The non-tuberculous mycobacterium (NTM) is a very troublesome opportunistic pathogen, placing a heavy burden on public health. The pathogenesis of NTM pulmonary infection is not well-revealed yet, and its diagnosis is always challenging. This study aimed to use a comprehensive proteomics analysis of plasma exosomes to distinguish patients with rapidly growing NTM M. abscessus (MAB), slowly growing NTM M. intracellulare (MAC), and Mycobacterium tuberculosis (MTB). The identified protein components were quantified with label-free proteomics and determined with a bioinformatics analysis. The complement and coagulation were significantly enriched in patients with Mycobacterium infection, and a total of 24 proteins were observed with up-regulation, which included C1R, C1S, C2, MASP2, C4B, C8B, C9, etc. Of them, 18 proteins were significantly up-regulated in patients with MAB, while 6 and 10 were up-regulated in patients with MAC or MTB, respectively. Moreover, MAB infection was also related to the HIF-1 signaling pathway and phagosome processes, and MTB infection was associated with the p53 signaling pathway. This study provided a comprehensive description of the exosome proteome in the plasma of patients infected with MAB, MAC, and MTB and revealed potential diagnostic and differential diagnostic markers.

We performed a prospective study to evaluate the diagnostic accuracy of nucleotide matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) in identifying nontuberculous mycobacterium (NTM) from clinical respiratory samples. A total of 175 eligible patients were prospectively enrolled, including 108 patients diagnosed with NTM pulmonary disease (NTM-PD) and 67 control patients with other diseases. All specimens were subjected to acid-fast staining, liquid culture combined with MPT64 antigen detection, and a nucleotide MALDI-TOF MS assay. NTM cultures were also subjected to the MeltPro Myco assay for species identification. Altogether, the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of nucleotide MALDI-TOF MS were 77.8% (95% CI: 68.6-85.0%), 92.5% (82.8-97.2%), 94.4% (86.8-97.9%), and 72.1% (61.2-81.0%), respectively; these results were not statistically different from the results of culture + MPT64 antigen testing (75.0% [65.6-82.6%], 95.5% [86.6-98.8%], 96.4% [89.2-99.1%], and 70.3% [59.7-79.2%], respectively). In the identification of NTM species, of the 84 nucleotide MALDI-TOF MS positive samples, 77 samples (91.7%) were identified at the species level. Using culture + MeltPro Myco assay as the reference standard, nucleotide MALDI-TOF MS correctly identified 77.8% (63/81) of NTM species. Our results demonstrated that the nucleotide MALDI-TOF MS assay was a rapid single-step method that provided the reliable detection of NTM and identification of NTM species. This new method had the same sensitivity and specificity as the culture + MPT64 antigen method, but was much more rapid.