NTM Identification by MALDI-TOF MS

Species identification was performed for all samples, by using matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS) (Vitek MS Mycobacterium/Nocardia Kit, bioMérieux SA, Marcy L’Étoile, France). Briefly, 1 μL of NTM colonies grown on LJ medium was transferred to a tube containing glass beads and 70% ethanol, vortexed for 15 min, and incubated for additional 10 min at room temperature. The dried pellets were then uniformly dispersed in 70% formic acid and acetonitrile. After centrifugation, 1 μL of the supernatant was transferred to a slide. After drying, 1 μL of CHCA matrix (bioMérieux SA, Marcy L’Eoile, France) was applied and allowed to dry before the analysis. Identification was performed automatically. For calibration and quality control, Escherichia coli (ATCC 8739; American Type Culture Collection, Manassas, VA, USA) was used for each run according to the manufacturer’s protocol [19 (link)].