Construction of Recombinant Pseudorabies Virus Coexpressing E and M Protein of Porcine Reproductive and Respiratory Syndrome Virus Fused with VP22 Protein of Bovine Herpesvirus 1 and Its Immunogenicity in Mice

In the present study,the ORF5M gene(the modified ORF5 gene) and ORF6 gene,the two major immunogenicity genes of PRRSV,were used as the target genes,and bovine herpesvirus 1(BHV-1)VP22 possessing protein transduction property was chosed as the immunoadjuvant.Based on homologous recombination,a recombinant PRV(rPRV) TK~-/gE~-/VP22E~+/VP22M~+ coexpressing PRRSV E and M protein fused with BHV-1 VP22 protein was constructed by using PRV TK~-/gE~-/LacZ~+ as parent strain.The rPRV was confirmed by PCR,Southern blot and Western blot.The results of TCID_(50) tests showed that the insertion of the foreign genes had no influence on the propagation of rPRV in IBRS-2 or PK15 cells.BALB/c mice were immunized with rPRV TK~-/gE~-/VP22E~+/VP22M~+,and compared with rPRV TK~-/gE~-/E~+ expressing ORF5M gene and TK~-/gE~-/E~+/M~+ coexpressing ORF5M and ORF6 genes respectively.Neutralizing antibodies and lymphocyte proliferative responses were evaluated at 10 weeks after primary immunization.Among three rPRVs,TK~-/gE~-/VP22E~+/VP22M~+ had the best effect on enhancing PRRSV-specific humoral and cellular immune responses,and VP22 showed adjuvant efficiencies on rPRV vaccine to a certain degree.In conclusion,the study established a base for the research of bivalent genetic engineering vaccines against PRRSV and PRV.