J Vet Sci.  2018 Jan;19(1):71-78. 10.4142/jvs.2018.19.1.71.

Isolation and characterization of a new porcine epidemic diarrhea virus variant that occurred in Korea in 2014

• Yang DK
• Kim HH
• Lee SH
• Yoon SS
• Park JW
• Cho IS

• KMID: 2402697
• DOI: http://doi.org/10.4142/jvs.2018.19.1.71

Abstract

Outbreaks of porcine epidemic diarrhea (PED) have resulted in significant economic losses in the swine industry, and another PED outbreak occurred in 2014 in Korea. Isolating and culturing PED virus (PEDV) allow investigations into its pathogenesis and the development of vaccines and diagnostic assays. In this study, we successfully isolated two PEDV isolates (QIAP1401 and QIAP1402) from naturally infected piglets at Jeju-do, Korea. Viral propagation was confirmed in Vero cells based on cytopathic effect, immunofluorescence assay, reverse transcription-polymerase chain reaction, and electron microscopic analyses. The QIAP401 isolate propagated well in Vero cells for 70 passages, with titers of 10(6.5) to 10(7.0) 50% tissue culture infectious dose/mL, which increased gradually with passaging. The nucleotide and amino acid sequences of the QIAP1401 isolate were determined and compared with those of other PEDV isolates. The QIAP1401 isolate was determined to be closely related to the USA/Minnesota271/2014 strain (> 99.9% nucleotide similarity) that was isolated in the USA in 2014. Phylogenetic analysis based on several PEDV genes suggested that a new PEDV variant is circulating in the Korean swine industry, with 93.08% similarity to the SM98 strain isolated in 1998. In addition, the QIAP1401 strain showed strong virulence in 3-day-old piglets and 11-week-old growing pigs.

Keyword

isolation; porcine epidemic diarrhea virus; swine

MeSH Terms

Amino Acid Sequence
Diarrhea
Disease Outbreaks
Fluorescent Antibody Technique
Jeju-do
Korea*
Porcine epidemic diarrhea virus*
Swine
Vaccines
Vero Cells
Virulence
Vaccines

Figure

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  Fig. 1 Cytopathic effects of (A–C), and immunofluorescence results for (D–F), porcine epidemic diarrhea virus isolates in infected Vero cells at 200×. Vero cells were infected with the QIAP1401 or QIAP1402 isolates and maintained in DMEM containing 1 µg/mL trypsin

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  Fig. 2 Virus titers according to number of the porcine epidemic diarrhea virus (PEDV) passages (A) and growth curves of the QIAP1401-P10, -P40, and -P70 strains according to time of harvesting in Vero cells (B). TCID, tissue culture infectious dose.

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  Fig. 3 Virus particles from the QIAP1401 strain propagated in Vero cells. Negatively stained porcine epidemic diarrhea virus particles of 80 to 100 nm in diameter are visible (100,000×). Scale bar = 100 nm.

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  Fig. 4 Phylogenetic analysis based on the nucleotide sequences of the spike (S) gene of porcine epidemic diarrhea viral strains. The QIAP1401 and QIAP1402 strains were predicted to belong to porcine epidemic diarrhea virus group G2. The phylogenetic tree was constructed based on aligned nucleotide sequences obtained by using the neighbor-joining method. KOR, Korea; CAN, Canada; JPN, Japan; CHN, China; BEL, Belgium; G1, genotype 1; G2a, genotype 2a; G2b, genotype 2b.

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  Fig. 5 Mean clinical symptom scores for 3-day-old piglets (A) and 11-week-old growing pigs (B) infected with the QIAP1401-P70 strain, and the mean score obtained by reverse transcription-polymerase chain reaction in feces following oral administration of porcine epidemic diarrhea viral strains. RT-PCR, reverse transcription-polymerase chain reaction.

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  Fig. 6 Jejunal sections of piglets infected with the QIAP1401 strain. Severe atrophy of villi and several cytoplasmic vacuoles were observed in intestinal epithelial cells following hematoxylin and eosin staining (20×) (A); brown staining, indicating a positive reaction, was identified by immunohistochemistry (20×) (B).

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  Fig. 7 Number of porcine epidemic diarrhea (PED) cases () and PED virus-infected piglets (◆) in Korea between January 2000 and December 2016.

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