Reverse transcription-PCR using a Primer Set Targeting the 3D Region Detects Foot-and-Mouth Disease Virus with High Sensitivity

Abstract

12Because foot-and-mouth disease (FMD) has the potential to spread extensively, methods used for its 13 diagnosis must be rapid and accurate. Therefore, reverse transcription-PCR (RT-PCR) plays an important 14 diagnostic role. Here we designed the primer set FM8/9 to amplify 644 bases of the conserved 3D region of 15 all seven serotypes of FMD virus (FMDV). We compared the performance of RT-PCR assays using FM8/9 16 with that using the primer set 1F/R targeting the 5'-UTR described in the manual of the World Organization 17for Animal Health. The detection limits of the RT-PCR assays were determined for 14 strains representing all 18 serotypes. Compared with the sensitivities of the RT-PCR assay using 1F/R, those using FM8/9 were 10 1 -to 19 10 4 -fold higher for eight strains. To assess the validity of the methods for analyzing clinical samples, sera and 20 saliva samples from pigs and cows infected with FMDV were collected daily and analyzed using the two PCR 21 assays. The FM8/9 assay detected FMDV from all infected pigs and cows for longer times compared with the 22 1F/R assay, therefore revealing higher sensitivity for the clinical samples. Our results suggest that the FM8/9 23 RT-PCR assay is highly sensitive and is therefore suitable for the diagnosis of FMD. 24 25