Establishment and Application of Triple-qPCR for HEV, PEDV and PDCoV in Pork and Its Products

Animal-derived food safety incidents caused by zoonotic diseases occur frequently. In order to control and ensure the safety of pork consumption from its raw material source and to control the cost of pork and its products consumption, this study constructed a triple real-time fluorescent quantitative PCR (qPCR) method that can simultaneously detect hepatitis E virus (HEV), porcine epidemic diarrhea virus (PEDV), and porcine δ corona virus (PDCoV) in pork and its products. The results showed that the triple qPCR method could only amplify the specific gene fragments of three target viruses, which had high specificity. The minimum detection limits of the three viruses (HEV, PEDV, and PDCoV) were 6.02, 6.98 and 6.92 copies/μL respectively. The coefficient of variation (CV%) within and between groups ranged from 0.10% to 3.0%, with good repeatability. The established method was applied to detect 248 samples of pork products and exports and 282 raw pig manure swabs, and parallel detection was also conducted using corresponding virus detection methods for comparison. The detection rates of porcine epidemic diarrhea virus, porcine deltacoronavirus, and hepatitis E virus in pork products and exports were all 0%, which was consistent with the results of the corresponding virus detection methods. The detection rates of the three viruses (PEDV, PDCoV, and HEV) using this method were 1.06%, 3.19% and 0.35%, respectively. The detection rates of the standard method for these three viruses were 1.06%, 3.19% and 0%, respectively. The study showed that the established triplex qPCR detection method can accurately and rapidly detect three viruses in pork and its products or pig samples, providing technical support for ensuring the market circulation of fresh pork and its products and blocking the transmission of viral foodborne diseases.