Detection of Viable Mycobacterium paratuberculosis in UK Retail Pasteurised Milk Using Phage-PCR.

Introduction:  Mycobacterium paratuberculosis (MAP) causes Johne's disease, a chronic enteritis of ruminants, and has been implicated in the development of Crohn's disease. Milk is a major vertical transmission route within herds and approximately 40-60% of herds in developed countries are infected. Commercial pasteurisation processes do not completely inactivate MAP; therefore, pasteurised milk has been highlighted as a key vector for entry of MAP into the human food chain. The slow growth of this organism prevents the use of standard culture; therefore, the use of alternative rapid methods is required to detect its presence.

Purpose:  This study completed a survey of commercial, pasteurised, semi-skimmed milk using a rapid detection method (phage-PCR).

Methods:  Samples of retail semi-skimmed milk (385) were purchased from local retailers. Viable Mycobacteria were detected using a previously described phage-PCR method. The presence of MAP in any plaque positive samples was confirmed by PCR amplification of IS900.

Results:   The phage-PCR assay detected viable MAP in 10.3% (38/368) of the pasteurised milk samples. Only 1.1% of the samples contained more than 10 detectable MAP cells; 3.5% of the samples contained more than two cells (1.1% with >10 and 2.4% with 3-9). The majority of samples (6.8%) contained only one or two detectable cells. Within the main survey, MAP was detected in a small number of filtered milk samples. A further survey, focusing on filtered milk alone, found that Mycobacteria can be detected in this type of milk at a high frequency.

Significance:  Viable MAP were detected at levels consistent with previous surveys of retail milk by culture or PCR. Detection of very low levels (one to two MAP cells) suggest the rapid phage PCR-method is more sensitive than existing methods. Mycobacteria can also be detected in filtered milk, which is potentially introduced in the cream fraction.