Preparation And Immunity Of American Type Prrsv Bivalent Vaccine In Mice

Porcine Reproductive and Respiratory Syndrome was first reported in the United States in 1987,and has caused serious economic losses to the pig industry since its outbreak.It was introduced into China in about 1995.The cause of the disease is Porcine Reproductive and Respiratory Syndrome Virus.American PRRSV and its variants are the main prevalent strains in China.In recent years,PRRSV ancestry in China have shown a trend of diversification,which increases the difficulty of PRRS prevention and treatment in China.JXA1-like strain is a HP-PRRSV that has been prevalent since the outbreak of PRRSV,and NADC3 0-like strain is a popular HP-PRRSV strain in recent years.The bivalent nucleic acid vaccine constructed by these two ancestries will be an ideal new vaccine for PRRSV of prevention and control.A PRRSV strain was collected from an infected pig farm and named YN-LQ strain.The whole gene was amplified,then Nsp2?GP5 genes and whole genome went on genetic evolution analysis and homology comparison,and its Nsp2 and GP5 genes went on Sequence alignment of amino acids encoded.The YN-LQ strain was confirmed to be a JXA1-like strain.It is a study on physicochemical characteristics of YN-LQ strain.The results showed that YN-LQ strain was not heat-resistant,acid-resistant and alkali-resistant,sensitive to ether,chloroform and ultraviolet rays,and insensitive to trypsin and ultrasound.It went on Connecting of ORF5 gene of YN-LQ strain with ORF5 gene of NADC30-Like by autolysis Linker.Then it went on Constructing of bivalent nucleic acid vaccine by connecting with pVAXl vector.It made vaccine transfect to BHK cells,which was proved by Western Blot and so on.The results showed that bivalent nucleic acid vaccine could express target protein of GP5(JXA1-Like)-Linker-GP5(NADC30-Like).It went on immunizating of mice with the constructed bivalent nucleic acid vaccine.The results showed that the combination of bivalent nucleic acid vaccine and adjuvant could improve the humoral and cellular immunity of mice,and had good immunogenicity.Detection of IL-4 and IFN-gamma cytokines showed that the level of the combination of bivalent nucleic acid vaccine and adjuvant group was higher than that of bivalent nucleic acid vaccine group 35 days after immunization.Testing neutralizing antibody showed that the titer of neutralizing antibody was 1:17.77 in the group of the combination of bivalent nucleic acid vaccine and adjuvant,which was higher than other groups.The results of CD3~+CD4~+and CD3~+CD8~+serum cytokines showed that the level of the combination of bivalent nucleic acid vaccine and adjuvant group was higher than that of bivalent nucleic acid vaccine group after 35d.The results of T lymphocyte proliferation test showed that the stimulation index(SI)of the specific stimulation group was higher than that of the non-specific stimulation group,the combination of bivalent nucleic acid vaccine and adjuvant and the bivalent nucleic acid vaccine groups were higher than others.the combination of bivalent nucleic acid vaccine and adjuvant group was higher than bivalent nucleic acid vaccine group in specific stimulation.the combination of bivalent nucleic acid vaccine and adjuvant group was higher than bivalent nucleic acid vaccine group in non-specific stimulation too.In conclusion,the combination of adjuvants can improve the immune effect of bivalent nucleic acid vaccine and lay a foundation for further research and development of PRRSV vaccine in America.