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Fibronectin-binding protein A (FnBPA) is a key adhesin of Staphylococcus aureus, and the protein binding to fibrinogen and elastin is mediated by its N-terminal A domain. Thus, FnBPA-A has been considered a potential vaccine candidate, but the relevant epitopes are not fully understood. Here, purified rabbit anti-FnBPA-A antibodies were produced and used to screen for peptides corresponding to or mimicking the epitope of native FnBPA-A protein by using a phage random 12-mer peptide library. After four rounds of panning, 25 randomly selected phage clones were detected by phage-ELISA and competition-inhibition ELISA. Then, eight anti-rFnBPA-A antibody-binding phage clones were selected for sequencing, and six different 12-mer peptides were displayed by these phages. Although these displayed peptides shared no more than three consecutive amino acid residues identical to the sequence of FnBPA-A, they could be recognized by the FnBPA-A-specific antibodies in vitro and could induce specific antibodies against FnBPA-A in vivo, suggesting that these displayed peptides were mimotopes of FnBPA-A. Finally, the protective efficiencies of these mimotopes were investigated by mouse vaccination and challenge experiments. Compared with that of control group mice, the relative percent survival of mice immunized with phage clones displaying a mimotope was 13.33% (C2 or C15), 0% (C8), 6.67% (C10), 26.67% (C19 or 1:2 mixture of C23 and C19), 53.33% (C23), 33.33% (1:1 mixture of C23 and C19), and 66.67% (2:1 mixture of C23 and C19). Overall, five peptides mimicking FnBPA-A protein epitopes were obtained, and a partially protective immunity against S. aureus infection could be stimulated by these mimotope peptides in mice., (© 2019 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)

Background: Infections caused by Staphylococcus aureus continue to plague surgical patients, whether as surgical site infections or other nosocomial infections that complicate surgical care. The only meaningful methods available to decrease the risk of developing such infections are topical skin antisepsis (pre-operative skin preparation) and peri-operative antibiotic prophylaxis, neither of which offer a panacea. Alternatives to the latter are sought so as to minimize antibiotic selection pressure as a factor in the increasing problem of antimicrobial drug resistance. This review considers the possibility that immunization against S. aureus may offer a viable alternative for prophylaxis. Methods: Review and synthesis of pertinent English-language medical literature. Results: Vaccination against viral pathogens has been in successful clinical use for more than two centuries and was instrumental in the eradication of smallpox and the near-elimination of diseases such as poliomyelitis. Vaccinations against a limited number of bacterial pathogens (e.g., Bordetella pertussis , Clostridium tetanii , Corynebacterium diphtheriae , Haemophilus influenzae type b, Neisseria meningiditis , Streptococcus pneumoniae ) have also been introduced with success, whereas others against bacteria are in development ( C. difficile , Pseudomonas aeruginosa , S. aureus ). Vaccination against S. aureus infection is in current veterinary use (e.g., to prevent mastitis among dairy cattle) but has not been successful to date in human beings despite multiple attempts, although development continues. Conclusions: Because of its complex microbiology, including multiple virulence factors and the ability to evade host immune surveillance, S. aureus presents numerous antigenic targets for vaccine development. Failure of two prior single-antigen vaccines in clinical trials has led to the consensus that future vaccine candidates must be directed against multiple antigens. Two distinct four-antigen vaccines are in clinical trials, but efficacy is yet to be determined.

Staphylococcus aureus infections are becoming a major global health issue due to the rapid emergence of multidrug-resistant strains. Therefore, there is an urgent need to develop an effective vaccine to prevent and control these infections. In order to develop a universal immunization strategy, we constructed a mutant derivative of S. aureus 132 which lacks the genes involved in D-alanine biosynthesis, a structural component of cell wall peptidoglycan. This unmarked deletion mutant requires the exogenous addition of D-alanine for in vitro growth. The aim of this study was to examine the ability of this D-alanine auxotroph to induce protective immunity against staphylococcal infection. Our findings demonstrate that this deletion mutant is highly attenuated, elicits a protective immune response in mice and generates cross-reactive antibodies. Moreover, the D-alanine auxotroph was completely eliminated from the blood of mice after its intravenous or intraperitoneal injection. We determined that the protective effect was dependent on antibody production since the adoptive transfer of immune serum into naïve mice resulted in effective protection against S. aureus bacteremia. In addition, splenocytes from mice immunized with the D-alanine auxotroph vaccine showed specific production of IL-17A after ex vivo stimulation. We conclude that this D-alanine auxotroph protects mice efficiently against virulent staphylococcal strains through the combined action of antibodies and IL-17A, and therefore constitutes a promising vaccine candidate against staphylococcal disease, for which no licensed vaccine is available yet.

The cytokine storm induced by staphylococcal enterotoxin B (SEB) describes the rapid and dramatic induction of mediators which are likely responsible for the toxin's deleterious effects. However despite the use of numerous animal models for investigating SEB related illness in humans, mechanisms of toxicity and correlates of protection remain unclear. In the present study, we used an LPS-potentiated model of SEB lethality to investigate the toxin-induced cytokine and chemokine responses in untreated and immunized mice. Of 30 separate mediators analyzed, serum levels for 28 or 27 of these cytokines and chemokines were elevated following administration of dosages of 3 or 30 LD 50 of native SEB, respectively. Mice immunized with a non-toxic SEB vaccine candidate expressed in either E. coli or transgenic soy expression systems were protected from lethality when challenged with potentiated SEB. The majority of SEB-induced cytokines and chemokines (21 of 28 or 23 of 27 following challenge with dosages of 3 or 30 LD 50 of native SEB, respectively) were significantly decreased in mice immunized with an SEB vaccine candidate when compared to control animals. Together, these studies provide the most comprehensive evaluation of the cytokine storm induced in this LPS-potentiated model of SEB lethality to date. As with other animal models, the identification of those mediators which are necessary and sufficient for SEB-induced toxicity remains unclear., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Staphylococcus aureus is both a commensal organism and also an important opportunistic human pathogen, causing a variety of community and hospital-associated pathologies, such as bacteremia-sepsis, endocarditis, pneumonia, osteomyelitis, arthritis and skin diseases. The resurgence of S. aureus during the last decade in many settings has been facilitated not only by bacterial antibiotic resistance mechanisms but also by the emergence of new S. aureus clonal types with increased expression of virulence factors and the capacity to neutralize the host immune response. Prevention of the spread of S. aureus infection relies on the use of contact precautions and adequate procedures for infection control that so far have not been fully effective. Prevention using a prophylactic vaccine would complement these processes, having the potential to bring additional, significant progress toward decreasing invasive disease due to S. aureus.

In a recent article, X. Li et al. [mBio 7(1):e02232-15, 2016, http://dx.doi.org/10.1128/mBio.02232-15] investigate the utility of a vaccine composed of the Staphylococcus aureus protein clumping factor A (ClfA) in protecting mice from S. aureus infection. ClfA, one of the first proteins to be identified as a potential vaccine antigen for S. aureus prophylaxis, is currently a component of several investigational vaccines. The authors conclude that ClfA may not be effective for S. aureus prophylaxis. In contrast, previously published papers reporting positive data suggested that ClfA was potentially an important vaccine target to prevent invasive S. aureus disease. This commentary addresses the observed differences between the findings of Li et al. and those from other publications, highlighting the importance for preclinical vaccine antigen assessments to reflect the biological role of said antigen in virulence and, consequently, the importance of choosing appropriate preclinical disease models to test such antigens., (Copyright © 2016 Anderson et al.)

Staphylococcus aureus is a Gram-positive pathogen that causes a wide range of infections in humans and animals. Bacterial ghosts are nonliving, empty cell envelopes and are well represented as novel vaccine candidates. In this study, we examined the immunogenicity and protective efficacy of S. aureus ghosts (SAGs) against a virulent challenge in rats. Nonliving SAGs were generated by using the MIC of sodium hydroxide. The formation of a transmembrane lysis tunnel structure in SAGs was visualized by scanning electron microscopy. To investigate these SAGs as a vaccine candidate, rats were divided into four groups, A (nonimmunized control), B (orally immunized), C (subcutaneously immunized), and D (intravenously immunized). The IgG antibody responses were significantly stronger in the SAG-immunized groups than in the nonimmunized control group (P < 0.05). Moreover, a significant increase in the populations of CD4(+) and CD8(+) T cells was observed in all three immunized groups (P < 0.05). We also found that serum bactericidal antibodies were significantly elicited in the SAG-immunized groups (P < 0.05). Most importantly, the bacterial loads in the immunized groups were significantly lower than those in the nonimmunized control group (P < 0.01). These results suggest that immunization with SAGs induces immune responses and provides protection against a virulent S. aureus challenge., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Staphylococcal disease represents a universal burden including acute, life-threatening infections as well as chronic infections usually associated with foreign materials. Infections occur notably in permanent carriers of Staphylococcus aureus. To date, all the attempts to develop an efficacious vaccine against S. aureus have failed. Failures in vaccine clinical trials might be related to a focus on single targets and development of humoral-based vaccines rather than vaccines with a combination of antigens stimulating both humoral and cellular immunity. The end points of these unsuccessful trials were a reduction in mortality or bacteremia, whereas the patient's decolonization was not assessed. Adopting the latter point of view, the aim of this article is to discuss nasal mucosal decolonization as a complementary marker of vaccine efficacy for clinical research in vaccine development.

In an effort to develop a sustainable platform for manufacturing protein-based vaccine candidates, we expressed a triple mutant of staphylococcal enterotoxin B carrying the L45R, Y89A, and Y94A modifications in transgenic soybean seeds (soy-mSEB). Soy-mSEB possessed no detectable superantigen activity in vitro. We found that this soybean-derived, nontoxic mutant of SEB could be stably expressed, stored in seeds for extended periods at room temperature without degradation, and easily purified from contaminating soy proteins. Vaccination of pigs with purified soy-mSEB, or the identical triple mutant expressed in Escherichia coli (E. coli-mSEB), resulted in high antibody titers against the native toxin in immunized animals. In fact, titers were indistinguishable regardless of the immunogen used, demonstrating the equivalence of soy-mSEB and E. coli-mSEB vaccinations. Antisera from either immunized group were able to block native SEB superantigen activity in an in vitro neutralization assay. Similar results were obtained when immunized animals were challenged with a sublethal dose of native toxin. Significant reductions in toxin-induced serum cytokine levels were observed in soy-mSEB- and E. coli-mSEB-immunized pigs compared to control animals. The reductions in SEB-induced cytokine responses were similar regardless of the immunogen used for vaccination. Surprisingly, however, some clinical symptoms, such as prostration, lethargy, emesis, and/or diarrhea, were still observed in all immunized animals. These studies demonstrate the potential for soybean-derived proteins as a platform technology for sustainable vaccine manufacturing and the usefulness of a sublethal challenge model in pigs for evaluating the efficacy of potential SEB vaccine candidates.

Aim: Assessment of characteristics of antigenic complexes of Staphylococcus aureus vaccine strains in different cultivation conditions., Materials and Methods: S. aureus vaccine strains (No. 5, 9, 1986, 1991) were grown in liquid nutrient media--full value and semi-synthetic--as well as on solid medium. Reactor cultivation was performed in the fermenter ANKUM-2M. Complex of antigens were obtained by water extraction method applied to staphylococcal biomass inactivated with acetone and assessed by common methods on protein and carbohydrate content; specific activity was assessed by minimal inhibitory dose in passive hemagglutination inhibition assay. Study of acute toxicity was performed on outbred mice., Results: Using strain no. 1991, model of reactor cultivation in full value medium with separation of biomass by microfiltration was validated on the basis of biomass and semiproduct of antigenic complex (acetone powder) yield as well as productivity of biomass cumulation. Study of antigenic complexes obtained from biomass of 4 strains during reactor cultivation compared with complexes extracted from cultures grown on solid medium revealed increased protein and decreased carbohydrate content but similar specific activity. It was demonstrated that complex of antigens obtained from cultures grown either by reactor cultivation or on solid medium were non-toxic., Conclusion: New technology for manufacturing staphylococcal complex of antigens with reactor cultivation of vaccine strains in full value medium with subsequent purification of antigenic complex from the biomass by microfiltration was developed. Results of the study demonstrated the usefulness of the developed technology for both further studies on a cellular staphylococcal vaccine and manufacture of staphylococcal component of "Immunovac" vaccine.

Methicillin-resistant Staphylococcus aureus (MRSA) has posed an immense problem for clinicians in the hospital setting for years, emerging as the most frequent nosocomial infection. To deal with this problem pathogen and others, infectious disease specialists have developed a variety of procedures for their control and prevention, involving options from preventative measures such as decolonization and isolation of MRSA-confirmed patients, to the more simple procedures of hand washing, expanding glove use, and reducing time in the hospital. With the realization that MRSA is now a community problem, there are expanded efforts toward more direct intervention, such as the use of anti-MRSA antibacterials and vaccines, in an attempt to reduce the overall burden of MRSA.

An in vitro protein selection method, ribosome display, has been applied to comprehensively identify and map the immunologically relevant proteins of the human pathogen Staphylococcus aureus. A library built up from genomic fragments of the virulent S. aureus COL strain (methicillin-resistant S. aureus) allowed us to screen all possible encoded peptides for immunoreactivity. As selective agents, human sera exhibiting a high antibody titer and opsonic activity against S. aureus were used, since these antibodies indicate the in vivo expression and immunoreactivity of the corresponding proteins. Identified clones cluster in distinct regions of 75 genes, most of them classifiable as secreted or surface-localized proteins, including previously identified virulence factors. In addition, 14 putative novel short open reading frames were identified and their immunoreactivity and in vivo mRNA expression were confirmed, underscoring the annotation-independent, true genomic nature of our approach. Evidence is provided that a large fraction of the identified peptides cannot be expressed in an in vivo-based surface display system. Thus, in vitro protein selection, not biased by the context of living entities, allows screening of genomic expression libraries with a large number of different ligands simultaneously. It is a powerful approach for fingerprinting the repertoire of immune reactive proteins serving as target candidates for active and passive vaccination against pathogens.

An alum-precipitated Staphylococcus aureus vaccine, composed of a formalin-inactivated whole culture of a strain which produces Smith surface antigen and combined with the whole culture of a highly toxigenic strain, was found to afford a good immunity to staphylococcal skin infection in rabbits. Three injections of the vaccine provided immunity which lasted for at least 6 months against a primarily pyogenic strain of S. aureus and for at least 3 months against a toxigenic strain. From experiments using vaccines prepared from cells or toxoid only, it was deduced that, although there is a measure of strain specific immunity, a good heterologous immunity can be established with a combined product provided that it contains adequate quantities of toxoid. The use of such a vaccine as a potential aid in the control of bovine staphylococcal mastitis is discussed.