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.
Reid
Scott M
SM
Institute for Animal Health, Ash Road, Pirbright, Woking, Surrey, GU24 0NF, UK.
Pierce
Kenneth E
KE
Mistry
Rohit
R
Bharya
Sukvinder
S
Dukes
Juliet P
JP
Volpe
Carmelo
C
Wangh
Lawrence J
LJ
King
Donald P
DP
D000818:Animals; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D005536:Foot-and-Mouth Disease / Q000175:diagnosis / Q000821:virology; D029544:Foot-and-Mouth Disease Virus / Q000145:classification / Q000201:enzymology / Q000235:genetics*; D016679:Genome, Viral / Q000235:genetics; D008969:Molecular Sequence Data; D012367:RNA, Viral / Q000235:genetics; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D012703:Serotyping; D013045:Species Specificity; D014764:Viral Proteins / Q000235:genetics*
D018076:DNA, Complementary; D012367:RNA, Viral; D014764:Viral Proteins; D012321:DNA-Directed RNA Polymerases