| BackgroundsBrucellosis is caused by Brucella, which is a kind of B class infectious diseases of zoonoses.Brucellosis can make ones get fever and other series of reaction after infected. The infertility and abortion of maternal livertock due to Brucellosis.Brucellosis is not easy to cure so that vaccine to brucellosis is more significance.Now.there are three kinds of brucellosis vaccines:first of all.live attenuated vaccine,is the most widely used in preventing Brucellosis.Because of its large side effect.which become to a security problem.Secondly, Inactivated vaccine can only provide a temporary protection. However. The component of subunits vaccine is clear, meanwhile,it is easy to product and have ittle side prospects in application.Yet,there is a few immunogenicity antigens have been recognized.which limits the development of subunits vaccine of brucellosis.ObjectivesIn this study, we used the theory of reverse vaccine to find immunogenicity antigens of brucellosis, including secretes proteins, poison proteins and outer membrane proteins. which were related with intracellular survival and protection of brucellosis. The number of the protein was 108.In order to obtain a certain concentration of protein, the protein genes was expressed and purificated in E.coli in certain condition.MethodsWe have stimulated splenocytes of mice with the protein in vitro; whicih had been immunizated with S19:live attenuated vaccine.Meanwhile; there is a group immunizated with PBS as negative control group, which were stimulated by the 103proteins in vitro in the same condition. The clear culture supernatants were collected. IFN-γwas determined by using an ELISA Quantikine Mouse kit (R&D Systems).If IFN-γin S19 were twice more than the negative group,it can induceimmunodominant T-cell response.There were 10 proteins could induce immunodominant T-cell response. To test the possible protection roles of 7 candidate proteins, the immunized mice was challenged with virulent strain B. abortus 544, and the significant reduction in the number of bacteria in spleen represent the protection efficiency. Data from protection experiments demonstrated that 4 candidate proteins immunizated with proteins plus adjuvant resulted in significantly higher degree of protection than the controls that received PBS plus adjuvant. These results indicated that BMEI0357, BMEII1048, BMEI0705 and BMEI1692 in adjuvant could confer a significant degree of protection against Brucella infection.The antibody responses were analyzed in the sera of the immunized mice at selected intervals post-immunization to monitor total IgG level by ELISA. We examined the proportion of Thl-associated IgG2a and Th2-associated IgG1 in the total serum IgG with ELISAs. These results indicate that both of them can induce humoral responses. Mice immunized with them plus adjuvants were prone to producing a Thl-type of immune response.in addition to BMEI1692.ResultsIn order to further analyze the cell-mediated immunity of the candidate protein, we investigated IFN-γsecretion by the splenocytes of immunized mice after stimulation with purified protein, complete medium or ConA as negative and positive controls, respectively. The results indicated immunization of BMEII1048. and BMEI0705 plus adjuvant could induce cellular immune response.ConclusionsAbove all, we had found two immunodominant antigens of brucellosis, whicih could induce humoral responses and cellular immune response. BMEII1048, and BMEI0705 had better immune protection effect, whicih could use for subunit vaccine of brucellosis. |
PDF Full Text Request