Immune response characterization and vaccine potential of a recombinant chimera comprising B-cell epitope of Aeromonas hydrophila outer membrane protein C and LTB

Abstract

Aeromonas hydrophila is one of the most virulent fish pathogens, causing colossal economic losses to the aquaculture industry annually. The absence of a safe and effective vaccine makes it very difficult to control this infection. Outer membrane proteins have been widely illustrated to confer protective immunity against a broad spectrum of gram negative bacteria. In the current study, we have analyzed the ability of B-cell epitopes of A. hydrophila's outer membrane protein C (OmpC) to confer protection against bacterial virulence. Bioinformatic algorithms were used to predict linear B-cell epitopes of OmpC and the corresponding nucleotide sequences were cloned in translational fusion with heat labile enterotoxin B subunit (LTB) of E. coli. Of the three recombinant LTB.epitope fusion proteins evaluated, antisera against the fusion protein comprising the epitope stretch of 143-175 amino acids gave maximum cross reactivity with the parent protein OmpC. The anti-fusion protein antisera contained both OmpC- and LTB-specific antibodies. The fusion proteins' LTB moiety retained its ability to bind to the GM₁ ganglioside receptor, an essential requirement for its adjuvanicity. Antibody isotyping, cytokine ELISA, and cytokine array analysis revealed a Th2 skewed type immune response along with the presence of some relevant Th17 and Th1 cytokines involved in conferring protective immunity. Surface exposure of the epitope_143-175 on live A. hydrophila membrane was investigated and validated using bacterial agglutination and flow cytometry analysis using anti-fusion protein antisera. Our results strongly support the potential of B-cell epitope_143-175 of OmpC of A. hydrophila, in fusion with the LTB, as an effective and promising vaccine candidate against this bacterium.

Keywords: Aeromonas hydrophila; Agglutination; Cytokines; GM(1) ganglioside receptor; Heat-labile enterotoxin B subunit; Recombinant protein.