ABSTRACT
Brucella spp. are intracellular bacteria that cause the most frequent zoonosis in the world. Although recent work has advanced the field of Brucella vaccine development, there remains no safe human vaccine. In order to produce a safe and effective human vaccine, the immune response to Brucella spp. requires greater understanding. Induction of Brucella-specific CD8+ T cells is considered an important aspect of the host response; however, the CD8+ T-cell response is not clearly defined. Discovering the epitope containing antigens recognized by Brucella-specific CD8+ T cells and correlating them with microarray data will aid in determining proteins critical for vaccine development that cover a kinetic continuum during infection. Developing tools to take advantage of the BALB/c mouse model of Brucella melitensis infection will help to clarify the correlates of immunity and improve the efficacy of this model. Two H-2(d) CD8+ T-cell epitopes have been characterized, and a group of immunogenic proteins have provoked gamma interferon production by CD8+ T cells. RYCINSASL and NGSSSMATV induced cognate CD8+ T cells after peptide immunization that showed specific killing in vivo. Importantly, we found by microarray analysis that the genes encoding these epitopes are differentially expressed following macrophage infection, further emphasizing that these discordant genes may play an important role in the pathogenesis of B. melitensis infection.
Subject(s)
Antigens, Bacterial/metabolism , Brucella melitensis/physiology , Brucellosis/immunology , CD8-Positive T-Lymphocytes/physiology , Animals , Antibodies, Bacterial/blood , Antibodies, Bacterial/immunology , Antibody Affinity , Epitopes , Female , Gene Expression Profiling , Gene Expression Regulation/immunology , Major Histocompatibility Complex , Mice , Mice, Inbred BALB C , Protein BindingABSTRACT
BACKGROUND: There is no safe, effective human vaccine against brucellosis. Live attenuated Brucella strains are widely used to vaccinate animals. However these live Brucella vaccines can cause disease and are unsafe for humans. Killed Brucella or subunit vaccines are not effective in eliciting long term protection. In this study, we evaluate an approach using a live, non-pathogenic bacteria (E. coli) genetically engineered to mimic the brucellae pathway of infection and present antigens for an appropriate cytolitic T cell response. METHODS: E. coli was modified to express invasin of Yersinia and listerialysin O (LLO) of Listeria to impart the necessary infectivity and antigen releasing traits of the intracellular pathogen, Brucella. This modified E. coli was considered our vaccine delivery system and was engineered to express Green Fluorescent Protein (GFP) or Brucella antigens for in vitro and in vivo immunological studies including cytokine profiling and cytotoxicity assays. RESULTS: The E. coli vaccine vector was able to infect all cells tested and efficiently deliver therapeutics to the host cell. Using GFP as antigen, we demonstrate that the E. coli vaccine vector elicits a Th1 cytokine profile in both primary and secondary immune responses. Additionally, using this vector to deliver a Brucella antigen, we demonstrate the ability of the E. coli vaccine vector to induce specific Cytotoxic T Lymphocytes (CTLs). CONCLUSION: Protection against most intracellular bacterial pathogens can be obtained mostly through cell mediated immunity. Data presented here suggest modified E. coli can be used as a vaccine vector for delivery of antigens and therapeutics mimicking the infection of the pathogen and inducing cell mediated immunity to that pathogen.
