DETECTION OF MYCOBACTERIUM-BOVIS IN MILK BY POLYMERASE-CHAIN-REACTION

A simple method was developed for DNA extraction of Mycobacterium bovi s in milk and further detection of the bacterium by polymerase chain r eaction (PCR). Milk previously seeded with M. bovis was used as the st arting material. The procedure involved overnight digestion of a milk sample with proteinase K at 56 degrees C and phenol extraction, follow ed by ethanol precipitation and PCR. The amplification pattern obtaine d was analyzed with primers BW8-BW9 which amplify a 248 bp in strains of M. bovis. By using the BW8-BW9 primers, 10(3) CFU were detected on silver-stained PAGE gels. The procedure was validated by PCR analysis of milk in tuberculin-positive animals. It is anticipated that this me thod can be used for routine diagnosis of M. bovis in milk samples.