title: Isolation and characterization of porcine epidemic diarrhea viruses associated with the 2013 disease outbreak among swine in the United States.
abstract: Porcine epidemic diarrhea virus (PEDV) was detected in May 2013 for the first time in U.S. swine and has since caused significant economic loss. Obtaining a U.S. PEDV isolate that can grow efficiently in cell culture is critical for investigating pathogenesis and developing diagnostic assays and for vaccine development. An additional objective was to determine which gene(s) of PEDV is most suitable for studying the genetic relatedness of the virus. Here we describe two PEDV isolates (ISU13-19338E and ISU13-22038) successfully obtained from the small intestines of piglets from sow farms in Indiana and Iowa, respectively. The two isolates have been serially propagated in cell culture for over 30 passages and were characterized for the first 10 passages. Virus production in cell culture was confirmed by PEDV-specific real-time reverse-transcription PCR (RT-PCR), immunofluorescence assays, and electron microscopy. The infectious titers of the viruses during the first 10 passages ranged from 6 × 10(2) to 2 × 10(5) 50% tissue culture infective doses (TCID50)/ml. In addition, the full-length genome sequences of six viruses (ISU13-19338E homogenate, P3, and P9; ISU13-22038 homogenate, P3, and P9) were determined. Genetically, the two PEDV isolates were relatively stable during the first 10 passages in cell culture. Sequences were also compared to those of 4 additional U.S. PEDV strains and 23 non-U.S. strains. All U.S. PEDV strains were genetically closely related to each other (≥99.7% nucleotide identity) and were most genetically similar to Chinese strains reported in 2011 to 2012. Phylogenetic analyses using different genes of PEDV suggested that the full-length spike gene or the S1 portion is appropriate for sequencing to study the genetic relatedness of these viruses.
authors.affiliation: Department of Veterinary Diagnostic and Production Animal Medicine, College of Veterinary Medicine, Iowa State University, Ames, Iowa, USA.
authors.forename: Qi
authors.initials: Q
authors.lastname: Chen
authors.forename: Ganwu
authors.initials: G
authors.lastname: Li
authors.forename: Judith
authors.initials: J
authors.lastname: Stasko
authors.forename: Joseph T
authors.initials: JT
authors.lastname: Thomas
authors.forename: Wendy R
authors.initials: WR
authors.lastname: Stensland
authors.forename: Angela E
authors.initials: AE
authors.lastname: Pillatzki
authors.forename: Phillip C
authors.initials: PC
authors.lastname: Gauger
authors.forename: Kent J
authors.initials: KJ
authors.lastname: Schwartz
authors.forename: Darin
authors.initials: D
authors.lastname: Madson
authors.forename: Kyoung-Jin
authors.initials: KJ
authors.lastname: Yoon
authors.forename: Gregory W
authors.initials: GW
authors.lastname: Stevenson
authors.forename: Eric R
authors.initials: ER
authors.lastname: Burrough
authors.forename: Karen M
authors.initials: KM
authors.lastname: Harmon
authors.forename: Rodger G
authors.initials: RG
authors.lastname: Main
authors.forename: Jianqiang
authors.initials: J
authors.lastname: Zhang
mesh_terms: D000818:Animals; D016000:Cluster Analysis; D018352:Coronavirus Infections / Q000453:epidemiology / Q000662:veterinary* / Q000821:virology; D004196:Disease Outbreaks; D016679:Genome, Viral; D042822:Genomic Instability; D005838:Genotype; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D010802:Phylogeny; D053485:Porcine epidemic diarrhea virus / Q000235:genetics / Q000302:isolation & purification* / Q000648:ultrastructure; D012367:RNA, Viral / Q000235:genetics; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D012692:Serial Passage; D013552:Swine; D013553:Swine Diseases / Q000453:epidemiology / Q000821:virology*; D014481:United States / Q000453:epidemiology; D014776:Virus Cultivation
chemical_list: D012367:RNA, Viral