Development of a rapid molecular assay for specific identification of Mycobacterium avium and Mycobacterium intracellulare.

In genetic testing of non-tuberculous mycobacteria, false positives due to cross-reactions and differences in detection performance among strains are problematic. A rapid genetic testing method with high specificity to distinguish only Mycobacterium avium and Mycobacterium intracellulare and high detection performance against genetic diversity among strains is desirable. We identified completely novel genes (MAA44156_02837 for M. avium and KN251_15665 for M. intracellulare) showing strong identity with M. avium and M. intracellulare respectively. We then developed a new M. avium/M. intracellulare simultaneous detection assay targeting these genes and evaluated their specificity and sensitivity. To evaluate cross-reactivity, the newly developed assay correctly identified all Mycobacterium spp. type strains, except for one examination of a duplicated evaluation with a Mycobacterium kyorinense type strain as a false positive at McFarland 1.0. All M. avium-positive and M. intracellulare-positive clinical isolates used in this study were accurately detected using the newly developed assay. The limits of detection for the newly developed assay were estimated to be 100 CFU/mL for M. avium and 30 CFU/mL for M. intracellulare. In addition, the detection performance was maintained even in samples containing both M. avium and M. intracellulare. The M. avium/M. intracellulare simultaneous detection assay developed in this study demonstrated high specificity, detection performance, and sensitivity for detecting M. avium and M. intracellulare.