Immunogenicity Of Recombinant BCG RBCG :: Ag85B - ESAT6 - Rv2608 And RBCG :: Ag85B - ESAT6 - Rv3620c

Tuberculosis (TB), as a serious infection disease, brings a long-term harm to human health, and it is estimated over one-third of the world’s population is infected with M. tuberculosis and almost3million people die from this disease annually. Moreover, the morbidity and mortality of TB patients have been increasing remarkably in recent decades due to TB and HIV co-infection and the emergence of drug-resistant TB, which pose new urgency to global TB control. UN has ranked tuberculosis as one of the three main diseases that greatly affect human’s health in the21st Century. Bacillus Calmette-Guerin (BCG) is the only available globally-used vaccine against TB currently, while some studies have shown it provides variable efficacy ranging from0to80%, especially confers almost no protection among adults. Therefore, developing new TB vaccines is a high worldwide priority.There are two main vaccination strategies, pre-exposure vaccination and postexposure vaccination. The former appeared earlier and has developed various kinds of vaccine candidates, including DNA vaccines, subunit peptide vaccines, recombinant BCG, auxotrophic mutants of BCG and virus-based vaccines. This study focuses on recombinant BCG (rBCG) with the hope to induce stronger immunogenicity than parental BCG rBCG is a class of vaccines that utilize BCG as a host organism to express genes encoding immunodominant antigens or immunostimulatory cytokines.It has been accepted that both Ag85B and ESAT-6are importantly immunodominant antigens of M. tuberculosis by the scientists all over the world. Previous studies showed that rBCG expressing the Ag85B-ESAT6fusion protein increased the protective efficacy of BCG in a mouse model of tuberculosis. Furthermore, the multigenic vaccine approach is attractive due to broad immune responses generated by simultaneously targeting several antigens. So we constructed two rBCG strains expressing Ag85B, ESAT-6, Rv2608(PPE42) and Rv3620c (EsxW), rBCG::Ag85B-ESAT6-Rv2608and rBCG::Ag85B-ESAT6-Rv3620c, within the shuttle plasmid pMV261. After vaccinating C57BL/6mice with the rBCG strains and the parental BCG separately, we studied the level of antibody response, IgG2b/IgGl and the amount of cytokines secreted, including IFN-γ, IL-2and IL-10by ELISA, calculated the number of splenocytes secreting TNF-a by ELISPOT, and detected CD4+/CD8+T cells by flow cytometry. It was found that rBCG::Ag85B-ESAT6-Rv2608or rBCG::Ag85B-ESAT6-Rv3620c induced stronger cell-mediated immune response than BCG in mice by increasing TNF-a and IL-2secrection and decreasing IL-10secrection, while rBCG::Ag85B-ESAT6-Rv3620c also induced a higher production of IFN-y. Either rBCG::Ag85B-ESAT6-Rv2608or rBCG::Ag85B-ESAT6-Rv3620c improved T-cell differentiation into CD4+T cells or CD8+T cells and enhanced specific cytotoxic T lymphocyte (CTL) response. Additionally, it was found that rBCG::Ag85B-ESAT6-Rv2608or rBCG::Ag85B-ESAT6-Rv3620c induced stronger humoral. These results suggest that the two rBCG strains are both promising candidates to control tuberculosis, and pave the way for further studies.