title: Evaluation of two real-time polymerase chain reaction assays for Porcine epidemic diarrhea virus (PEDV) to assess PEDV transmission in growing pigs.
abstract: In April 2013, a Porcine epidemic diarrhea virus (PEDV) epidemic began in the United States. As part of the response, real-time reverse transcription polymerase chain reaction (RT-PCR) assays to detect PEDV were developed by several veterinary diagnostic laboratories. Our study evaluated RT-PCR PEDV assays that detect the N gene (gN) and S gene (gS) for their ability to detect PEDV infection and the transmission potential of pigs experimentally exposed to PEDV. Detection limits and quantification cycle (Cq) values of real-time RT-PCR were assayed for PEDV samples and positive controls for both gN and gS. The limit of detection for the gN assay was 10(-6) (mean Cq: 39.82 ± 0.30) and 10(-5) (mean Cq: 39.39 ± 0.72) for the gS assay with PEDV strain USA/Colorado/2013. Following recommended guidelines, rectal swabs (n = 1,064) were tested; 354 samples were positive by gN assay and 349 samples were positive by gS assay (Cq ≤ 34.99), 710 samples were negative by gN assay and 715 were negative by gS assay (Cq > 34.99) of which 355 and 344 were "undetermined" (i.e., undetected within a threshold of 40 RT-PCR cycles, by gN and gS assays, respectively). The coefficient of variation (intra-assay variation) ranged from 0.00% to 2.65% and interassay variation had an average of 2.75%. PEDV could be detected in rectal swabs from all pigs for ~2 weeks postinfection at which time the prevalence began to decrease until all pigs were RT-PCR negative by 5 weeks postinfection. Our study demonstrated that RT-PCR assays functioned well to detect PEDV and that the gN assay was slightly better.
authors.affiliation: U.S. Department of Agriculture, Agricultural Research Service, National Animal Disease Center, Virus and Prion Research Unit, Ames, IA laura.miller@ars.usda.gov.
authors.forename: Laura C
authors.initials: LC
authors.lastname: Miller
authors.affiliation: U.S. Department of Agriculture, Agricultural Research Service, National Animal Disease Center, Virus and Prion Research Unit, Ames, IA.
authors.forename: Kimberly K
authors.initials: KK
authors.lastname: Crawford
authors.affiliation: U.S. Department of Agriculture, Agricultural Research Service, National Animal Disease Center, Virus and Prion Research Unit, Ames, IA.
authors.forename: Kelly M
authors.initials: KM
authors.lastname: Lager
authors.affiliation: U.S. Department of Agriculture, Agricultural Research Service, National Animal Disease Center, Virus and Prion Research Unit, Ames, IA.
authors.forename: Steven G
authors.initials: SG
authors.lastname: Kellner
authors.affiliation: U.S. Department of Agriculture, Agricultural Research Service, National Animal Disease Center, Virus and Prion Research Unit, Ames, IA.
authors.forename: Susan L
authors.initials: SL
authors.lastname: Brockmeier
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D018352:Coronavirus Infections / Q000662:veterinary* / Q000821:virology; D018562:Disease Transmission, Infectious / Q000662:veterinary*; D005243:Feces / Q000821:virology; D053485:Porcine epidemic diarrhea virus / Q000235:genetics* / Q000472:pathogenicity; D060888:Real-Time Polymerase Chain Reaction / Q000662:veterinary; D048348:Reverse Transcription; D013552:Swine; D013553:Swine Diseases / Q000453:epidemiology / Q000635:transmission / Q000821:virology*; D014481:United States / Q000453:epidemiology
keywords: Porcine epidemic diarrhea virus; real-time reverse transcription polymerase chain reaction; swine