Molecular And Serological Epidemiology Of Porcine Epidemic Diarrhea Virus In Hunan Province

Porcine epidemic diarrhea (PED) is caused by porcine epidemic diarrhea virus (PEDV) which belongs to family Coronaviridae and genera Coronavirus. It is a highly contagious disease which causes vomiting, diarrhea and dehydration in pig of various age groups. It is also a disease spreading widely around the world including China, which caused severe economic damage to the pig industry.In this study, anatomy was applied in60piglets with diarrhea, and RT-PCR was conducted on intestine and its content, the result showed that65%piglets (39out of60) were infected with PEDV. Partial nucleotide sequence of S gene was subjected to sequencing and alignment with PEDV reference strain, the results indicated that PEDV strains that prevalent in Hunan were similar to strains in other provinces such as Henan, Heilongjiang and Guangdong of China.S protein encoded by S gene of PEDV is the main structural protein, which is also the spike protein that present outside the envelope of the virus, it recognizes the target cell, promotes infusion of the virus with the host cell and plays important roles in virus entry, neutralizing antibody producing and induction of the immune system. Thus fragment spanning between2983nt and3927nt of S gene was amplified by PCR, which was named S’gene and cloned into prokaryotic expression system pET-28a, the resulting recombinant pET-28a-S’was transferred into host cell BL21(DE3) for expression. The result showed that the recombinant partial S protein was expressed in the host cell and the expressed protein was further purified. The purified protein achieved a putridity of above90%, which demonstrated good immunoreactivity by Western blot. This recombinant S protein was used to coating the ELISA plate as coating antigen, after optimizing the working conditions for indirect ELISA, an indirect ELISA for detection of PEDV antibodies was successfully established. This indirect ELISA was applied to sera collected in Hunan, the positive rate for PEDV indicated by our ELISA was53.1%.