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Background: Ventilator associated pneumonia (VAP) is a serious infection among patients in the intensive care unit (ICU). Methods: We reviewed the medical charts of all patients admitted to the adult intensive care units of the Massachusetts General Hospital that went on to develop VAP during a five year period. Results: 200 patients were included in the study of which 50 (25%) were infected with a multidrug resistant pathogen. Increased age, dialysis and late onset (≥5 days from admission) VAP were associated with increased incidence of resistance. Multidrug resistant bacteria (MDRB) isolation was associated with a significant increase in median length of ICU stay (19 vs. 16 days, p = 0.02) and prolonged duration of mechanical ventilation (18 vs. 14 days, p = 0.03), but did not impact overall mortality (HR 1.12, 95% CI 0.51–2.46, p = 0.77). However, age (HR 1.04 95% CI 1.01–1.07, p = 0.003) was an independent risk factor for mortality and age ≥65 years was associated with increased incidence of methicillin-resistant Staphylococcus aureus (MRSA) infections (OR 2.83, 95% CI 1.27–6.32, p = 0.01). Conclusions: MDRB-related VAP is associated with prolonged ICU stay and mechanical ventilation. Interestingly, age ≥ 65 years is associated with MRSA VAP.

The objectives of this study were to investigate the immune response to intradermal immunization with wall teichoic acid (WTA) and the effect of MBL deficiency in a murine model of infection with methicillin-resistant Staphylococcus aureus (MRSA). WTA is a bacterial cell wall component that is implicated in invasive infection. We tested susceptibility to MRSA infection in wild type (WT) and MBL deficient mice using two strains of MRSA: MW2, a community-associated MRSA (CA-MRSA); and COL, a healthcare-associated MRSA (HA-MRSA). We also performed in vitro assays to investigate the effects of anti-WTA IgG containing murine serum on complement activation and bacterial growth in whole blood. We found that MBL knockout (KO) mice are relatively resistant to a specific MRSA strain, MW2 CA-MRSA, compared to WT mice, while both strains of mice had similar susceptibility to a different strain, COL HA-MRSA. Intradermal immunization with WTA elicited and augmented an anti-WTA IgG response in both WT and MBL KO mice. WTA immunization significantly reduced susceptibility to both MW2 CA-MRSA and COL HA-MRSA, independent of the presence of MBL. The protective mechanisms of anti-WTA IgG are mediated at least in part by complement activation and clearance of bacteria from blood. The significance of these findings is that 1) Intradermal immunization with WTA induces production of anti-WTA IgG; and 2) This anti-WTA IgG response protects from infection with both MW2 CA-MRSA and COL HA-MRSA even in the absence of MBL, the deficiency of which is common in humans.

Background: Patterns of methicillin-resistant S. aureus (MRSA) nasal carriage over time and across the continuum of care settings are poorly characterized. Knowledge of prevalence rates and outcomes associated with MRSA nasal carriage patterns could help direct infection prevention strategies. The VA integrated health-care system and active surveillance program provides an opportunity to delineate nasal carriage patterns and associated outcomes of death, infection, and conversion in carriage. Methods/Findings: We conducted a retrospective cohort study including all patients admitted to 5 acute care VA hospitals between 2008–2010 who had nasal MRSA PCR testing within 48 hours of admission and repeat testing within 30 days. The PCR results were used to define a baseline nasal carriage pattern of never, intermittently, or always colonized at 30 days from admission. Follow-up was up to two years and included acute, long-term, and outpatient care visits. Among 18,038 patients, 91.1%, 4.4%, and 4.6% were never, intermittently, or always colonized at the 30-day baseline. Compared to non-colonized patients, those who were persistently colonized had an increased risk of death (HR 2.58; 95% CI 2.18;3.05) and MRSA infection (HR 10.89; 95% CI 8.6;13.7). Being in the non-colonized group at 30 days had a predictive value of 87% for being non-colonized at 1 year. Conversion to MRSA colonized at 6 months occurred in 11.8% of initially non-colonized patients. Age >70 years, long-term care, antibiotic exposure, and diabetes identified >95% of converters. Conclusions: The vast majority of patients are not nasally colonized with MRSA at 30 days from acute hospital admission. Conversion from non-carriage is infrequent and can be risk-stratified. A positive carriage pattern is strongly associated with infection and death. Active surveillance programs in the year following carriage pattern designation could be tailored to focus on non-colonized patients who are at high risk for conversion, reducing universal screening burden.

Staphylococcus aureus is a major cause of nosocomial and community-acquired infections for which a vaccine is greatly desired. Antigens found on the S. aureus outer surface include the capsular polysaccharides (CP) of serotype 5 (CP5) or 8 (CP8) and/or a second antigen, a β-(1→6)-polymer of N-acetyl-D-glucosamine (PNAG). Antibodies specific for either CP or PNAG antigens have excellent in vitro opsonic killing activity (OPKA), but when mixed together have potent interference in OPKA and murine protection. To ascertain if this interference could be abrogated by using a synthetic non-acetylated oligosaccharide fragment of PNAG, 9GlcNH2, in place of chemically partially deacetylated PNAG, three conjugate vaccines consisting of 9GlcNH2 conjugated to a non-toxic mutant of alpha-hemolysin (Hla H35L), CP5 conjugated to clumping factor B (ClfB), or CP8 conjugated to iron-surface determinant B (IsdB) were used separately to immunize rabbits. Opsonic antibodies mediating killing of multiple S. aureus strains were elicited for all three vaccines and showed carbohydrate antigen-specific reductions in the tissue bacterial burdens in animal models of S. aureus skin abscesses, pneumonia, and nasal colonization. Carrier-protein specific immunity was also shown to be effective in reducing bacterial levels in infected lungs and in nasal colonization. However, use of synthetic 9GlcNH2 to induce antibody to PNAG did not overcome the interference in OPKA engendered when these were combined with antibody to either CP5 or CP8. Whereas each individual vaccine showed efficacy, combining antisera to CP antigens and PNAG still abrogated individual OPKA activities, indicating difficulty in achieving a multi-valent vaccine targeting both the CP and PNAG antigens.

The increasing frequency, severity and antimicrobial resistance of Staphylococcus aureus infections has made the development of immunotherapies against this pathogen more urgent than ever. Previous immunization attempts using monovalent antigens resulted in at best partial levels of protection against S. aureus infection. We therefore reasoned that synthesizing a bivalent conjugate vaccine composed of two widely expressed antigens of S. aureus would result in additive/synergetic activities by antibodies to each vaccine component and/or in increased strain coverage. For this we used reductive amination, to covalently link the S. aureus antigens clumping factor A (ClfA) and deacetylated poly-N-β-(1–6)-acetyl-glucosamine (dPNAG). Mice immunized with 1, 5 or 10 µg of the dPNAG-ClfA conjugate responded in a dose-dependent manner with IgG to dPNAG and ClfA, whereas mice immunized with a mixture of ClfA and dPNAG developed significantly lower antibody titers to ClfA and no antibodies to PNAG. The dPNAG-ClfA vaccine was also highly immunogenic in rabbits, rhesus monkeys and a goat. Moreover, affinity-purified, antibodies to ClfA from dPNAG-ClfA immune serum blocked the binding of three S. aureus strains to immobilized fibrinogen. In an opsonophagocytic assay (OPKA) goat antibodies to dPNAG-ClfA vaccine, in the presence of complement and polymorphonuclear cells, killed S. aureus Newman and, to a lower extent, S. aureus Newman ΔclfA. A PNAG-negative isogenic mutant was not killed. Moreover, PNAG antigen fully inhibited the killing of S. aureus Newman by antisera to dPNAG-ClfA vaccine. Finally, mice passively vaccinated with goat antisera to dPNAG-ClfA or dPNAG-diphtheria toxoid conjugate had comparable levels of reductions of bacteria in the blood 2 h after infection with three different S. aureus strains as compared to mice given normal goat serum. In conclusion, ClfA is an immunogenic carrier protein that elicited anti-adhesive antibodies that fail to augment the OPK and protective activities of antibodies to the PNAG cell surface polysaccharide.

Staphylococcus epidermidis (SE) causes late onset sepsis and significant morbidity in catheterized preterm newborns. Animal models of SE infection are useful in characterizing disease mechanisms and are an important approach to developing improved diagnostics and therapeutics. Current murine models of neonatal bacterial infection employ intraperitoneal or subcutaneous routes at several days of age, and may, therefore, not accurately reflect distinct features of innate immune responses to bacteremia. In this study we developed, validated, and characterized a murine model of intravenous (IV) infection in neonatal mice <24 hours (h) old to describe the early innate immune response to SE. C57BL/6 mice <24 h old were injected IV with 106, 107, 108 colony-forming units (CFU) of SE 1457, a clinical isolate from a central catheter infection. A prospective injection scoring system was developed and validated, with only high quality injections analyzed. Newborn mice were euthanized between 2 and 48 h post-injection and spleen, liver, and blood collected to assess bacterial viability, gene expression, and cytokine production. High quality IV injections demonstrated inoculum-dependent infection of spleen, liver and blood. Within 2 h of injection, SE induced selective transcription of TLR2 and MyD88 in the liver, and increased systemic production of plasma IL-6 and TNF-α. Despite clearance of bacteremia and solid organ infection within 48 h, inoculum-dependent impairment in weight gain was noted. We conclude that a model of IV SE infection in neonatal mice <24 h old is feasible, demonstrating inoculum-dependent infection of solid organs and a pattern of bacteremia, rapid and selective innate immune activation, and impairment of weight gain typical of infected human neonates. This novel model can now be used to characterize immune ontogeny, evaluate infection biomarkers, and assess preventative and therapeutic modalities.