Immunogenicity and protective efficacy of Brucella abortus recombinant protein cocktail (rOmp19 + rP39) against B. abortus 544 and B. melitensis 16M infection in murine model

• Intraperitoneal immunization with protein cocktail rO + rP elicited IgG2a, IFN-γ, IL-2 and IL-12 greater than individual proteins.
• The rO + rP immunogen has significantly reduced the bacterial burden in spleen and liver at 8th week post challenge.
• The present study reveals that the multivalent cocktail protein consisting Omp19 and P39 can render better protection than Omp19 and P39 alone against brucellosis.

In this study, the immunogenicity and protective efficacy of recombinant proteins Omp19 (rO) and P39 (rP) from Brucella abortus were evaluated individually and compared with the cocktail protein (rO + rP) against B. abortus 544 and Brucella melitensis 16M infection in BALB/c mouse model. Intra-peritoneal (I.P.) immunization with rO + rP cocktail developed substantially higher antibody titers predominant with Th1 mediated isotypes (IgG2a/2b). Western blot analysis using anti-rO + rP antibodies showed specific reactivity with native Omp19 (19 kDa) and P39 (39 kDa) among whole cell proteins of B. abortus and B. melitensis. Splenocytes extracted from rO + rP immunized mice induced significantly (P < 0.001) higher proliferative responses at 30 μg/ml with considerable expression of pro-inflammatory cytokines (IFN-γ, IL-2 and IL-12) than rO and rP. Macrophage cell (RAW 264.7) monolayer supplemented with anti-rO + rP polysera exhibited enhanced viability against challenge with B. abortus 544 (72.27%) and B. melitensis 16 M (68.57%). On the other hand, individual anti-rO and anti-rP polysera resulted in relatively lesser protection against the pathogens (64.79%, 54.45% and 47.13%, 45.11%, respectively). Immunized group of mice when I.P. challenged with 5 × 104 CFU of B. abortus 544 and B. melitensis 16M were found significantly (P < 0.001) protected in the rO + rP group (log units of protection, spleen: 2.38, 2.12; liver: 1.04, 0.81, respectively) than in rO (spleen: 1.43, 1.21; liver: 0.7, 0.47) and rP (spleen: 1.24, 1.17; liver: 0.65, 0.34). Findings from this study depicted that rO + rP cocktail is highly immunogenic with the Th1 predominant serum antibody titers and T-cell mediated immune protection, would be a valuable intervention in the development of a safer and improved Brucella vaccine.