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The present study employs the  Brucella abortus  L7/L12 antigen in a  Salmonella  secretion platform and investigates its ability to induce protective immune responses against wild type challenge in BALB/c mice. The highly conserved L7/L12 open reading frame was PCR amplified from  B. abortus  and cloned into a prokaryotic expression vector, pJHL65, directly under the beta-lactamase secretory signal. The plasmid constructs pJHL65::L7/L12 was then transformed into an attenuated  Salmonella  Typhimurium strain, JOL1800 ( ∆lon ,  ∆cpxR ,  ∆asd , and  ∆rfaL ), and protein secretion was verified by Western blot. Three mice groups were inoculated with either phosphate-buffered saline (PBS), vector-only control, or the vaccine strain secreting L7/L12 antigen. Assessment of humoral and cell-mediated immune responses revealed successful elicitation of  Brucella  antigen-specific Th1 and Th2 immune responses that were significantly higher than PBS and vector control groups. The immune responses were confirmed by splenocyte proliferation assay, flow cytometry analysis for CD4+ and CD8+ markers, and RT-PCR based cytokine profiling upon restimulation with L7/L12 purified antigen. Results indicate that immunization with  Salmonella  secreting L7/L12 antigen demonstrated significant enhancement of cell-mediated immune (CMI) responses in immunized mice. The overall effectiveness of the immunization was evaluated by challenging with virulent  B. abortus  that revealed significant reduction in  Brucella  colonization in spleen and liver tissues in  Salmonella  L7/L12 immunized mice. Delivery of  Brucella  protective antigen L7/L12 using the  Salmonella  secretion system can effectively accomplish immunogenic advantages of both  Salmonella  and L7/L12 to derive robust CMI responses and induce humoral immunity to protect against  Brucella  infection in the mouse model.

veterinary research

Live vaccine consisting of attenuated Salmonella secreting and delivering Brucella ribosomal protein L7/L12 induces humoral and cellular immune responses and protects mice against virulent Brucella abortus 544 challenge