Design of Staphylococcus aureus New Vaccine Candidates with B and T Cell Epitope Mapping, Reverse Vaccinology, and Immunoinformatics.

An effective vaccine against Staphylococcus aureus infection is a major planetary heath priority, particularly with increasing antibiotic resistance worldwide. Previous efforts for a highly effective S. aureus vaccine were largely unsuccessful, in part, because the vaccine designs have tended to target mainly the B cell immunity and development of opsonic antibodies. In contrast, recent observations suggest that cell mediated immunity may be critical for protection against S. aureus. In addition, the S. aureus surface proteins are among the key immunodominant antigens because they are the first molecules to interact with the host organism cells and tissues. We report here an original vaccinomics study in which we used a reverse vaccinology and immunoinformatics in silico strategy integrated with genomics. After analyzing 2767 proteins, we defined 16 proteins of S. aureus as promising subunit vaccine candidates. Phosphatidylinositol phosphodiesterase (Plc) is secreted by extracellular pathogens such as S. aureus. We mapped the B and T cell epitopes for the Plc protein, tested the reactivity of the synthesized epitopes by Western blotting, and verified our findings in a pilot study of 10 patients with S. aureus infection. The peptides were then tested for their protective effect in groups of mice challenged with pathogenic S. aureus strain, which showed high protection level. These findings warrant further translational research for development of novel vaccines against S. aureus infection. Reverse vaccinology is an advanced approach that can be applied to identify new vaccine candidates against a host of microorganisms, including S. aureus.

Interleukin-10 (IL-10) Produced by Mutant Toxic Shock Syndrome Toxin 1 Vaccine-Induced Memory T Cells Downregulates IL-17 Production and Abrogates the Protective Effect against Staphylococcus aureus Infection.

Development of long-term memory is crucial for vaccine-induced adaptive immunity against infectious diseases such as Staphylococcus aureus infection. Toxic shock syndrome toxin 1 (TSST-1), one of the superantigens produced by S. aureus, is a possible vaccine candidate against infectious diseases caused by this pathogen. We previously reported that vaccination with less toxic mutant TSST-1 (mTSST-1) induced T helper 17 (Th17) cells and elicited interleukin-17A (IL-17A)-mediated protection against S. aureus infection 1 week after vaccination. In the present study, we investigated the host immune response induced by mTSST-1 vaccination in the memory phase, 12 weeks after the final vaccination. The protective effect and IL-17A production after vaccination with mTSST-1 were eliminated because of IL-10 production. In the presence of IL-10-neutralizing monoclonal antibody (mAb), IL-17A production was restored in culture supernatants of CD4+ T cells and macrophages sorted from the spleens of vaccinated mice. Vaccinated mice treated with anti-IL-10 mAb were protected against systemic S. aureus infection in the memory phase. From these results, it was suggested that IL-10 produced in the memory phase suppresses the IL-17A-dependent vaccine effect through downregulation of IL-17A production.

Immunogenicity of a Staphylococcus aureus-cholera toxin A2/B vaccine for bovine mastitis.

Staphylococcus aureus causes a chronic, contagious disease of the udder, or mastitis, in dairy cows. This infection is often refractory to antibiotic treatment, and has a significant economic impact on milk production worldwide. An effective vaccine to prevent S. aureus mastitis would improve animal health, reduce antibiotic dependence and inform human vaccine approaches. The iron-regulated surface determinant A (IsdA) and clumping factor A (ClfA) are conserved S. aureus extracellular-matrix adhesins and target vaccine antigens. Here we report the results of two bovine immunogenicity trials using purified IsdA and ClfA-cholera toxin A2/B chimeras (IsdA-CTA2/B and ClfA-CTA2/B). Cows were intranasally inoculated with IsdA-CTA2/B + ClfA-CTA2/B at dry off and followed for 70 days. Trial 1 utilized three groups with one or two booster doses at a total concentration of 600 or 900 µg. Trial 2 utilized two groups with one booster at a total concentration of 1200 µg. Humoral immune responses in serum and milk were examined by ELISA. Responses in serum were significant between groups and provide evidence of antigen-specific IgG induction after vaccination in both trials. Cellular proliferation was detected by flow cytometry using antigen-stimulated PBMCs from day 60 of Trial 2 and revealed an increase in CD4+ T cells from vaccinated cows. IsdA and ClfA stimulation induced IL-4 expression, but not IFN-γ or IL-17, in PBMCs from day 60 as determined by cytokine expression analysis. Opsonophagocytosis of S. aureus confirmed the functional in vitro activity of anti-IsdA antibodies from Trial 2 serum and milk. The vaccine was well tolerated and safe, and results support the potential of mucosally-delivered CTA2/B chimeras to protect cows from mastitis caused by S. aureus.

SA4Ag, a 4-antigen Staphylococcus aureus vaccine, rapidly induces high levels of bacteria-killing antibodies.

BACKGROUND: Staphylococcus aureus is a leading cause of healthcare-associated infections. No preventive vaccine is currently licensed. SA4Ag is an investigational 4-antigen S. aureus vaccine, composed of capsular polysaccharide conjugates of serotypes 5 and 8 (CP5 and CP8), recombinant surface protein clumping factor A (rmClfA), and recombinant manganese transporter protein C (rMntC). This Phase 1 study aimed to confirm the safety and immunogenicity of SA4Ag produced by the final manufacturing process before efficacy study initiation in a surgical population. METHODS: Healthy adults (18-<65years) received one intramuscular SA4Ag injection. Serum functional antibodies were measured at baseline and Day 29 post-vaccination. An opsonophagocytic activity (OPA) assay measured the ability of vaccine-induced antibodies to CP5 and CP8 to kill S. aureus clinical isolates. For MntC and ClfA, antigen-specific immunogenicity was assessed via competitive Luminex® immunoassay (cLIA) and via fibrinogen-binding inhibition (FBI) assay for ClfA only. Reactogenicity and adverse event data were collected. RESULTS: One hundred participants were vaccinated. SA4Ag was well tolerated, with a satisfactory safety profile. On Day 29, OPA geometric mean titers (GMTs) were 45,738 (CP5, 95% CI: 38,078-54,940) and 42,652 (CP8, 95% CI: 32,792-55,477), consistent with 69.2- and 28.9-fold rises in bacteria-killing antibodies, respectively; cLIA GMTs were 2064.4 (MntC, 95% CI: 1518.2-2807.0) and 3081.4 (ClfA, 95% CI: 2422.2-3920.0), consistent with 19.6- and 12.3-fold rises, respectively. Similar to cLIA results, ClfA FBI titers rose 11.0-fold (GMT: 672.2, 95% CI: 499.8-904.2). The vast majority of participants achieved the pre-defined biologically relevant thresholds: CP5: 100%; CP8: 97.9%, ClfA: 87.8%; and MntC 96.9%. CONCLUSIONS: SA4Ag was safe, well tolerated, and rapidly induced high levels of bacteria-killing antibodies in healthy adults. A Phase 2B efficacy trial in adults (18-85years) undergoing elective spinal fusion is ongoing to assess SA4Ag's ability to prevent postoperative invasive surgical site and bloodstream infections caused by S. aureus. Clinicaltrials.gov Identifier: NCT02364596.

Protective efficacy of the chimeric Staphylococcus aureus vaccine candidate IC in sepsis and pneumonia models.

Staphylococcus aureus causes serious sepsis and necrotic pneumonia worldwide. Due to the spread of multidrug-resistant strains, developing an effective vaccine is the most promising method for combating S. aureus infection. In this study, based on the immune-dominant areas of the iron surface determinant B (IsdB) and clumping factor A (ClfA), we designed the novel chimeric vaccine IsdB151-277ClfA33-213 (IC). IC formulated with the AlPO4 adjuvant induced higher protection in an S. aureus sepsis model compared with the single components alone and showed broad immune protection against several clinical S. aureus isolates. Immunisation with IC induced strong antibody responses. The protective effect of antibodies was demonstrated through the opsonophagocytic assay (OPA) and passive immunisation experiment. Moreover, this new chimeric vaccine induced Th1/Th17-skewed cellular immune responses based on cytokine profiles and CD4(+) T cell stimulation tests. Neutralisation of IL-17A alone (but not IFN-γ) resulted in a significant decrease in vaccine immune protection. Finally, we found that IC showed protective efficacy in a pneumonia model. Taken together, these data provide evidence that IC is a potentially promising vaccine candidate for combating S. aureus sepsis and pneumonia.

Staphylococcal enterotoxin B mutants (N23K and F44S): biological effects and vaccine potential in a mouse model.

Superantigens produced by Staphylococcus aureus can cause food poisoning and toxic shock syndrome. The biological activities and vaccine potential of mutant staphylococcal enterotoxin B (SEB) proteins, N23K and F44S, were studied in a lipopolysaccharide-potentiated mouse model. Although 10 micrograms of SEB per mouse is equivalent to 30 LD50, the same intraperitoneal dose of either mutant protein was nonlethal and did not elevate serum levels of tumor necrosis factors (TNF). N23K, F44S, and SEB were serologically identical in an enzyme-linked immunosorbent assay with polyclonal anti-SEB. Immunization with alum containing N23K, F44S, or SEB elicited an anti-SEB response that protected 80-87% of the mice against a 10 micrograms SEB challenge. Controls lacking an anti-SEB titer did not survive. Pooled sera from immunized mice effectively blocked SEB-induced T-cell proliferation in vitro. Naive mice survived a lethal SEB challenge when given pooled antisera 1, 2, or 4 h later, whereas the antisera failed to protect animals when administered 6 or 8 h after the toxin. Lethality at the later times was consistent with increased serum levels of TNF observed 6 h after SEB injection. These studies suggest that the N23K and F44S mutant proteins of SEB are less biologically active than the wild-type toxin, yet retain epitopes useful for eliciting a protective antibody response.