Abstract: This paper aims to establish a triple fluorescence quantitative PCR method for the simultaneous detection of porcine epidemic diarrhea virus (PEDV), porcine rotavirus type A (PoRVA) and porcine deltacoronavirus (PDCoV). Primers and probes were designed for PEDV-N gene, PoRVA-VP6 gene and PDCoV-M gene, and their performance was evaluated after the conditions were optimized, and compared with commercial kits. The results showed that the triple RT-qPCR method established in this study had good specificity and did not amplify positive nucleic acids such as PRRSV, PCV2 and PRV. The detection limits of PEDV, PoRVA and PDCoV were all 1 copies·μL^-1. The coefficient of variation within and between groups was less than 1%. The sample has wide adaptability, and the coefficient of variation is less than 1% when different sample types are detected. Compared with commercial kits, the detection coincidence rates of PEDV and PoRVA were 92.5% and 97.5%, and the detection range of PDCoV was wider, and the detection effect of PDCoV variant strains was still good. The detection method established in this study has the advantages of good specificity, high sensitivity, strong stability and wide sample adaptability, which provides a good detection method for clinical diarrhea samples.

Key words: porcine epidemic diarrhea virus, porcine rotavirus type A, porcine deltacoronavirus, triple RT-qPCR