-related difference in mutational frequency per gene family. These findings suggest an age-related change in light chain gene usage in response to PPS4 and PPS14.The incidence of pneumococcal pneumonia is significantly increased in individuals Ͼ65 years of age (13). Several studies (8,18) have demonstrated a significant impairment in the immune response to pneumococcal polysaccharides in the elderly, particularly those Ͼ77 years of age, associated with a markedly decreased efficacy of the pneumococcal vaccine (6, 23).The molecular mechanisms responsible for the decreased immune response in the elderly remain poorly understood. Nicoletti et al. (15) studied the immune response to phosphorylcholine (PC) in aged mice vaccinated with Streptococcus pneumoniae. These studies demonstrated a significant functional impairment in the immune response to PC despite normal antibody concentrations. Molecular analysis of the anti-PC-specific antibody repertoire in aged mice revealed a significantly altered V gene usage compared to young immunized mice (14).To date, studies examining the structure-function relationship of antipneumococcal antibodies have been performed using human monoclonal antibodies (1, 22, 26), combinatorial libraries (12,17,27,28), and monoclonal antibodies derived from a transgenic mouse strain with human immunoglobulin loci (2, 19). All studies, with the exception of those using the transgenic mouse, have analyzed the V repertoire using peripheral blood mononuclear cells obtained from young high responders. Although these studies have enriched our knowledge concerning the human immune response to pneumococcal polysaccharide (PPS), they were not designed to represent the in vivo antibody repertoire response to PPS in the...