Pan-serotypic detection of foot-and-mouth disease virus by RT linear-after-the-exponential PCR

A reverse transcription Linear-After-The-Exponential polymerase chain reaction (RT LATE-PCR) assay was evaluated for detection of foot-and-mouth disease virus (FMDV). This pan-serotypic assay targets highly conserved sequences within the 3D (RNA polymerase) region of the FMDV genome, and uses end-point hybridisation analysis of a single mismatch-tolerant low temperature probe to confirm the identity of the amplicons. An Armored RNA® served as an internal control to validate virus negative results. The ability of the assay to identify FMDV was directly compared to a real-time RT-PCR assay routinely used by reference laboratories. The analytical sensitivity of the RT LATE-PCR assay was 10 genomic copies and the dynamic range of the test was identical to real-time RT-PCR based on decimal dilutions of an FMDV-positive sample. This pan-serotypic assay was able to detect FMDV in a broad range of clinical samples collected from field cases of FMD (n = 121), while samples of other viruses causing vesicular disease in livestock and genetic relatives of FMDV were negative. In addition to the laboratory-based utility of this diagnostic test, the RT LATE-PCR assay format has potential application for use in a portable (“point-of-care”) device designed to achieve rapid detection of FMDV in the field.