Identification of Nontuberculous Mycobacteria Species Isolated from Water Samples Using Phenotypic and Molecular Methods and Determination of their Antibiotic Resistance Patterns by E- Test Method, in Isfahan, Iran

INTRODUCTION Many studies have shown epidemiological links between strains isolated in tap water, and those isolated from patients. Molecular methods linked to PCR are more reliable and faster for identification of non- tuberculous mycobacteria (NTM). In this study molecular methods were used for identification and typing of NTM. MATERIALS AND METHODS Five hundred ml of 85 water samples was passed through 0.45 μm filters. The filters were transferred directly onto 7H10 Middle Brook solid media, containing 15% OADC. PCR for 16S rRNA was done and the PCR product (1500 bp) was sequenced. PRA of the hsp65 gene was investigated to identify the species of isolates. For evaluation of susceptibility of NTM to antimycobacterial agents, E-test method was used. RESULT The genus of 26 isolated NTM was confirmed by 16s rRNA sequence based method. Nineteen isolates of Mycobacteria were differentiated using hsp65 genes PRA. The dominant isolates were M. fortuitum (26.7%), M. chelonae like organism (13.3%) and M. mucogenicum (13.3%). Seventy one percent of NTM species were resistant to isoniazid, 64% to rifampin, 57% to ethambutol, 35% to tetracycline, 14 % to azithromycin and 7.1 % to amikacin. CONCLUSION The results showed that E-test method is not a proper technique for antimycobacterial assay because some NTM species are slow in growing and have no growth on Muller Hinton agar. Regarding the 16S rRNA sequence analysis, the identification of isolates was restricted to the genus level, because 99% similarity within 16S rRNA of two isolates may or may not determine the same species.