In silico prediction and experimental evaluation of LIP3228 of pathogenic Leptospira as a potential subunit vaccine target against leptospirosis.

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A protein subunit vaccine comprising conserved surface-exposed outer membrane proteins (SE-OMPs) is considered a promising platform for leptospirosis vaccine. The search for novel vaccine candidates that confer high protective efficacy against leptospirosis is ongoing. The LIP3228 protein was previously identified as a conserved and abundant SE-OMP with the potential to serve as an effective vaccine candidate. However, it is crucial to explore the immunological properties of this vaccine antigen before proceeding with animal experiments. This study aimed to assess the immunological characteristics of LIP3228 through in silico prediction and to validate its immunogenicity and protective efficacy in a hamster model of acute leptospirosis. The LIP3228 vaccine candidate was predicted in silico to be immunogenic, with strong binding B-cell and T-cell epitopes. In the immune simulator, it demonstrated stable interactions with Toll-like receptors 2 (TLR2) and 4 (TLR4) and induced immune responses, potentially stimulating host immune responses in vivo. The animal experiment showed that immunization with recombinant LIP3228 protein, formulated with AddaVax adjuvant, induced high and specific IgG responses in hamsters, with IgG2 being the predominant subclass. Although no significant improvement in survival was observed compared to the negative control after a homologous challenge with virulent leptospires, the vaccinated hamsters showed a reduction in histopathological changes and severity of lesions in target organs compared to unvaccinated hamsters. These results suggest that the immunoinformatic prediction is effective in predicting immunogenicity but not protective efficacy. Therefore, LIP3228 could be considered a potential vaccine candidate for mitigating severe tissue damage. These findings may have significant implications for the development of subunit vaccines in the future.