Reverse transcription‐PCR using a primer set targeting the 3D region detects foot‐and‐mouth disease virus with high sensitivity

The potential of foot-and-mouth disease (FMD) to spread extensively means that rapid and accurate methods are needed for its diagnosis. Therefore, reverse transcription-PCR (RT-PCR) plays an important diagnostic role. Here, we designed the primer set FM8/9 to amplify 644 bases in the conserved 3D region of all seven serotypes of the FMD virus (FMDV). We compared the performance of RT-PCR assays using FM8/9 with those using the primer set 1F/R, which targets the 5′-UTR, and real-time RT-PCR (rRT-PCR) assays described in the World Organization for Animal Health manual. Detection limits of the RT-PCR assays were determined for 24 strains, representing all serotypes. The sensitivities of RT-PCR assays using FM8/9 were 100.6- to 103.8-fold higher than those of 1F/R assays for 21 strains. To assess the validity of the methods for analysing clinical samples, sera and saliva samples collected daily from pigs and cows infected with FMDV were analysed using the four PCR assays. FM8/9 assays detected FMDV from all infected pigs and cows for longer periods than 1F/R assays, indicating that FM8/9 assays have higher sensitivity for the clinical samples. Our results suggest that the FM8/9 RT-PCR assay is highly sensitive and is therefore suitable for the diagnosis of FMD.