Adaptation of an Invader assay for the detection of African swine fever virus DNA.
A closed tube isothermal Invader assay (Third Wave Technologies Inc., Madison, Wisconsin, USA) was adapted for the detection of African swine fever virus (ASFV) DNA. Several ASFV Invader assays were designed successfully and tested on a real-time PCR instrument (iCycler, BioRad). The assay exhibiting the lowest signal/noise ratio (VP73 ASFV Invader Assay) was analysed further using serial 10-fold dilutions of Lisbon 60 ASFV viral genome. The assay sensitivity was determined to be in the order of 2500 copies of ASFV DNA and showed a dynamic range of 4 logs, from 2.5x10(6) to 2500 copies. The high specificity of the test was demonstrated by the lack of cross-reactivity to the clinically similar but heterologous virus, classical swine fever virus. The sensitivity of the Invader assay is sufficient for the testing of acutely infected viremic animals in which the viral load will be high. The robustness and ease of use of the ASFV Invader assay, combined with the possibility to run and read the assay using simple and relatively inexpensive equipment, makes it suitable for laboratories lacking containment facilities and/or real-time PCR instrumentation or on a regional basis for on-site diagnosis close to putative sites of ASFV outbreaks.
Hjertner
Bernt
B
Department of Veterinary Science, Queen's University Belfast, Stormont, Belfast BT4 3SD, UK. b.hjertner@qub.ac.uk
Meehan
Brian
B
McKillen
John
J
McNeilly
Francis
F
Belák
Sándor
S
D000358:African Swine Fever Virus / Q000235:genetics*; D001483:Base Sequence; D004279:DNA, Viral / Q000032:analysis*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D004279:DNA, Viral