Yuko Kazumi1, Satoshi Mitarai2. 1. Molecular Epidemiology Division, Department of Mycobacterium Reference and Research, Research Institute of Tuberculosis, Tokyo, Japan. 2. Bacteriology Division, Department of Mycobacterium Reference and Research, Research Institute of Tuberculosis, Tokyo, Japan. Electronic address: mitarai@jata.or.jp.
Abstract
BACKGROUND: Conventional biochemical tests are the standard for the identification of Mycobacterium species, but molecular identifications are becoming more prevalent. The rpoB gene encodes the β-subunit of RNA polymerase and is utilized for the identification of Mycobacterium species. In the present study, a stepwise Mycobacterium species identification algorithm using the 16S rRNA encoding gene and rpoB analysis was evaluated for its effectiveness. METHODS: A total of 172 clinical Mycobacterium isolates were tested, and concordant results were obtained with 108 strains by using the conventional method and molecular methods (AccuProbe or DDH method). RESULTS: In these 108 strains, 4 strains were identified by 16S rRNA gene analysis, but rpoB indicated no identical Mycobacterium species with more than 99% similarity. The remaining 64 strains were not identified by conventional method and commercial kits. Forty-two showed concordant results with 16S rRNA and rpoB analysis, and 13 strains were identified by 16S rRNA gene analysis although rpoB indicated no identical Mycobacterium species. On the other hand, 4 strains included 2 strains of Gordona and 2 strains of M. celatum type II which were identified by rpoB but not by 16S rRNA gene analysis. Finally, 5 strains could not be identified by analysis of either gene. The rpoB analysis can differentiate M. kansasii from M. gastri; M. malmoense from M. szulgai; M. abscessus from M. chelonae; M. peregrinum from M. septicum; M. porcinum from M. fortuitum; and M. farucinogense from M. senegalense-pairs that are not differentiated by 16S rRNA analysis. Additionally, Nocardia asteroids, Rhodococcus equi, Gordona aichiense, G. aurantiaca, G. bronchialis and G. terrae are able to be analyzed by using rpoB. CONCLUSIONS: The 16S rRNA gene identification is a rapid and prevalent method but still has some limitations. Therefore, the stepwise combination of rpoB with 16S rRNA gene analysis is an effective system for the identification of Mycobacterium species.
BACKGROUND: Conventional biochemical tests are the standard for the identification of Mycobacterium species, but molecular identifications are becoming more prevalent. The rpoB gene encodes the β-subunit of RNA polymerase and is utilized for the identification of Mycobacterium species. In the present study, a stepwise Mycobacterium species identification algorithm using the 16S rRNA encoding gene and rpoB analysis was evaluated for its effectiveness. METHODS: A total of 172 clinical Mycobacterium isolates were tested, and concordant results were obtained with 108 strains by using the conventional method and molecular methods (AccuProbe or DDH method). RESULTS: In these 108 strains, 4 strains were identified by 16S rRNA gene analysis, but rpoB indicated no identical Mycobacterium species with more than 99% similarity. The remaining 64 strains were not identified by conventional method and commercial kits. Forty-two showed concordant results with 16S rRNA and rpoB analysis, and 13 strains were identified by 16S rRNA gene analysis although rpoB indicated no identical Mycobacterium species. On the other hand, 4 strains included 2 strains of Gordona and 2 strains of M. celatum type II which were identified by rpoB but not by 16S rRNA gene analysis. Finally, 5 strains could not be identified by analysis of either gene. The rpoB analysis can differentiate M. kansasii from M. gastri; M. malmoense from M. szulgai; M. abscessus from M. chelonae; M. peregrinum from M. septicum; M. porcinum from M. fortuitum; and M. farucinogense from M. senegalense-pairs that are not differentiated by 16S rRNA analysis. Additionally, Nocardia asteroids, Rhodococcus equi, Gordona aichiense, G. aurantiaca, G. bronchialis and G. terrae are able to be analyzed by using rpoB. CONCLUSIONS: The 16S rRNA gene identification is a rapid and prevalent method but still has some limitations. Therefore, the stepwise combination of rpoB with 16S rRNA gene analysis is an effective system for the identification of Mycobacterium species.