Quantitative Real-Time PCR for the detection of Mycobacterium avium subsp. paratuberculosis in bovine fecal samples and liquid culture samples

摘要

The objectives of this study were: to evaluate a novel fecal DNA extraction method, combined with a quantitative real-time PCR (QRTPCR) amplification method (VetAlert~(TM), Tetracore, Inc.) for the detection of MAP in bovine fecal samples from naturally infected cows, and 2) evaluate the use of the QRTPCR for confirmation of liquid culture of MAP. Fecal samples from multiple cattle shedding MAP at various levels were tested. The samples were processed with a DNA extraction and purification method which employed bead-beating and chaotropic solid phase extraction. Fecal DNA samples were then subjected to the QRTPCR amplification procedure, which is a real-time fluorescent probe hydrolysis assay.