Proteomics-based identification of anchorless cell wall proteins as vaccine candidates against Staphylococcus aureus.

Staphylococcus aureus is an important human pathogen with increasing clinical impact due to the extensive spread of antibiotic-resistant strains. Therefore, development of a protective polyvalent vaccine is of great clinical interest. We employed an intravenous immunoglobulin (IVIG) preparation as a source of antibodies directed against anchorless S. aureus surface proteins for identification of novel vaccine candidates. In order to identify such proteins, subtractive proteome analysis (SUPRA) of S. aureus anchorless cell wall proteins was performed. Proteins reacting with IVIG but not with IVIG depleted of S. aureus-specific opsonizing antibodies were considered vaccine candidates. Nearly 40 proteins were identified by this preselection method using matrix-assisted laser desorption ionization--time of flight analysis. Three of these candidate proteins, enolase (Eno), oxoacyl reductase (Oxo), and hypothetical protein hp2160, were expressed as glutathione S-transferase fusion proteins, purified, and used for enrichment of corresponding immunoglobulin Gs from IVIG by affinity chromatography. Use of affinity-purified anti-Eno, anti-Oxo, and anti-hp2160 antibodies resulted in opsonization, phagocytosis, and killing of S. aureus by human neutrophils. High specific antibody titers were detected in mice immunized with recombinant antigens. In mice challenged with bioluminescent S. aureus, reduced staphylococcal spread was measured by in vivo imaging. The recovery of S. aureus CFU from organs of immunized mice was diminished 10- to 100-fold. Finally, mice immunized with hp2160 displayed statistically significant higher survival rates after lethal challenge with clinically relevant S. aureus strains. Taken together, our data suggest that anchorless cell wall proteins might be promising vaccine candidates and that SUPRA is a valuable tool for their identification.

The interferon-inducible p47 (IRG) GTPases in vertebrates: loss of the cell autonomous resistance mechanism in the human lineage.

BACKGROUND: Members of the p47 (immunity-related GTPases (IRG) family) GTPases are essential, interferon-inducible resistance factors in mice that are active against a broad spectrum of important intracellular pathogens. Surprisingly, there are no reports of p47 function in humans. RESULTS: Here we show that the p47 GTPases are represented by 23 genes in the mouse, whereas humans have only a single full-length p47 GTPase and an expressed, truncated presumed pseudo-gene. The human full-length gene is orthologous to an isolated mouse p47 GTPase that carries no interferon-inducible elements in the promoter of either species and is expressed constitutively in the mature testis of both species. Thus, there is no evidence for a p47 GTPase-based resistance system in humans. Dogs have several interferon-inducible p47s, and so the primate lineage that led to humans appears to have lost an ancient function. Multiple p47 GTPases are also present in the zebrafish, but there is only a tandem p47 gene pair in pufferfish. CONCLUSION: Mice and humans must deploy their immune resources against vacuolar pathogens in radically different ways. This carries significant implications for the use of the mouse as a model of human infectious disease. The absence of the p47 resistance system in humans suggests that possession of this resistance system carries significant costs that, in the primate lineage that led to humans, are not outweighed by the benefits. The origin of the vertebrate p47 system is obscure.