Direct use of cell lysates in PCR-based diagnosis of bovine leukemia virus infection.
Polymerase chain reaction (PCR) has been used for direct detection of bovine leukemia virus (BLV) proviral DNA in cattle, but it is still mainly used for experimental research. One bottleneck for routine diagnosis of BLV by PCR has always been the isolation and purification of DNA. We compare the use of not purificated with highly-purified DNA in the PCR-based diagnosis of BLV infection. DNA extracted from whole blood by chloroform extraction (CP-DNA) and DNA prepared only by osmotic shock, washing, heating and freezing procedures (RPoS-DNA), were utilized. Fifteen cattle well characterized serologically were investigated for BLV-provirus with PCR using this different DNA preparations. With both methods all but one investigated animal were correctly identified. It was estimated that in case of CP-DNA PCR 10 BLV-provirus copies were sufficient to obtain a positive result. The sensitivity of RPoS-DNA PCR was similar. Because of the greater practicability of the latter technique we used it in a small field study with ten cattle. All serologically positive animals were correctly identified by the PCR. In addition one seronegative animal was found to carry BLV-provirus. Therefore RPoS-DNA PCR might be a good tool for the routine diagnosis of BLV-infected cattle.
Fechner
H
H
Institute of Virology, Faculty of Veterinary Medicine, Free University of Berlin, Germany.
Kurg
A
A
Blankenstein
P
P
Mewes
G
G
Geue
L
L
Albrecht
C
C
Ebner
D
D
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D002417:Cattle; D017931:DNA Primers; D004279:DNA, Viral / Q000032:analysis; D016583:Enzootic Bovine Leukosis / Q000175:diagnosis* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D001909:Leukemia Virus, Bovine / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000662:veterinary; D015203:Reproducibility of Results
D000914:Antibodies, Viral; D017931:DNA Primers; D004279:DNA, Viral