This Master's thesis project was designed to compare molecular techniques with traditional methods of culture processing in detecting Mycobacterium bovis in environmental substrates. A polymerase chain reaction (PCR) assay was applied to environmental substrates previously inoculated with varying concentrations of M. bovis and stored for various times under a range of temperatures. PCR was statistically equivalent to bacterial culture for detection of M. bovis in samples of water and hay but overall more effective than bacterial culture for detection of M. bovis in soil samples. In a second experiment, M. bovis was detected by PCR for up to 11 months in water samples exposed to natural weather conditions. In contrast, M. bovis was not detected by culture longer than 2 months post-inoculation in any substrate. PCR techniques were applied to a final set of environmental substrates that were sampled from cattle farms with previous occurrences of M. bovis infection. However, none of the analyzed samples tested positive for M. bovis DNA.In epidemiologic investigations of TB farms and wildlife sites, PCR-based assays may be useful for parallel testing with bacterial culture to enhance detection of M. bovis in the environment. The data presented supports the fact that M. bovis can persist in the environment long enough to pose a potential infection risk to susceptible animals. Molecular techniques can be a useful and efficient tool in investigations concerning the role of indirect transmission in the epidemiology of bovine tuberculosis.
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