The diversity of strains observed underscores the importance of studying the distribution of the vaccine antigen itself rather than relying on common epidemiological surrogates such as MLST.

Staphylococcus aureus can cause severe life threatening invasive diseases. The principal immune effector mechanism by which humans are protected from Gram positive bacteria such as S. aureus is antigen specific antibody- and complement-dependent opsonophagocytosis. This process can be measured in vitro using the opsonophagocytic antibody assay (OPA), which is a complex assay composed of live S. aureus bacteria, a complement source, phagocytic effector cells such as differentiated HL-60 cells, and test serum. In this report, we investigated the impact on the OPA of S. aureus surface antigens capsular polysaccharides (CP) and protein A (SpA). We demonstrated that higher CP expression renders bacteria more resistant to non-specific opsonophagocytic killing than increased SpA expression, suggesting that the expression of capsular polysaccharides may be the more important immune evasion strategy for S. aureus. Bacteria that were not fully encapsulated were highly susceptible to non-specific killing in the assay in the absence of immune serum. This non-specific killing was prevented by growing the bacteria under conditions that increased capsular polysaccharide levels on the surface of the bacteria. In contrast, the level of SpA expression had no detectable effect on non-specific killing in OPA. Using anti-CP antibodies we demonstrated type-specific killing in OPA of both MRSA and MSSA clinical isolates. SpA expression on the cell surface did not interfere with OPA activity, providing evidence that despite the role of SpA in sequestering antibodies by their Fc region, killing is easily accomplished in the presence of high titered anti-capsular polysaccharide antibodies. This highlights the role of CP as an important immune evasion mechanism and supports the inclusion of capsular polysaccharide antigens in the formulation of multi-component prophylactic vaccines against S. aureus.

LP2086 is a family of outer membrane lipoproteins fromNeisseria meningitidis, which elicits bactericidal antibodies and are currently undergoing human clinical trials in a bivalent formulation where each antigen represents one of the two known LP2086 subfamilies. Here we report the NMR structure of the recombinant LP2086 variant B01, a representative of the LP2086 subfamily B. The structure reveals a novel fold composed of two domains: a "taco-shaped" N-terminal ␤-sheet and a C-terminal ␤-barrel connected by a linker. The structure in micellar solution is consistent with a model of LP2086 anchored to the outer membrane bilayer through its lipidated N terminus. A long flexible chain connects the folded part of the protein to the lipid anchor and acts as spacer, making both domains accessible to the host immune system. Antibodies broadly reactive against members from both subfamilies have been mapped to the N terminus. A surface of subfamily-defining residues was identified on one face of the protein, offering an explanation for the induction of subfamily-specific bactericidal antibodies.Neisseria meningitidis is a Gram-negative bacterial pathogen, which colonizes the upper respiratory tract, occasionally invading the bloodstream, causing sepsis, and crossing the blood-brain barrier, resulting in meningitis. Despite the availability of effective antibiotic treatment, the rapid progression of meningococcal disease still results in substantial morbidity and mortality (1). Five meningococcal serogroups, categorized according to the chemical structure of the bacterial capsular polysaccharides, A, B, C, Y, and W135, account for most of the disease (2). Although a vaccine against four of the five major serogroups of meningococci is currently available, a vaccine for the prevention of serogroup B disease is still an unmet clinical need (3). The development of vaccines against serogroup B meningococci has focused on subcapsular antigens, in order to avoid the risk of autoimmunity arising from structural similarities between the capsular polysaccharides and the sialic acidmodified surface of developing human brain (1,4,5).Recently, a new family of lipidated outer membrane proteins, LP2086, was identified as a potential vaccine target (6). Members of the LP2086 family have been divided into two subfamilies, subfamily A and B, based on their genetic variation (6, 7). Since recombinant LP2086 (rLP2086) 3 elicits a bactericidal response that is largely subfamily-specific, a bivalent vaccine containing one protein from each subfamily will offer protection against serogroup B meningococci (6, 8 -11). LP2086 lipoproteins are lipidated at the N-terminal Cys with a tripalmitoyl lipid tail, which anchors the protein to the bacterial membrane (12). More recently, LP2086 was found to induce serum resistance via binding with human Factor H, a key regulator of the alternative complement pathway that prevents autologous complement attack (13).Our work seeks to understand the structural elements of LP2086 responsible for inducing the subf...

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