TY - JOUR T1 - Laboratory validation of two real-time RT-PCR methods with 5'-tailed primers for an enhanced detection of foot-and-mouth disease virus. JF - J Virol Methods Y1 - 2017 A1 - Frank Vandenbussche A1 - David Lefebvre A1 - Ilse De Leeuw A1 - Steven Van Borm A1 - Kris De Clercq KW - 5' Untranslated Regions KW - Animals KW - DNA Primers KW - Foot-and-Mouth Disease KW - Foot-and-Mouth Disease Virus KW - Reproducibility of Results KW - Reverse Transcriptase Polymerase Chain Reaction KW - RNA, Viral KW - Sensitivity and Specificity KW - Sheep KW - Swine AB -

The 3D and 5UTR real-time RT-PCR assays (RT-qPCR) from Callahan et al. (2002) and Reid et al. (2002) are commonly used reference methods for the detection of foot-and-mouth disease virus (FMDV). For an optimal detection of FMDV in clinical samples, it is advised to use both assays simultaneously (King et al., 2006). Recently, Vandenbussche et al. (2016) showed that the addition of 5'-tails to the FMDV-specific primers enhances the detection of FMDV in both the 3D and the 5UTR RT-qPCR assay. To validate the 3D and 5UTR RT-qPCR assays with 5'-tailed primers for diagnostic purposes, both assays were run in parallel in a triplex one-step RT-qPCR protocol with beta-actin as an internal control and synthetic RNA as an external control. We obtained low limits of detection and high linearity's, high repeatability and reproducibility, near 100% analytical specificity and >99% diagnostic accuracy for both assays. It was concluded that the 3D and 5UTR RT-qPCR assays with 5'-tailed primers are particularly suited for the detection of FMDV as well as to exclude the presence of FMDV.

VL - 246 U1 - https://www.ncbi.nlm.nih.gov/pubmed/28457784?dopt=Abstract M3 - 10.1016/j.jviromet.2017.04.014 ER -