Viliam Pavliak

Heterogeneous in vivo expression of clumping factor A and capsular polysaccharide by Staphylococcus aureus: Implications for vaccine design

There is a clear unmet medical need for a vaccine that would prevent infections from Staphylococcus aureus (S. aureus). To validate antigens as potential vaccine targets it has to be demonstrated that the antigens are expressed in vivo. Using murine bacteremia and wound infection models, we demonstrate that the expression of clumping factor A (ClfA) and capsular polysaccharide antigens are heterogeneous and dependent on the challenge strains examined and the in vivo microenvironment. We also demonstrate opsonophagocitic activity mediated by either antigen is not impeded by the presence of the other antigen. The data presented in this report support a multiantigen approach for the development of a prophylactic S. aureus vaccine to ensure broad coverage against this versatile pathogen.

Identification of the common antigenic determinant shared by Streptococcus pneumoniae serotypes 33A, 35A, and 20 capsular polysaccharides

In order to better understand cross-reactions of serogroup 33 polysaccharides and the typing sera, the structure of pneumococcal capsular polysaccharide serotype 33A was elucidated. Serotype 33A has been shown to have an identical polysaccharide backbone as that of serotype 33F, with two additional sites of O-acetylation at C5, and C6 of the 3-β-Galf residue in serotype 33A. This finding is consistent with the presence of an additional functional acetyltransferase gene (wcjE) in the cps biosynthetic locus of serotype 33A compared to 33F. The identical polysaccharide backbone with at least one common O-acetylation site (C2 of 5-β-Galf) shared by serotype 33A and 33F polysaccharides is proposed to be the epitope recognized by typing serum 33b. In addition, a 5,6-di-O-acetylated →3)-β-d-Galf5,6Ac-(1→3)-β-d-Glcp-(1→ disaccharide unit, a common structural motif present in serotypes 33A, 20, and 35A polysaccharides, is proposed to be the antigenic determinant recognized by typing serum 20b.

NMR structural analysis of the capsular polysaccharide from Streptococcus pneumoniae serotype 6C.

The original structure of Streptococcus pneumoniae capsular polysaccharide (CPS) serotype 6C was proposed based on chemical degradation and tandem mass analysis [J. Clin. Microbiol.2007, 45, 1225-1233]. In order to confirm the repeat unit structure and assign the stereochemical structure, the CPS 6C and the known CPS 6A were fully characterized by NMR spectroscopy. Full (1)H and (13)C NMR spectra assignments of CPS 6C and CPS 6A were achieved based on DQCOSY, TOCSY, HSQC, HMBC, and NOESY analysis. These analyses confirmed the published structure of CPS 6A and established the repeat unit structure of the CPS 6C as: →2)-α-D-Glcp-(1→3)-α-D-Glcp-(1→3)-α-L-Rhap-(1→3)-D-Ribitol-(5→phosphate-.

Structure elucidation of capsular polysaccharides from Streptococcus pneumoniae serotype 33C, 33D, and revised structure of serotype 33B

We report herein the previously unknown structures of the pneumococcal capsular polysaccharides serotype 33C and 33D, and a revised structure of serotype 33B. The syntenic pair 33B/33D has nearly identical polysaccharide repeat units with the exception of one sugar residue (→2-α-Glcp in 33B and →2-α-Galp in 33D). Serotype 33C is structurally more similar to 33B/33D than 33A/33F, in that it also possesses a backbone ribitol-phosphate group and a →3-β-GalpNAc residue, both of which are absent in the repeat units of 33A/33F. Serotype 33C is notably different from all other serogroup 33 polysaccharides, as there is no →3-β-Glcp residue and the location of the O-acetylation of the →5-β-Galf residue (O-6) differs from the other serogroup 33 polysaccharides (O-2). This completes the structural assignments of polysaccharides within serogroup 33 and provides a framework for understanding the recognition of epitopes by serogroup 33 typing sera based on observed cross-reactivities reported in the literature.

O-Acetylation is essential for functional antibody generation against Staphylococcus aureus capsular polysaccharide

ABSTRACT Staphylococcus aureus produces an antiphagocytic polysaccharide capsule to evade neutrophil-mediated killing. Many vaccines against encapsulated bacterial pathogens require generation of functional anti-capsular antibodies to mediate protection against infection and disease. Here it is shown that the generation of such antibody responses to S. aureus in vivo and in vitro requires the presence of O-acetyl modifications on the capsular polysaccharides. O-acetylation of S. aureus capsular polysaccharide therefore should be monitored carefully during vaccine development and production. This finding may provide additional insight into the previous failure of a S. aureus capsular polysaccharide conjugate vaccine.