Abstract: Objective To investigate the prevalence status of non-tuberculous mycobacteria(NTM) in old town of Guangzhou during the last 10 years.Methods One thousand and fifty-eight samples of NTM confirmed by culture and strain identification from Guangzhou Research Institute for Tuberculosis Prevention and Treatment were analyzed.Results Seven thousand five hundred and forty-one samples were performed mycobacterium culture and identified between Mycobacterium tuberculosis(Mtb) and NTM, in which 20.00% (1508/7541) of samples were NTM and 228 samples were positive twice by culture. Samples which presented over 1 time of culture positive and same strain identification in one patient in a year were analyzed as one sample, 6797 samples were mycobacterium, in which 1120 were NTM. Constituent ratio of NTM rose to 24.22% (232/958) in 2012 from 12.87% (135/1049) in 2003. Seven hundred and sixty-two of 1120 samples were performed strain identification and 22 species were found. The top of 11 species in NTM constituent ration included 43.18% (329/762) of M. chelonae-abscessus complex, 18.24% (139/762) of M. avium-intracellulare complex, 8.53% (65/762) of M. gordonae, 6.96% (53/762) of M. fortuitum, 4.33% (33/762) of M. scrofulaceum, 3.67% (28/762) of M. kansasii, 3.15% (24/762) of M. smegmatis, 2.62% (20/762) of M. nonchromogenicum, 1.57% (12/762) of M. szulgal, 1.57% (12/762) of M. simiae, and 1.57% (2/762) of M. malmoense. 60.23% (459/762) were male and 39.77% (303/762) were female. The age groups of the highest load rate of bacteria for both male and female was 25-<35 years old, 16.99% (78/459) and 20.13% (61/303), respectively. One hundred and eighteen samples were confirmed as NTM containing 11 species, in which included 66.95% (79/118) of M. chelonae-abscessus complex, 16.95% (20/118) of M. avium-intracellulare complex, 4.24% (5/118) of M. fortuitum, 3.39% (4/118) of M. kansasii, 2.54% (3/118) of M. gordonae, 1.69% (2/118) of M. smegmatis, 0.85% (1/118) of M. scrofulaceum, 0.85% (1/118) of M. nonchromogenicum, 0.85% (1/118) of M. simiae, 0.85% (1/118) of M. simiae, and 0.85% (1/118) of M. chelonae- fortuitum.Conclusion Guangzhou is a high-prevalence area of NTM, and M. chelonae-abscessus complex and M. avium-intracellulare complex are main pathogenic bacteria. Patients with NTM are younger and on rise. It is more important for acid-fast bacillus culture and strain identification to make differential diagnosis and treatment.