COMPARISON OF METHODS FOR ISOLATION OF MYCOBACTERIA FROM WATER

Twelve methods for the isolation of mycobacteria were compared by appl ying them in parallel to 26 samples of surface water and 109 samples o f treated water. Each method was defined by a particular combination o f decontamination method, growth medium, and incubation temperature. F or the decontamination of surface water, we used cetylpyridinium chlor ide (CPC) (30 min, 0.05%), as well as sample preincubation in tryptic soy broth (TSB) followed by decontamination with a cocktail of NaOH, c ycloheximide, and malachite green. Treated water was decontaminated wi th 0.005 and 0.05% CPC (30 min). After enrichment by filtration, all s amples were incubated on Lowenstein-Jensen medium (LJ), Ogawa egg yolk medium (OEY), and Ogawa whole-egg medium containing ofloxacin and eth ambutol (OEOE) at temperatures of 30 and 37 degrees C. The efficacy of each method was determined by calculating the positivity rate, negati vity rate, contamination rate, mean number of mycobacterial colonies g rown, and mean number of different mycobacterial strains isolated. The last value was determined by subjecting the isolates to PCR restricti on analysis and mycolic acid thin-layer chromatography. Statistical an alysis demonstrated that both the TSB method and 0.05% CPC were approp riate for the decontamination of surface water, Decontamination with 0 .005% CPC was best for treated water. The results for incubation on LJ were at least equal to those for incubation on OEY and always superio r to the results with OEOE, At an incubation temperature of 30 degrees C, all methods achieved higher yields than at 37 degrees C.