Immunoinformatics design and in vivo evaluation of a multiepitope vaccine targeting OMPL1, Lipl32, Lipl41, and Lipl46 for leptospirosis in a male ICR mouse model.

Leptospirosis is the most widespread and under-recognized zoonotic disease with around 1.03 million cases and ∼ 60,000 reported fatalities occurring each year. It is a threat to humans as well as animals and can only be avoided by appropriate immunization. In this study, epitopes from proteins present on the exterior membrane of selected pathogenic Leptospira species were used as immunogens viz. OMPL1, LipL32, LipL41 and LipL46. Conserved epitopes from each of the proteins across 8 species of Leptospira were screened and used to generate a multi-epitope vaccine that activates B cells, CD4+ and CD8+ cells. Six vaccine constructs were analyzed employing various immunoinformatics tools for physicochemical properties, structure prediction followed by validation, docking with immune receptors and molecular dynamics simulation. The vaccine construct 4 (LH) was found to comply with all the desired parameters to further consider for in vivo efficacy evaluation. For in vivo validation, gene for LH was cloned in a pVAX1 vector to be used as a DNA vaccine and in pET30a vector to express protein vaccine. DNA and protein vaccines were administered intramuscularly in ICR male mice along with adjuvant alhydrogel. Both types of vaccines elicited strong immune response evidenced from significantly increased serum IgG level evaluated by ELISA post the duration of 14 to 42 days. IFN-γ cytokine producing T cells were significantly stimulated in protein vaccine as revealed by flow cytometry suggesting the priming of CD4+ and CD8+ T cells by vaccine. The neutralizing antibody response to Leptospira serovars was confirmed with microscopic agglutination test. In summary, this study illustrates the prospective of multi-epitope DNA and protein vaccines incorporating epitopes from four outer membrane proteins to generate a strong immune response, paving the way forward for protection during challenge study against virulent Leptospira pathogens in animal model.