Protective Efficacy Of Mucosal Immunuization With Classical Swine Fever Virus Vaccine

Classical swine fever virus (CSFV) can infect animals through mucosa-alimentary tract and respiratory tract, therefore, preventing infection from local mucosal invision by mucosal immunization is possible. After mucosal immunization, the antigen can bind the receptor and block infection of wild virus, eliciting better immunoprophylaxis effect. In this study, we investigated the feasibility of mucosal immunization of CSFV vaccine along with mucosa adjuvant based on existent CSFV vaccine.The low toxical double-mutants LTKG and LTRG were obtained by fermentation of recombination strains. The two recombinant proteins were purified by affinity ehromatography and the concentrations were determined to be750μg/ml and660μg/ml by BCA kit, respectively.CSFV vaccine along with LTKG or LTRG were injected intranasally (i.n.), intragastric administration (i.g.) or peroral administration (p.o.), and intramuscular inoculation (i.m.), into mice every10-15days, respectively. Sera, nasal elution and stool were collected after the third immunization. The IgG in the serum and IgA of local mucosa antibody reponses were detected by indirect ELISA. For IgG in the serum, the results showed that there were no significant difference (P>0.05) between i.n. group and i.g. group, but the titers of them were lower than i.m. group before the third immunization. After the third immunization, IgG titers of the mice immunized with CSFV vaccine+LTRG through i.n. were the highest, and significantly higher (0.01<P<0.05) than mice only immunized with CSFV vaccine via i.n.. The IgA antibody of mucosa was tested in every group of mice. It had significant difference (P<0.01) between i.g. groups and i.n. groups immunized with CSFV vaccine+LTKG or CSFV+LTRG after the third immunization. Moreover, i.g. groups of mice immunizede with CSFV vaccine+LTKG and CSFV+LTRG developed higher (0.01<P<0.05) IgA antibody titers than i.m. groups. The results suggested that mice immunized CSFV vaccine along with LTRG, LTKG elicit the best local mucosal immunization effect. CSFV vaccine along with LTKG or LTRG were injected intranasally (i.n.), intragastric administration (i.g.) or peroral administration (p.o.), and intramuscular inoculation (i.m.), into pigs at days1,16and31, respectively. Sera, nasal elution and stool were collected after the third immunization. The IgG in the serum and IgA of local mucosa antibody reponses were examed by indirect ELISA. The titers of IgG antibody in the Sera were always high during the immunization process. The p.o. group of pigs immunized with CSFV vaccine+LTRG developed significantly (0.01<P<0.05) higher IgG antibody titers than i.m. or i.n. groups only immunized with CSFV vaccine at the14th day after the third immunization. And at the28th day after third immunization, IgG titers of pigs immunized with CSFV vaccine+LTRG in p.o. group were the highest. Pigs immunized with with CSFV vaccine+LTKG and CSFV+LTRG in the i.n. and p.o. groups showed significant (0.01<P<0.05) higher mucosal IgA antibody titers than pigs of i.m. groups only immunized with CSFV vaccine after the third immunization. In addition, pigs of i.n. group immunized with CSFV vaccine+LTKG developed the highest mucosal IgA antibody titers and showed the significantly (0.01<P<0.05) difference with i.n. group only immunized with CSFV vaccine. In the MTT assay, the results showed that T lymphocyte proliferation ability of i.n. group immunized with CSFV vaccine+LTKG was stronger than i.n. group only immunized with CSFV vaccine and also p.o. group immunized with CSFV vaccine+LTKG was stronger than p.o. group only immunized with CSFV vaccine. However, intranasally immunization was better than peroral administration immunization for lymphocyte proliferation.The results suggested that CSFV vaccine with LTKG or LTRG could elicit humoral, cellular and local mucosal immune response through mucosal immunization both in mice and pigs. In particular, the mucosal secreting antibody provided protective barrier for local neutralizing infection of wild virus and made immunoprophylaxis function developed effectively.