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ARS Home » Midwest Area » Ames, Iowa » National Animal Disease Center » Virus and Prion Research » Research » Publications at this Location » Publication #308500

Title: Performance assessment of a real-time polymerase chain reaction assay for porcine epidemic diarrhea virus to assess PEDV transmission in growing pigs

item Miller, Laura
item CRAWFORD, KIMBERLY - Orise Fellow
item Lager, Kelly

Submitted to: Conference Research Workers Disease Meeting
Publication Type: Abstract Only
Publication Acceptance Date: 9/26/2014
Publication Date: N/A
Citation: N/A

Interpretive Summary:

Technical Abstract: PEDV was first diagnosed in the U.S. in April 2013 as sporadic cases of diarrhea in young piglets with high mortality. Real-time RT-PCR is a high throughput test system that has potential to detect PEDV during the acute phase of the infection or pre-seroconversion. A study in nursery pigs was conducted to assess the transmission potential of young pigs experimentally-infected or experimentally-exposed to PEDV. On day 0 (D0), a 4-week-old pig was challenged with PEDV and 14 naïve contacts were comingled. On D7, 9 contact pigs were moved to a new room to serve as the principal virus reservoir group (PG), and comingled with 1 naïve age-matched sentinel (S1). Three days later, the S1 pig was moved to a separate room until necropsy. This process was repeated on D14, 21 and 28 with pigs S2, S3 and S4. On day 49, 5 naïve age-matched pigs (N) and the PG were challenged (N/C, PG/C) with homologous virus and euthanized on D78. A daily rectal swab was collected from each pig and tested for PEDV using real-time RT-PCR to detect the N gene (gN) and the S gene (gS) using commercial chemistry. Detection limits and threshold cycle (Ct) values of real-time RT-PCR were assayed for PEDV samples and positive controls for both gN and gS. The coefficient of variation determined based on the replicates (intra-assay variation) ranged from 0.00% to 2.65% and inter-assay variation had an average of 2.75%. Real-time RT-PCR PEDV assay results are visualized in a heat map where positive is Ct = or < 34.99. All PG pigs were real-time RT-PCR-positive from D3-11, with some intermittently positive to D42. Following challenge at D49, all PG pigs were negative post-challenge (PC) and all N/C pigs were positive by D52 days. 3/5 pigs until 13 days PC. PEDV RNA was detected in S1 and S2 within 1 day of contact, but not detected in S3 or S4. Real-time RT-PCR positive rectal swabs collected after D21 were tested for infectious virus using bioassay techniques. A critical need for the current PEDV surveillance program is the rapid detection of PEDV. The present study evaluated the real-time RT-PCR assay to detect PEDV infection in the transmission potential of young pigs experimentally-exposed to PEDV.