Detection of African swine fever virus by loop-mediated isothermal amplification.
A loop-mediated isothermal amplification (LAMP) assay was developed for the detection of African swine fever virus (ASFV). This assay targets the topoisomerase II gene of ASFV and its specificity was confirmed by restriction enzyme digestion of the reaction products. The analytical sensitivity of this ASFV LAMP assay was at least 330 genome copies, and the test was able to detect representative isolates of ASFV (n=38) without cross-reacting with classical swine fever virus. The performance of the LAMP assay was compared with other laboratory tests used for ASF diagnosis. Using blood and tissue samples collected from pigs experimentally infected with ASFV (Malawi isolate), there was good concordance between the LAMP assay and real-time PCR. In addition to detecting the reaction products using either agarose gels or real-time PCR machines, it was possible to visualise dual-labelled biotin and fluorescein ASFV LAMP amplicons using novel lateral flow devices. This assay and detection format represents the first step towards developing a practical, simple-to-use and inexpensive molecular assay format for ASF diagnosis in the field which is especially relevant to Africa where the disease is endemic in many countries.
James
Heather E
HE
Institute for Animal Health, Ash Road, Pirbright, Surrey GU24 0NF, United Kingdom.
Ebert
K
K
McGonigle
R
R
Reid
Scott M
SM
Boonham
Neil
N
Tomlinson
Jennifer A
JA
Hutchings
Geoffrey H
GH
Denyer
Mick
M
Oura
Chris A L
CA
Dukes
Juliet P
JP
King
Donald P
DP
D000357:African Swine Fever / Q000175:diagnosis*; D000358:African Swine Fever Virus / Q000235:genetics / Q000302:isolation & purification*; D000818:Animals; D006692:Classical Swine Fever Virus / Q000235:genetics; D003429:Cross Reactions; D004250:DNA Topoisomerases, Type II / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics / Q000302:isolation & purification*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D012680:Sensitivity and Specificity; D013552:Swine; D013696:Temperature; D014764:Viral Proteins / Q000235:genetics
D004279:DNA, Viral; D014764:Viral Proteins; D004250:DNA Topoisomerases, Type II