Objective To determine the clinical significance of nontuberculous mycobacteria (NTM) isolated from respiratory specimens. Methods Clinical data of patients with NTM strains isolated from the respiratory tract between January 2014 and February 2017 were retrospectively analyzed. Clinical significance of NTM isolated strains was evaluated based on diagnostic criteria of NTM pulmonary diseases from American Thoracic Society (ATS). Quantitative data of two groups were analyzed by independent t test. Categorical data were analyzed by Pearson χ2 test or Fisher exact test. Results Totally 352 NTM strains from 257 patients had been isolated between January 2014 and February 2017. Among 15 identified NTM species, M. intracellular (51.7%, 182/352), M. abscessus (25.6%, 90/352), and M. avium (10.8%, 38/352) were predominant. Of the 157 patients with full clinical data involved in the analysis, 58 (36.9%) patients were determined to have definite disease, and 34 patients (21.7%) were designated as probable disease candidates, and 16 (10.2%) patients were regarded as uncertain disease, and 49(31.2%) patients were diagnosed as unlikely disease. The age of 58 patients with definite disease was (63.9±12.7) years, and 48.3% (28/58) were female. M. intracellulare (55.2%, 32/58) was the main cause of pulmonary NTM disease, followed by M. abscessus (25.9%, 15/58) and M. avium (12.1%, 7/58), while other NTM species only accounted for 6.8% (4/58). Definite cases with M. intracellulare, M. abscessus, M. avium, M. kansasii, M. marseillense, and M. columbia accounted for 35.2% (32/91), 57.7% (15/26), 7/15, 2/3, 1/3 and 1/2, respectively, among patients with corresponding isolations, while patients with other species of isolation did not meet the diagnostic criteria. Patients with clinical significant isolation of NTM were older than those without clinical significance (χ2=3.603, P=0.000), and proportion of anti-acid staining positivity of patients with clinical significance was higher than that of patients without clinical significance (χ2=18.815, P=0.000). The proportion of M. abscessus in patients with clinical significance was higher than that in patients without clinical significance (χ2=6.313, P=0.012). However, there was no significant difference between the two groups in the isolation of M. Gordon (Fisher exact test, P=0.028). The proportion of M. abscessus lung disease in women was 11/15, which was higher than that of M. intracellulare lung disease (41.5%, 17/41), and the difference between the two groups was statistically significant (χ2=4.462, P=0.035). There was no significant difference of clinical symptoms and underlying diseases in NTM lung disease among different groups (all P>0.05). In these patients with definite disease, 39.7% (23/58) of them manifested the upper lobe cavitary form, 53.4% (31/58) exhibited nodular bronchiectatic form, and only 6.9% (4/58) exhibited unclassified form. The upper lobe cavitary form (43.8%, 14/32) and the nodular bronchiectatic form (53.1%, 17/32) dominated in patients with M. intracellulare lung disease. M. abscessus lung disease was dominated by the nodular bronchiectatic form (11/15) while the upper lobe cavitary form only accounts for 3/15. There was no significant difference of image characteristics between NTM lung disease and other different groups (all P>0.05). Conclusions 36.9% of the patients with NTM isolates met the ATS diagnostic criteria for NTM lung disease. Different species have different clinical significance. M. intracellulare and M. abscessus are the most predominant NTM isolated species that cause NTM lung disease. Majority of patients manifest as the upper lobe cavitary form, followed by the nodular bronchiectatic form. Key words: Respiratory; Nontuberculous mycobacteria; Clinical relevance; Clinical significance