Comparative Opsonic and Protective Activities of Staphylococcus aureus Conjugate Vaccines Containing Native or Deacetylated Staphylococcal Poly-N-Acetyl- -(1-6)-Glucosamine
ABSTRACT Staphylococcus aureus and Staphylococcus epidermidis both synthesize the surface polysaccharide poly-N-acetyl-beta-(1-6)-glucosamine (PNAG), which is produced in vitro with a high level (>90%) of the amino groups substituted by acetate. Here, we examined the role of the acetate substituents of PNAG in generating opsonic and protective antibodies. PNAG and a deacetylated form of the antigen (dPNAG; 15% acetylation) were conjugated to the carrier protein diphtheria toxoid (DT) and used to immunize animals. Mice responded in a dose-dependent fashion to both conjugate vaccines, with maximum antibody titers observed at the highest dose and 4 weeks after the last of three weekly immunizations. PNAG-DT and dPNAG-DT vaccines were also very immunogenic in rabbits. Antibodies raised to the conjugate vaccines in rabbits mediated the opsonic killing of various staphylococcal strains, but the specificity of the opsonic killing was primarily to dPNAG, as this antigen inhibited the killing of S. aureus strains by both PNAG- and dPNAG-specific antibodies. Passive immunization of mice with anti-dPNAG-DT rabbit sera showed significant levels of clearance of S. aureus from the blood (54 to 91%) compared to control mice immunized with normal rabbit sera, whereas PNAG-specific antibodies were ineffective at clearing S. aureus. Passive immunization of mice with a goat antiserum raised to the dPNAG-DT vaccine protected against a lethal dose of three different S. aureus strains. Overall, these data show that immunization of animals with a conjugate vaccine of dPNAG elicit antibodies that mediated opsonic killing and protected against S. aureus infection, including capsular polysaccharide types 5 and 8 and an untypable strain.
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ABSTRACT: Polysaccharide intercellular adhesin (PIA), also known as poly-N-acetyl-β-(1-6)-glucosamine (PIA/PNAG) is an important component of Staphylococcus aureus biofilms and also contributes to resistance to phagocytosis. The proteins IcaA, IcaD, IcaB, and IcaC are encoded within the intercellular adhesin (ica) operon and synthesize PIA/PNAG. We discovered a mechanism of phase variation in PIA/PNAG expression that appears to involve slipped-strand mispairing. The process is reversible and RecA-independent, and involves the expansion and contraction of a simple tetranucleotide tandem repeat within icaC. Inactivation of IcaC results in a PIA/PNAG-negative phenotype. A PIA/PNAG-hyperproducing strain gained a fitness advantage in vitro following the icaC mutation and loss of PIA/PNAG production. The mutation was also detected in two clinical isolates, suggesting that under certain conditions, loss of PIA/PNAG production may be advantageous during infection. There was also a survival advantage for an icaC-negative strain harboring intact icaADB genes relative to an isogenic icaADBC deletion mutant. Together, these results suggest that inactivation of icaC is a mode of phase variation for PIA/PNAG expression, that high-level production of PIA/PNAG carries a fitness cost, and that icaADB may contribute to bacterial fitness, by an unknown mechanism, in the absence of an intact icaC gene and PIA/PNAG production.PLoS Pathogens 07/2014; 10(7):e1004292. DOI:10.1371/journal.ppat.1004292 · 8.14 Impact Factor
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ABSTRACT: As an immune-privileged site, the eye, and particularly the outer corneal surface, lacks resident mature immune effector cells. Physical barriers and innate mediators are the best-described effectors of immunity in the cornea. When breached, infection can result in rapid tissue destruction leading to loss of visual acuity and frank blindness. To determine the cellular and molecular components needed for effective adaptive immunity on the corneal surface we investigated which immune system effectors were required for protection against Staphylococcus aureus corneal infections in mice, which is a serious cause of human eye infections. Both systemically-injected and topically-applied antibody to the conserved cell surface polysaccharide poly-N-acetyl glucosamine (PNAG) were effective at mediating reductions in corneal pathology and bacterial levels. Additional host factors impacting protection included ICAM1-dependent PMN recruitment, functional CD4(+) T cells, signaling via the IL-17 receptor, and IL-22 production. In germ-free mice there was no protective efficacy of antibody to PNAG due to lack of LY6G(+) inflammatory cell co-effector recruitment to the cornea. Protection was manifest after three weeks of exposure to conventional mice and acquisition of a resident microbiota. We conclude that in the anterior eye, ICAM-1 mediated PMN recruitment to the infected cornea along with endogenous microbiota-matured CD4(+) T cells producing both IL-17 and IL-22 are required for antibody to PNAG to protect against S. aureus infection.Infection and Immunity 06/2014; 82(8). DOI:10.1128/IAI.01951-14 · 4.16 Impact Factor
Advances in Experimental Medicine and Biology 01/2015; 831:69-91. DOI:10.1007/978-3-319-09782-4_6 · 2.01 Impact Factor