Sensitive detection of African swine fever virus using real-time PCR with a 5' conjugated minor groove binder probe.
The design of a 5' conjugated minor groove binder (MGB) probe real-time PCR assay is described for the rapid, sensitive and specific detection of African swine fever virus (ASFV) DNA. The assay is designed against the 9GL region and is capable of detecting 20 copies of a DNA standard. It does not detect any of the other common swine DNA viruses tested in this study. The assay can detect ASFV DNA in a range of clinical samples. Sensitivity was equivalent to the Office International des Epizooties (OIE) recommended TaqMan assay. In addition the assay was found to have a detection limit 10-fold more sensitive than the conventional PCR recommended by the OIE. Linear range was ten logs from 2x10(1) to 2x10(10). The assay is rapid with an amplification time just over 2h. The development of this assay provides a useful tool for the specific diagnosis of ASF in statutory or emergency testing programs or for the detection of ASFV DNA in research applications.
McKillen
John
J
Veterinary Sciences Division, Agri-Food and Biosciences Institute, Stormont, Belfast BT4 3SD, UK. john.mckillen@afbini.gov.uk
McMenamy
Michael
M
Hjertner
Bernt
B
McNeilly
Francis
F
Uttenthal
Ase
A
Gallardo
Carmina
C
Adair
Brian
B
Allan
Gordon
G
D000357:African Swine Fever / Q000175:diagnosis* / Q000821:virology; D000358:African Swine Fever Virus / Q000235:genetics / Q000302:isolation & purification*; D000818:Animals; D015345:Oligonucleotide Probes / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013552:Swine; D013997:Time Factors; D014773:Virology / Q000379:methods*
D015345:Oligonucleotide Probes