title: Effect of saliva stabilisers on detection of porcine reproductive and respiratory syndrome virus in oral fluid by quantitative reverse transcriptase real-time PCR.
abstract: This study evaluated the effect of extraction-amplification methods, storage temperature and saliva stabilisers on detection of porcine reproductive and respiratory syndrome virus (PRRSV) RNA by quantitative reverse transcriptase real-time PCR (qRT-PCR) in porcine oral fluid. The diagnostic performance of different extraction-amplification methods was examined using a dilution series of oral fluid spiked with PRRSV. To determine RNA stability, porcine oral fluid, with or without commercially available saliva stabilisers, was spiked with PRRSV, stored at 4°C or room temperature and tested for the presence of PRRSV RNA by qRT-PCR. PRRSV RNA could be detected in oral fluid using all extraction-amplification combinations, but the limit of detection varied amongst different combinations. Storage temperature and saliva stabilisers had an effect on the stability of PRRSV RNA, which could only be detected for 7 days when PRRSV spiked oral fluid was kept at 4°C or stabilised at room temperature with a commercial mRNA stabiliser.
authors.affiliation: Operational Direction Viral Diseases, Enzootic and (Re)emerging Diseases, CODA-CERVA, Groeselenberg 99, 1180 Ukkel, Belgium. Inge.Decorte@coda-cerva.be
authors.forename: Inge
authors.initials: I
authors.lastname: Decorte
authors.forename: Yves
authors.initials: Y
authors.lastname: Van der Stede
authors.forename: Hans
authors.initials: H
authors.lastname: Nauwynck
authors.forename: Nick
authors.initials: N
authors.lastname: De Regge
authors.forename: Ann Brigitte
authors.initials: AB
authors.lastname: Cay
mesh_terms: D000818:Animals; D005079:Excipients / Q000737:chemistry*; D019316:Porcine respiratory and reproductive syndrome virus / Q000302:isolation & purification*; D012367:RNA, Viral / Q000302:isolation & purification; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D012463:Saliva / Q000737:chemistry / Q000821:virology*; D013048:Specimen Handling; D013552:Swine
chemical_list: D005079:Excipients; D012367:RNA, Viral
keywords: Oral fluid; Porcine reproductive and respiratory syndrome virus; Quantitative reverse transcriptase real-time PCR