Assessment of efficacy of bifunctional entities as vaccine candidates to combat typhoidal salmonellosis

Abstract

Present study has explored the effect of dual immunization with two different antigenic entities (i.e. flagellin epitope and cytolethal distending toxin subunit B (CdtB) protein) on the protective response against lethal challenge of two typhoidal serovars, i.e. Salmonella enterica serovar Typhi and Salmonella enterica serovar Paratyphi A. Flagellin epitope , present in both the antigenic phases of flagellin protein of typhoidal serovars were identified bioinformatically, synthesized and then conjugated to Bovine serum albumin. CdtB protein was purified using Nickel affinity chromatography. After confirming the in-vitro immunogenicity of flagellin epitope-BSA conjugate and CdtB protein using Indirect ELISA with typhoid positive patients sera, a combination of both entities was administered intraperitoneally in mice followed by lethal challenge of typhoidal Salmonellae. Active immunization studies using the combination of both entities demonstrated improved mice survival after lethal challenge with typhoidal Salmonellae, reduced bacterial burden in organs, expression of immunophenotypic markers in splenocytes and restored tissue histoarchitecture. When used in combination, the effective doses of both the candidates reduced which may be attributed to multiprong approach used by the immune system to recognize Salmonella. Serum samples obtained from the immunized mice were used for passive immunization studies which further determined the protective efficacy of generated antibodies by different mechanisms such as complement mediated bactericidal action, swarming inhibition and increased phagocytic uptake. Overall, the present study is the first phase of the proof-of-concept which may prove to be beneficial in developing an effective bi-functional vaccine candidate to render protection against both Vi-positive as well as Vi-negative Salmonella strains. newline