Detection of African swine fever virus by a biotinylated DNA probe: assay on cell cultures and field samples.
African swine fever virus was detected in various samples using a molecular hybridization technique. A fragment located in a constant area of the viral genome was biotin-labelled. This probe, when present at a concentration of 100 ng/ml of the hybridization solution, could detect 10 pg of target DNA immobilized on nitrocellulose with cellular DNA and RNA. The virus was evidenced after being passaged on monkey kidney cells, either 8 h post-inoculation (pi) if the multiplicity of infection (MOI) was at least 1 hemadsorbing unit (HAd) per cell, or 24 h later if the inoculum was diluted up to 10(-3) HAd per cell. When passaged on pig leukocytes with a MOI of 0.1 HAd per cell, the virus was evidenced 12 h pi, or 24 h pi with a MOI of 10(-2) HAd per cell. The probe did not hybridize with another DNA virus passaged on cells, neither did it react with non-infected blood or ham, but did so if African swine fever virus was resuspended with the samples. The spleen from uninfected pig and the lymph nodes from a pig which had died from hog cholera were found to be negative, whereas the spleen from a pig which had died of African swine fever was positive. These samples were also tested with a 32P-labelled probe whose sensitivity was 10-fold higher. A non-radioactive probe could be used both for the sensitive and specific diagnosis of African swine fever and the detection of the virus in an epidemiological survey.
Petit
F
F
Ecole Nationale Vétérinaire, Service des Maladies Contagieuses, Toulouse, France.
Boucraut-Baralon
C
C
Py
R
R
Benazet
F
F
Picavet
D P
DP
Chantal
J
J
D000357:African Swine Fever / Q000382:microbiology*; D000358:African Swine Fever Virus / Q000235:genetics / Q000302:isolation & purification*; D000818:Animals; D001710:Biotin; D002460:Cell Line; D015342:DNA Probes; D004279:DNA, Viral / Q000032:analysis*; D008198:Lymph Nodes / Q000382:microbiology; D009693:Nucleic Acid Hybridization; D013154:Spleen / Q000382:microbiology; D013552:Swine
D015342:DNA Probes; D004279:DNA, Viral; D001710:Biotin