An inter-laboratory ring trial for the detection and isolation of Mycobacterium avium subsp. paratuberculosis from raw milk artificially contaminated with naturally infected faeces Original Research Article

There is a need for standardised, robust, reproducible molecular and culture methods to achieve clarification of the inactivation of Mycobacterium avium subsp. paratuberculosis (Map), the causative microbial agent of Johneʹs disease, in (faecally) contaminated milk and other food products such as meat. This study assessed the performance of a commercially available Map DNA extraction kit for milk (Adiapure®) and accompanying PCR detection kit (Adiavet®) alongside ‘in-house’ molecular and culture methods in an inter-laboratory ring trial using raw milk spiked with Map-infected faeces. The combined Adiapure-Adiavet® Map DNA extraction and detection kit consistently detected 30 copies of IS900 (equivalent to approximately 2 cells) ml−1 raw milk, when used in four different laboratories. Improvements in sensitivity and ease of use for ‘in-house’ Map detection were observed when the Adiapure extraction kit was combined with ‘in-house’ detection assays. Detection by real-time PCR methods, using the commercial extraction and detection systems, resulted in an overall detection rate of 100%, 90%, 85% and 25% for respective Map concentrations of 300, 30, 3 and 0.3 copies of IS900 ml−1 raw milk.