Recent Outbreaks and emergence of mutants of Porcine Epidemic Diarrhea Viruses (PEDV) in Korea

Abstract

Porcine epidemic diarrhea (PED) virus was first reported in South Korea in 1992 and has since been circulating and exhibiting considerable genetic diversity. During 2013, PED outbreaks reoccurred in South Korea; however, the emerging PEDVs in these outbreaks were not variants of previous Korean isolates or attenuated vaccine strains. During October 2013-June 2014, 30 intestinal samples of dead piglets and 16 fecal swabs were found to be PEDV positive. One field strain of PEDV (named BM1) was successfully adapted for growth on Vero cells. This virus was isolated from BM farm of 60 sows, which had not vaccinated against PEDV. Pigs of all ages from the farm showed clinical symptoms of diarrhea with 100% deaths among suckling piglets and 10% among sows. Examination at necropsy revealed that the dead piglets of BM farm were covered with brown blotches of dried scour material and their stomachs were filled with undigested milk. Thin, translucent small intestines containing yellow fluid were also observed. The BM1 PEDV field isolate induced cytopathic effects of rounded shape within 48 hours at passage 10. The presence of PEDV in the cell culture was confirmed by immunofluorescence assay, which showed the specific fluorescence signal. Beside of microscopic observation, the increment in the quantity of viral RNA was observed to increase as the number of passages increased, from 30,325 copies/mu l (C-T = 16.11) at passage 2 to 418,000 copies/mu l (C-T = 13.77) at passage 10. The infective titers of the BM1 isolate increased from 10(4.7)TCID(50)/ml (at passage 2) to 10(7.9) TCID50/ml (at passage 10). The complete S gene of BM1 (KP861982) was sequenced for genetic characterization. The S gene has 4,161 nt in length and encodes 1,386 aa. The spike protein of the BM1 isolate showed substitutions at neutralizing SS6 epitope from LQDGQVKI to SQSGQVKI but identity at the SS2 and 2C10 neutralizing epitopes. The genetic relationship of the BM1 isolate with other PEDVs in the world was inferred from a codon-based alignment of 409 sequences of the complete S gene. The maximum likelihood phylogenetic tree was constructed by the FastTree program with the general time reversible nucleotide substitution model. The phylogeny constructed on the basis of the complete S gene showed that the BM1 isolate belongs to subgroup 2a, genogroup 2 of PEDV. This isolate clustered closely with emergent PEDV strains in the United States, showing 99.2%-99.7% identity with PEDVs of North American strains. This observation was repeated by the phylogenetic inference of the complete N gene. The branching pattern clearly showed that BM1 is genetically less related (92.9-93.4% identity) to the live vaccine strains derived from genogroup 1 and used currently to prevent PEDV infections in South Korea. And as an additional study, since 1992, porcine epidemic diarrhea virus (PEDV) has become one of the most important porcine diarrhea associated viruses in South Korea. We performed large-scale study of the incidence of PEDV in pigs with diarrhea in South Korea, and consequently identified and characterized a novel PEDV variant with large genomic deletion.