Application of an F57 Sequence-Based Real-Time PCR Assay for Mycobacterium paratuberculosis Detection in Bulk Tank Raw Milk and Slaughtered Healthy Dairy Cows

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Abstract
A light cycler–based real-time PCR assay that targets the F57 sequence was used to collect data on the prevalence of Mycobacterium avium subsp. paratuberculosis (MAP) in 100 bulk tank raw milk samples and in a population of 101 slaughtered dairy cattle. The assay's reproducible detection limit in total genomic DNA templates isolated from 10-ml samples of MAP-spiked raw milk was 100 cells per ml. Similarly, the evaluation of MAP-spiked bovine feces also demonstrated that the assay had a reproducible detection limit of 100 cells if they were contained within 200 mg of fecal sample material. Among the 100 bulk tank milk samples that were tested, we found 3 samples (3%) to be positive for MAP. In the slaughterhouse part of the study, 8.9% (9 of 101) of the cows were positive for MAP DNA in fecal samples, 4.9% (5 of 101) in mesenteric lymph nodes, 0.9% (1 of 101) in ileum tissue, and 3.6% (3 of 84) in milk. Meanwhile, for 2.9% (3 of 101) of the culled cows, MAP DNA was detected in samples of diaphragmatic muscles.

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