The aim of the study was to develop an assay which can distinguish between classical and variant strains of porcine epidemic diarrhea virus (PEDV), and understand the recent epidemiology knowledge of porcine epidemic diarrhea (PED) in China. Two pairs of primers based on the S gene differentiated between variant and classical PEDV strains were redesigned. A nest PCR associated with a novel method of nanoparticle constructed nano-nest PCR was developed. Seventy-eight cases collected in 9 different areas in China from 2015 to 2016 were tested using the nano-nest PCR assay. The results indicated that the lowest detection limit of the nano-nest PCR assay was 2.21×10^-7 ng/µL which was 100-fold more sensitive than common RT-PCR. This assay was highly specific to PEDV, which did not amplify DNA or cDNA of pseudorabies virus, porcine reproductive and respiratory syndrome virus, classical swine fever virus, porcine rotavirus, porcine transmissible gastroenteritis virus. The average positive rate was 74.36% (58/78) for the PED, and the positive rate of the variant strains of PEDV was 79.31% (46/58). Our results demonstrated that the nano-nest PCR allows for accurate and sensitive detection of variant and classical PEDV infection. PEDV infection was mainly the variant strains of PEDV in China.

Key words: Detection, Epidemiology, Nano-nest PCR, Porcine epidemic diarrhea virus (PEDV)