Molecular detection of Mycobacterium bovis in milk of cattle and buffaloes suspected for bovine tuberculosis
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Date
2016
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Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana
Abstract
The aim of this study was to diagnose bovine tuberculosis (TB) in live animals using comparative intradermal tuberculin test (CITT) and bovine Gamma Interferon assay (?-IFN), detection of antibody response and molecular diagnosis of Mycobacterium bovis by multiplex polymerase chain reaction (mPCR) and real time PCR. One hundred forty animals (84 cattles and 56 buffaloes) were selected from an organized dairy farm, while 60 (52 cattle and 8 buffaloes) were from four unorganized dairy farms. The animals of age group between 2-12 years were included in the study. Blood and milk samples were collected from the animals to be tested by CITT, ?-IFN, detection of antibody response against M. bovis and molecular detection of M. bovis. A total of 29 (14.5%) animals were positive by CITT, 42 (21.0%) animals gave inconclusive results and 129 (64.5%) were negative reactors. A total of 23 animals were tested positive by ?-IFN assay (11 cattle, and 12 buffaloes). The mPCR assay targeting simultaneously M. bovis and Mycobacterium tuberculosis complex (MTC) was standardized which detected amplicons of 245bp for MTC and 500bp for M. bovis from the milk samples. One milk sample was positive for M. bovis and two samples for MTC by conventional PCR. Forty four DNA samples of milk were selected from the animals which were positive by either of the test i.e. CITT, ?-IFN assay, antibody kit test, mPCR or positive in more than one test and subjected to Taqman real-time PCR. Two samples which were positive for MTC by mPCR were diagnosis to be positive for M. bovis by this technique. Thus it is evident that to control bovine tuberculosis in a herd, diagnosis should be made by using two or more tests to detect the infected animals and segregate them