Immunogenicity Of Baculovirus Expression System Expressing PEDV S1 Protein

Porcine epidemic diarrhea(PED)is one of the most threatening global pandemic swine diseases,caused by porcine epidemic diarrhea virus(PEDV)infection,which has a high mortality rate.The frequent PED outbreaks have caused a great loss to the global pig industry.But so far there is no security and effective vaccine to prevent this disease.It is needed to develop PEDV vaccine by new means.The S protein is the main structure protein of PEDV,and composed of two domains: S1(aa 1-789)and S2(aa 790-1,383).S1 protein contains the major antigen epitopes that mediate producing neutralizing antibody post infection.Therefore,S1 is often used as the most important target protein in PEDV genetically engineered vaccine study.In this study,we intended to construct recombinant baculoviruses including rvAc-S1,rvAc-SP-S1,rvAc-SP-S1-TMD,rvAc-SP-S1-eGFP,rvAc-SP-S1-eGFP-TMD to express S1 protein in Sf9 cells for subunit vaccine preparation,or display S1 protein on viral particle surface for using as virus live vector vaccine.The preliminary immune effect evaluation of recombinant S1 subunit vaccine and S1-displaying baculovirus vaccine was conducted in a mouse model.The expression and position of S1 in Sf9 cell or on viral particle surface was analyzed by direct fluorescence,indirect immunofluorescence assay,western blotting and immune electron microscopy.Results showed that S1 protein can be efficiently expressed in infected Sf9 cells of rvAc-SP-S1-eGFP and rvAc-SP-S1-eGFP-TMD,and the S1 expression level in infected Sf9 cells of rvAc-SP-S1-eGFP is higher than rvAc-SP-S1-eGFP-TMD;S1 is displayed along the membrane of infected Sf9 cells and on the surface of rvAc-SP-S1-eGFP-TMD.Recombinant S1 protein immunogen and S1-displaying baculovirus particles immunogen were prepared and then subcutaneously immunized BALB/c mice.The humoral and cellular immunological effects were identified by indirect ELISA,micro cell culture neutralization test and spleen lymphocyte proliferation test.Results showed that specific antibody in mice serum of S1-displaying baculovirus immunogen was detected and up to 1:5,000;and the immune serum has the capacity to neutralize PEDV with a titre of 1:26;spleen cells from mice immunized with recombinant viruses exhibited more significant proliferation than the control groups(P>0.05).But it does not achieve the ideal effect of mice immuned with S1.In conclusion,rvAc-SP-S1-eGFP-TMD can induce efficient humoral and cellular immunity post mice immunization.Taken together,we first reported the expression of full-length S1 protein of PEDV CV777 in insect cells using baculovirus and displayed this protein on the surface of viral particles,which elicited efficient immunity on vaccinated mice.This work paves a way for developing new vaccines against PEDV infection.