Expression Of Protein VP1 Of Foot-and-mouth Disease Virus In Lactococcus Lactic And Mucosal Immune Effect

Foot-and-mouth disease(FMD) is a highly contagious disease that infects cloven-hoofed animals and leads to significant economic losses. Mucosal vaccination is an effective strategy for inducing specific immune responses against infectious diseases invaded through mucosa, such as FMD. In this study, we expressed the IFN-α gene of bovine and the VP1 gene of FMDV in Lactococcus lactis(L.lactis), respectively. Then, we evaluate the immune effects of the recombinant L.lactis via oral vaccination in mice. The specific research methods and results are as follows:We optimized and synthesized the bovine interferon-α(IFN-α) gene and FMDV VP1 gene according to the codon bias of L.lactis, and constructed the recombinant plasmid pNZ8149-IFNα and pNZ8148-SPVP1 respectively. The recombinant plasmids were respectively transformed into L.lactis NZ3900 and NZ9000 and identified by PCR amplification, double enzyme digestion and gene sequencing. SDS-PAGE and Western blotting analysis showed that the two kinds of protein were expressed in L.lactis successfully.Then we evaluated the mucosal immune effects of NZ9000/pNZ8148-SPVP1 in mice via oral administration. Meanwhile, NZ9000/pNZ8148 was used as empty vector control, inactivated vaccine was used as positive control, and PBS was used as mock. Antibody test results showed that anti-FMDV IgA and IgG in serum and mucosal sIgA reached high levels in the L.lactis NZ9000/pNZ8148-SPVP1 groups, and no signification anti-FMDV antibodies were observed in the empty vector control group and mock group. These results indicated that orally administered NZ9000/pNZ8148-SPVP1 were able to elicit both FMDV-specific systemic and mucosal immune responses. CD4 + and CD8 + T cells in peripheral blood in the experimental group rose a higher level than the mock group. Furthermore, compared to the mock group, T spleen lymphocyte proliferated significantly after stimulation with antigen in NZ9000/pNZ8148-SPVP1 group and in positive control group, while no obvious proliferation occurred in the NZ9000/pNZ8148 group. Neutralizing antibody test showed that the serum of mice orally immunized with NZ9000/pNZ8148-SPVP1 exhibited higher level of FMDV-neutralizing antibody than the empty vector control group and mock group. Cytokines detection showed that recombinant NZ9000/pNZ8148-SPVP1 generated much more IL-2, IFN-γ, IL-4, IL-5 and IL-10 than the other three groups, while IL-4, IL-5 and IL-10 were more significant. These results indicated that recombinant L.lactis could stimulate organism to produce both humoral immune response and cellular immune response. In conclusion, recombinant L.lactis as a live vector vaccine has a promising perspective in the development of novel FMD vaccines.