No abstract

Aims: One possible route of transmission of Mycobacterium paratuberculosis from cattle to humans is via contaminated water supplies. The aim of this work was to determine whether this organism can survive standard water treatment processes. Methods and Results: Two strains of M. paratuberculosis (bovine strain, NCTC 8578 and human strain Linda, ATCC 43015) were subjected to various chlorine concentrations (0á5, 1á0 and 2á0 lg ml ±1 ) for 15 and 30 min. Chlorine test solutions were made up in two types of water, sterile water that had been deionized and subjected to reverse osmosis (DRO) and DRO water containing MgCl 2 , CaCl 2 , NaHCO 3 and bovine serum albumin (0á3% w/v), the latter to mimic conditions the organism would experience in commercial water treatment operations. Conclusions: The data showed that when initial inoculum levels were high (10 6 cfu ml ±1 ) neither M. paratuberculosis strain was completely killed at the free chlorine concentrations and contact times applied. Log 10 reductions in the range 1á32±2á82 were observed. The greatest log 10 reduction in cell numbers (2á82 and 2á35 for the bovine and human strains, respectively) was observed at the highest chlorine concentration (2 lg ml ±1 ) and longest contact time (30 min). Signi®cance and Impact of the Study: This work highlights the need for further research into the survival of M. paratuberculosis during water treatment.

Mycobacterium avium subsp. paratuberculosis is the known cause of Johne's disease of both domestic and wild ruminants and has been implicated as a possible cause of Crohn's disease in humans. The organism is shed in the feces of infected animals and can survive for protracted periods in the environment and hence could be present in catchment areas receiving agricultural runoff. A limited survey was undertaken in Northern Ireland to test for M. avium subsp. paratuberculosis in untreated water entering nine water treatment works (WTWs) over a 1-year period. Three detection methods were employed, viz., immunomagnetic separation-PCR and culture on Herrold's egg yolk medium (HEYM) and BACTEC 12B medium, the latter both supplemented with mycobactins. Of the 192 untreated water samples tested, 15 (8%) tested M. avium subsp. paratuberculosis positive by one or more of the three detection methods. M. avium subsp. paratuberculosis was successfully isolated from eight untreated water samples, three by BACTEC culture and five by culture on HEYM. Although the highest incidence of M. avium subsp. paratuberculosis was found in spring, overall, there was no statistically significant difference between the seasons. No significant correlation was found between numbers of coliforms or fecal coliforms and the presence of M. avium subsp. paratuberculosis. In general, a higher incidence of M. avium subsp. paratuberculosis was found in untreated water entering those WTWs that had a high mean water pH value over the sampling period. This work indicates the need to determine the efficacy of water treatment processes to either kill or remove M. avium subsp. paratuberculosis from untreated water and the possible risks posed by contact with recreational water sources.

Background: Interactions between Mycobacterium avium subsp. paratuberculosis (Map) and freeliving protozoa in water are likely to occur in nature. The potential impact of ingestion of Map by two naturally occurring Acanthamoeba spp. on this pathogen's survival and chlorine resistance was investigated.

Aims: To develop a sensitive detection method for Mycobacterium avium ssp. paratuberculosis (Map) in water by modifying and optimizing an existing immunomagnetic separation polymerase chain reaction (IMS-PCR) technique. Methods and Results: Sterile distilled water (50 ml) spiked with 10 6 Map ml )1 was subjected to either filtration (0AE45 lm pore size) followed directly by IS900 PCR (method 1) or centrifugation (2500 g for 20 min) followed by IMS and IS900 PCR (method 2). Method 2 permitted the detection of Map, whereas method 1 did not. Method 2 was then optimized by adding different concentrations of Tween 80 (0AE05, 0AE1, 0AE2, 0AE4 and 0AE6% v/v) to water samples spiked with Map (10 6 -1 CFU ml )1 ) prior to centrifugation, and assessing the impact of this action on the detection sensitivity of subsequent IMS-PCR. The optimum Tween 80 concentration was found to be 0AE4%, which permitted the detection of 10 Map CFU ml )1 in spiked water samples by IMS-PCR.Conclusions: This method will be used to determine the incidence of Map in water destined for domestic use in future studies. Significance and Impact of the Study: A sensitive method for the detection of Map in water involving addition of 0AE4% Tween 80, centrifugation and IMS-PCR was developed.