DIRECT DETECTION OF BOVINE LEUKEMIA-VIRUS INFECTION - PRACTICAL APPLICABILITY OF A DOUBLE POLYMERASE CHAIN-REACTION

A double polymerase chain reaction (PCR) assay has been devised for th e direct detection of bovine leukemia virus (BLV). The assay was direc tly performed on blood leukocytes, avoiding the DNA-purification proce dures. The PCR products were identified by gel-electrophoresis and the specificity of the test was confirmed by hybridization with a biotiny lated oligonucleotide probe. When testing the sensitivity of PCR, less than eight genome copies of the provirus were detected in the backgro und of two million negative lymphocytes. In a BLV infected herd 22 ani mals of various age groups were examined by the indirect (serological) diagnostic tests of agar-gel immunodiffusion and indirect ELISA as we ll as by the direct detection method of PCR. The tests were repeated a t monthly intervals on five occasions. When examining the specimens fr om cows and heifers, a close agreement was found between the results o f the various methods. The newborn calves, which were the offspring of BLV infected mothers, were consequently negative in PCR throughout th e experimental period. However, in the indirect tests the calves were positive during the first samplings and became negative only around fo ur months of age. Since the indirect tests can not discriminate infect ion from colostral immunity, PCR proved to be a useful complementary a ssay for the safe diagnosis of BLV infection in young calves.