Development Of Porcine Transmissible Gastroenteritis-Porcine Epidemic Diarrhea Bivalent Inactivated Vaccine

Since October2010, cute diarrhea outbreaks caused by porcine epidemicdiarrhea virus (PEDV) has been observed in Southeast Asian countries, resμLting inmahor losses of suckling piglets. Morbidity rates between90and100%have beenobserved, with mortalities of70-100%on affected pig farms. Currently SoutheastAsia, Germany and the United States have reported outbreaks of PED. In the eventof epidemic diarrhea in pigs, porcine transmissible gastroenteritis virus also has avery high detection rate. Research on the genetic characters of current PEDVChinese isolates showed that there were significant differences, especially the Sgene, between these isolates and PEDV vaccine strain XXX株, which has been usedfor several years in China. The fact that pig herds immunized with vaccine based onXXXstrain still got infected with current Chinese isolates, suggests that current PEDvaccine cannot provide complete protection against current isolates. In this paper, Sgene N-terminal hypervariable region of porcine epidemic diarrhea virus isolated inChina XXXwas expressed in the E.coli BL21, and purified S protein wasincoperated into porcine transmissible gastroenteritis-porcine epidemic diarrheabivalent vaccine. Immunogenicity of the prepared TGE-PED vaccine was evaluated.This research laid a foundation for further development of efficient vaccines foreffective prevention and control of porcine epidemic diarrhea caused by PEDV andporcine transmissible gastroenteritis.1. Establishment of PEDV indirect ELISA antibody detection methodpET-28a-PEDV-S recombinant positive bacteria expressing PEDVXXXvaccine strain S protein N-terminal1077bp (YSA) were induced underconditions of37℃with40uM IPTG. SDS-PAGE analysis showed that the proteinwas highly expressed with a molecμLar weight of42KD, and mainly in the form ofa fusion protein. Western blotting analysis further showed that the expressed proteinhas good immune reactivity. A PEDV indirect ELISA was developed by using thepurified protein as coating antigen.The optimized amount of antigen was0.5ug/well,work dilution of serum sample was1:100, and that of HRP-labeledsecondary antibody was1:2000.Tested serum with OD450values≥0.25wasconsidered as anti-PEDV antibody positive. Coincidence rate between developedand commercial ELISA was100%. 2. Development of TGE-PED bivalent inactivated vaccineTo develop TGE-PED bivalent inactivated vaccine, S protein N-terminalhypervariable region of PEDV epidemic strain XXXwas cloned into prokaryoticexpression vector pBV220, the expressed protein designed as LSA was expressedmainly in the form of inclusion bodies. Purified LSA protein was incoperated intoTGE-PED bivalent vaccine. Immunogenicity of the prepared TGE-PED vaccinewas evaluated. The data showed that there were significant differences inanti-PEDV antibody levels between vaccine with or without LSA protein. Theanti-PEDV antibody level in animals immunized with TGE-PED bivalent vaccinewith LSA protein was higher than those of in animals immunized with commercialTGE-PED vaccine. Sera from TGE-PED plus LSA protein vaccine group coμLdneutralize XXXvaccine strain.