The dynamics of Chinese variant porcine epidemic diarrhea virus production in Vero cells and intestines of 2-day old piglets

Abstract

A severe porcine epidemic diarrhea (PED) epizootic has been affecting pigs of all ages that are characterized by high mortality among suckling piglets in China since late 2010, causing significant economic losses. Obtaining a current-circulating PEDV variant isolate that can grow efficiently in cell culture is prerequisite for the development of efficient vaccines. In this study, PEDV strain HN1303 was isolated successfully on Vero cells with supplemental trypsin, and the isolate has been serially propagated in cell culture for over 95 passages. The infectious titers of the virus during the first 10 passages ranged from 10(2.6) to 10(5.8) 50% tissue culture infective doses (TCID50)/ml, and the titers of 20-95 passages ranged from 10(6.2) to 10(8.0)TCID50/ml. The growth curve of Vero cell-adapted HN1303 in cell culture was determined, and dynamics of virus production was confirmed by immunoperoxidase monolayer assay (IPMA). Sequence and phylogenetic analysis based on spike gene indicate that the HN1303 strain belongs to genotype IIa. In addition, the fourth passage cell-culture HN1303 was subjected to 2-day old piglets. All piglets orally inoculated developed severe watery diarrhea and vomiting within 24 hours post-inoculation (hpi) and died within 72 hpi. The results of animal experiments reveal that this strain is highly pathogenic to 2-day old piglets.

Keywords: Dynamics of virus production; Immunoperoxidase monolayer assay; Porcine epidemic diarrhea virus; Serial propagation in cell culture.