Evaluation of immunogenicity of Streptococcus equi ssp. zooepidemicus surface protein FIPP

【Objective】This study aimed to investigate the immune efficacy of the surface protein FIPP(fibrinogenand Ig-binding protein precursor) of Streptococcus equi ssp. zooepidemicus(SEZ) and to provide a new candidate vaccine antigen for SEZ. 【Method】According to the FIPP gene sequence published in the NCBI database, the primer was designed for PCR amplification, and the amplified fragment was cloned into the prokaryotic expression vector pColdⅠ, and then transformed into Escherichia coli BL21(DE3) receptor cells. The FIPP protein was expressed and purified in E. coli through IPTG induction. Identification by SDS-PAGE. BALB/c mice were immunized with purified recombinant FIPP(rFIPP). 14 d later, the mice were immunologically protected and serum was collected. The liver, spleen and lungs of the mice infected with SEZ were dissected for histopathological observation. Enzyme-linked immunosorbent assay(ELISA) was used to detect specific antibody IgG levels in serum of mice, the reactivity of recombinant protein was detected by Western blotting analysis, and the bactericidal activity of serum was detected by whole blood bactericidal test. 【Result】The protein fragments of about 96 kD were observed by SDS-PAGE electrophoresis, which was consistent with the theoretical molecular weight of FIPP protein. The protection rate of rFIPP against SEZ attack was 70.0% in mice immunized with RFIPP. ELISA assay results showed that rFIPP immunized mice with high titer specific antibody immunoglobulin G(IgG), which was extremely significantly higher than that of negative control(P<0.01, the same below). By IgG subtype analysis, the antibody level of IgG1 was significantly higher than that of IgG2a(P<0.05). These results indicated that IgG1 was the main antibody subtype produced after rFIPP immunization. The results of whole blood bactericidal test showed that rFIPP superimmune serum had an extremely significant killing effect on SEZ. The histopathological observation showed that rFIPP immunization could effectively reduce the pathological damage of various organs in mice. 【Conclusion】In this study,recombinant protein rFIPP obtained by expression purification has good immunoprotective efficacy in a mouse infecting SEZ model and can be used as a valid candidate antigen for SEZ subunit vaccine.