The evaluation of different molecular methods for rapid identification of non-tuberculous Mycobacterium

Abstract

Objective To evaluate three molecular methods for rapid identification of nontuberculous Mycobacterium(NTM).Methods Forty-one clinical NTM isolates were collected and 16S rRNA gene sequencing was used as the standard method for NTM identification.Meanwhile,the restriction fragment length polymorphism of hsp65 PCR-RFLP and hsp65 gene sequencing were used to identify NTM strains and compared with 16S rRNA gene sequencing.Results The results of 16S rRNA sequencing showed that there were nine Mycobacterium chelonae complex strains,seven Mycobacteriumfortuitum strains,seven Mycobacterium intracellulare strains,three Mycobacterium avium strains,three Mycobacterium kansasii complex strains, three Mycobacterium smegmatis strains, three Mycobacterium terrae strains, two Mycobacterium phlei strains,two Mycobacterium nonchromogenicum strains,one Mycobacterium scrofulaceum strain and one Mycobacterium arupense strain.Compared with 16S rRNA gene sequencing,hsp65 PCR-RFLP could identify nine Mycobacterium chelonae complexes and three Mycobacterium kansasii complexes to subspecies Mycobacterium abscessus and Mycobacterium kansasii,respectively; One Mycobacterium fortuitum strain and one Mycobacterium nonchromogenicum strain were different from 16S rRNA gene sequencing results ,but other isolates were the same.The coincidence was 95.1%.By hsp65 gene sequencing,only one identification of Mycobacterium hiberniae strain was different from 16S rRNA gene sequencing and the coincidence was 97.6%.And hsp65 gene sequencing could further identify nine Mycobacterium chelonae complexes and three Mycobacterium kansasii complexes to subspecies Mycobacterium abscessus and Mycobacterium kansasii,respectively.Conclusions All three molecular methods can identify NTM strains rapidly.Compared with 16S rRNA gene sequencing,hsp65 gene sequencing and hsp65 PCR-RFLP are easier to identify clinical common NTM strains(such as Mycobacterium kansasii and Mycobacterium abscessus),and can be widely used in clinical practice. Key words: Mycobacterium; RNA,ribosomal,16S; Bacterial proteins; Heat-shock proteins; Polymerase chain reaction; Polymorphism,restriction fragment length