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Immunoinformatics analysis and evaluation of recombinant chimeric triple antigen toxoid (r-HAB) against Staphylococcus aureus toxaemia in mouse model.

ABSTRACT: Staphylococcus aureus is a serious pathogen unleashing its virulence through several classes of exotoxins such as hemolysins and enterotoxins. In this study, we designed a novel multi-antigen subunit vaccine which can induce innate, humoral and cellular immune responses. Alpha hemolysin, enterotoxins A and B were selected as protective antigens for combining into a triple antigen chimeric protein (HAB). Immunoinformatics analysis predicted HAB protein as a suitable vaccine candidate for inducing both humoral and cellular immune responses. Tertiary structure of the HAB protein was predicted and validated through computational approaches. Docking studies were performed between the HAB protein and mice TLR2 receptor. Furthermore, we constructed and generated recombinant HAB (r-HAB) protein in E. coli and studied its toxicity, immunogenicity and protective efficacy in a mouse model. Triple antigen chimeric protein (r-HAB) was found to be highly immunogenic in mouse as the anti-r-HAB hyperimmune serum was strongly reactive to all three native exotoxins on Western blot. In vitro toxin neutralization assay using anti-r-HAB antibodies demonstrated > 75% neutralization of toxins on RAW 264.7 cell line. Active immunization with r-HAB toxoid gave ~ 83% protection against 2 × lethal dosage of secreted exotoxins. The protection was mediated by induction of strong antibody responses that neutralized the toxins. Passive immunization with anti-r-HAB antibodies gave ~ 50% protection from lethal challenge. In conclusion, in vitro and in vivo testing of r-HAB found the molecule to be nontoxic, highly immunogenic and induced excellent protection towards native toxins in actively immunized and partial protection to passively immunized mice groups. KEY POINTS: • HAB protein was computationally designed to induce humoral and cellular responses. • r-HAB protein was found to be nontoxic, immunogenic and protective in mouse model. • r-HAB conferred protection against lethal challenge in active and passive immunization.

SUBMITTER: Kota RK

PROVIDER: S-EPMC8490849 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundTSST-1 is a secretory and pyrogenic superantigen that is being responsible for staphylococcal mediated food poisoning and associated clinical manifestations. It is one of the main targets for the construction of vaccine candidates against Staphylococcus aureus. Most of the vaccines have met failure due to adverse reactions and toxicity reported during late clinical studies. To overcome this, an immunoinformatics approach is being used in the present study for the design of a multi-epitope vaccine to circumvent the problems related to toxicity and allergenicity.ResultsIn this study, a multi-epitope vaccine against Staphylococcus aureus targeting TSST-1 was designed through an immunoinformatics approach. B cell and T cell epitopes were predicted in silico and mapped with linkers to avoid junctional immunogenicity and to ensure the efficient presentation of exposed epitopes through HLA. β-defensin and PADRE were adjusted at the N-terminal end of the final vaccine as adjuvants. Physiochemical parameters, antigenicity, and allergenicity of the vaccine construct were determined with the help of online servers. The three-dimensional structure of the vaccine protein was predicted and validated with various tools. The affinity of the vaccine with TLR-3 was studied through molecular docking studies and the interactions of two proteins were visualized using LigPlot+. The vaccine was successfully cloned in silico into pET-28a (+) for efficient expression in E. coli K12 system. Population coverage analysis had shown that the vaccine construct can cover 83.15% of the global population. Immune simulation studies showed an increase in the antibody levels, IL-2, IFN-γ, TGF-β, B cell, and T cell populations and induced primary, secondary, and tertiary immune responses.ConclusionMulti-epitope vaccine designed through a computational approach is a non-allergic and non-toxic antigen. Preliminary in silico reports have shown that this vaccine could elicit both B cell and T cell responses in the host as desired.

Project description:Staphylococcus aureus is a leading cause of significant morbidity and mortality and an enormous economic burden to public health worldwide. Infections caused by methicillin-resistant S. aureus (MRSA) pose a major threat as MRSA strains are becoming increasingly prevalent and multi-drug resistant. To this date, vaccines targeting surface-bound antigens demonstrated promising results in preclinical testing but have failed in clinical trials. S. aureus pathogenesis is in large part driven by immune destructive and immune modulating toxins and thus represent promising vaccine targets. Hence, the objective of this study was to evaluate the safety and immunogenicity of a staphylococcal 4-component vaccine targeting secreted bi-component pore-forming toxins (BCPFTs) and superantigens (SAgs) in non-human primates (NHPs). The 4-component vaccine proved to be safe, even when repeated vaccinations were given at a dose that is 5 to 10- fold higher than the proposed human dose. Vaccinated rhesus macaques did not exhibit clinical signs, weight loss, or changes in hematology or serum chemistry parameters related to the administration of the vaccine. No acute, vaccine-related elevation of serum cytokine levels was observed after vaccine administration, confirming the toxoid components lacked superantigenicity. Immunized animals demonstrated high level of toxin-specific total and neutralizing antibodies toward target antigens of the 4-component vaccine as well as cross-neutralizing activity toward staphylococcal BCPFTs and SAgs that are not direct targets of the vaccine. Cross-neutralization was also observed toward the heterologous streptococcal pyogenic exotoxin B. Ex vivo stimulation of PBMCs with individual vaccine components demonstrated an overall increase in several T cell cytokines measured in supernatants. Immunophenotyping of CD4 T cells ex vivo showed an increase in Ag-specific polyfunctional CD4 T cells in response to antigen stimulation. Taken together, we demonstrate that the 4-component vaccine is well-tolerated and immunogenic in NHPs generating both humoral and cellular immune responses. Targeting secreted toxin antigens could be the next-generation vaccine approach for staphylococcal vaccines if also proven to provide efficacy in humans.

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Immunoinformatics analysis and evaluation of recombinant chimeric triple antigen toxoid (r-HAB) against Staphylococcus aureus toxaemia in mouse model.

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