A one-step reverse transcriptase loop-mediated isothermal amplification assay for simple and rapid detection of swine vesicular disease virus.
This report describes the development of a one-step reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) assay for the detection of swine vesicular disease virus (SVDV). The assay detects the virus rapidly, within 30-60 min and the result is visualised either by gel-electrophoresis or by the naked eye through the addition of SybrGreen. A collection of 28 SVDV isolates were tested positive, while heterologous viruses such as foot-and-mouth disease virus and vesicular stomatitis virus remained negative. The performance of the RT-LAMP was compared directly with real-time PCR using RNA from clinical samples including nasal swabs, serum and faeces. For nasal swabs and serum the sensitivity of the RT-LAMP was shown to be at least equivalent to real-time PCR. Interestingly, for faecal samples the RT-LAMP assay was shown to be even more sensitive than real-time PCR, possibly because it is less sensitive to inhibitory substances. This RT-LAMP assay provides a number of benefits for the diagnosis of SVD, since the assay is sensitive and rapid, and the isothermal amplification strategy used is not reliant upon expensive equipment it is particularly suited for "front line" diagnosis of SVD in modestly equipped laboratories, in field stations or in mobile diagnostic units.
Blomström
Anne-Lie
AL
Department of Biomedical Sciences and Veterinary Public Health, Section of Parasitology and Virology, Swedish University of Agricultural Sciences, S-750 07 Uppsala, Sweden. Anne-Lie.Blomstrom@bvf.slu.se
Hakhverdyan
Mikhayil
M
Reid
Scott M
SM
Dukes
Juliet P
JP
King
Donald P
DP
Belák
Sándor
S
Berg
Mikael
M
D000818:Animals; D029822:Enterovirus B, Human / Q000302:isolation & purification*; D012367:RNA, Viral / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013552:Swine; D013555:Swine Vesicular Disease / Q000175:diagnosis* / Q000821:virology
D012367:RNA, Viral