title: Cell culture isolation and sequence analysis of genetically diverse US porcine epidemic diarrhea virus strains including a novel strain with a large deletion in the spike gene.
abstract: The highly contagious and deadly porcine epidemic diarrhea virus (PEDV) first appeared in the US in April 2013. Since then the virus has spread rapidly nationwide and to Canada and Mexico causing high mortality among nursing piglets and significant economic losses. Currently there are no efficacious preventive measures or therapeutic tools to control PEDV in the US. The isolation of PEDV in cell culture is the first step toward the development of an attenuated vaccine, to study the biology of PEDV and to develop in vitro PEDV immunoassays, inactivation assays and screen for PEDV antivirals. In this study, nine of 88 US PEDV strains were isolated successfully on Vero cells with supplemental trypsin and subjected to genomic sequence analysis. They differed genetically mainly in the N-terminal S protein region as follows: (1) strains (n=7) similar to the highly virulent US PEDV strains; (2) one similar to the reportedly US S INDEL PEDV strain; and (3) one novel strain most closely related to highly virulent US PEDV strains, but with a large (197aa) deletion in the S protein. Representative strains of these three genetic groups were passaged serially and grew to titers of ∼5-6log10 plaque forming units/mL. To our knowledge, this is the first report of the isolation in cell culture of an S INDEL PEDV strain and a PEDV strain with a large (197aa) deletion in the S protein. We also designed primer sets to detect these genetically diverse US PEDV strains.
authors.affiliation: Food Animal Health Research Program, Department of Veterinary Preventive Medicine, The Ohio State University, Wooster, OH, United States; Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.
authors.forename: Tomoichiro
authors.initials: T
authors.lastname: Oka
authors.affiliation: Food Animal Health Research Program, Department of Veterinary Preventive Medicine, The Ohio State University, Wooster, OH, United States. Electronic address: saif.2@osu.edu.
authors.forename: Linda J
authors.initials: LJ
authors.lastname: Saif
authors.affiliation: University of Minnesota Veterinary Diagnostic Laboratory, Saint Paul, MN, United States.
authors.forename: Douglas
authors.initials: D
authors.lastname: Marthaler
authors.affiliation: Food Animal Health Research Program, Department of Veterinary Preventive Medicine, The Ohio State University, Wooster, OH, United States.
authors.forename: Malak A
authors.initials: MA
authors.lastname: Esseili
authors.affiliation: Molecular and Cellular Imaging Center, Ohio Agricultural Research and Development Center, The Ohio State University, Wooster, OH, United States.
authors.forename: Tea
authors.initials: T
authors.lastname: Meulia
authors.affiliation: Food Animal Health Research Program, Department of Veterinary Preventive Medicine, The Ohio State University, Wooster, OH, United States.
authors.forename: Chun-Ming
authors.initials: CM
authors.lastname: Lin
authors.affiliation: Food Animal Health Research Program, Department of Veterinary Preventive Medicine, The Ohio State University, Wooster, OH, United States.
authors.forename: Anastasia N
authors.initials: AN
authors.lastname: Vlasova
authors.affiliation: Food Animal Health Research Program, Department of Veterinary Preventive Medicine, The Ohio State University, Wooster, OH, United States.
authors.forename: Kwonil
authors.initials: K
authors.lastname: Jung
authors.affiliation: Animal Disease Diagnostic Laboratory, Ohio Department of Agriculture, Reynoldsburg, OH, United States.
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Food Animal Health Research Program, Department of Veterinary Preventive Medicine, The Ohio State University, Wooster, OH, United States. Electronic address: wang.655@osu.edu.
authors.forename: Qiuhong
authors.initials: Q
authors.lastname: Wang
mesh_terms: D000818:Animals; D001483:Base Sequence; D018929:Cell Culture Techniques / Q000662:veterinary; D002522:Chlorocebus aethiops; D018352:Coronavirus Infections / Q000453:epidemiology / Q000662:veterinary*; D017931:DNA Primers / Q000235:genetics; D019084:Fluorescent Antibody Technique, Indirect / Q000662:veterinary; D014644:Genetic Variation; D046529:Microscopy, Electron, Transmission / Q000662:veterinary; D015146:Midwestern United States / Q000453:epidemiology; D008969:Molecular Sequence Data; D010802:Phylogeny; D053485:Porcine epidemic diarrhea virus / Q000235:genetics* / Q000302:isolation & purification / Q000472:pathogenicity; D060888:Real-Time Polymerase Chain Reaction / Q000662:veterinary; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000662:veterinary; D017422:Sequence Analysis, DNA / Q000662:veterinary; D017384:Sequence Deletion / Q000235:genetics; D013045:Species Specificity; D064370:Spike Glycoprotein, Coronavirus / Q000235:genetics*; D013552:Swine; D013553:Swine Diseases / Q000453:epidemiology* / Q000821:virology*; D014709:Vero Cells; D014774:Virulence
chemical_list: D017931:DNA Primers; D064370:Spike Glycoprotein, Coronavirus
keywords: Cell culture isolation; Complete genome analysis; Large deletion in S protein; PEDV variants; Porcine epidemic diarrhea virus (PEDV)