Isolation Phenotypic and Genotypic Characterization of Non Tuberculous Mycobacteria isolated from Clinical Specimens

Abstract

newlineNon tuberculous mycobacteriais an opportunistic pathogen reported in the recent years among both immunodeficient and immunocompetent hosts. The aim of this study is to know the incidence of NTM species in pulmonary and extrapulmonary tuberculosis cases in a tertiary care hospital in Pondicherry and to evaluate different phenotypic and genotypic methods in identifying NTM and finally to determine their DST patterns. According to PCR-RFLP analysis of hsp65 gene, 253 (56.22%) were identified as M.abscessus followed by M.simiae 86 (19.11%), 58 (12.88%) as M.fortuitum, 49 (10.88%) M.chelonae and 4(0.88) were identified as M.gordonae. PCR RFLP of 16S-23S rRNA ITS gene results were contradictory, of the 452 isolates 2 isolates were identified as M.tb complex, ITS gene of M.abscessus has no restriction sites for both BstEII and HaeIII enzymes. The ITS gene of 86 isolates of M.simiae produced restriction fragments with HaeIII and the gene was not digested with BstEII enzyme.Hsp5 gene sequencing of the 450 NTM species, 221 were classified as M.abscessus sub species abscessus, 32 were classified as M.abscessus, 86 were determined as M.simiae, 58 as M.fortuitum, 49 were identified as M.chelonae and 4 isolates as M.gordonae. All the isolates of NTM were 100% resistant to first line antitubercular drugs tested. M.simiae exhibited 100% resistance to doxyxycline and Sulfamethoxazole and hundred percent sensitive to the Aminoglycosides like Amikacin, macrolides like clarithromycin quinolones like moxifloxacin, levofloxacin, oxazolidinone like linezolid and cotimoxazole. And M.fortuitum exhibited 100% sensitivity to Amikacin and clarithromycin. Overall rapid isolation and differentiation to the level of species using molecular techniques had guided in targeted therapy and management of diseases with NTM.