Background and Objectives: Non-tuberculous mycobacteria are one of the major causes of death, which are often a form of progressive lung disease. Diagnosis of NTM infections can be quite challenging. It is well known that the determination of the drug sensitivity of clinical isolates of NTM is one of the key points in the treatment of these diseases. Most NTM clinical isolates are resistant to first-line anti-TB drugs. The aim of this study was to determine the resistance pattern of isolated mycobacterial strains from clinical samples within 2011-2018 to inform and focus the scientific community, especially specialists in this field to the importance of these bacteria of different types of lung disease. Materials and Methods: In total, 15829 samples including different clinical specimens were collected at NRITLD center in Iran. The Drug-resistant tests of first-line anti-TB drugs for INH (isoniazid) and RIF (rifampin) were performed for mycobacterial clinical isolates using MAS-PCR .The genes involved were katG315, inhA for INH and rpoB516, rpoB526 and rpoB531 for RIF. Results: As revealed, (7528/15829, 47.56%) mycobacterial isolates were obtained, including 6937 (43.82%) MTBC and 591 (3.73%) NTM, respectively. Out of the 7528 MTB and NTM isolates, the average resistance to INH ratio was 21% and increased from 15.98% in 2011 to 18.76% in 2018. In the case of rifampin, the same resistance trend has been gradually increasing, that is, from 12.45% in 2011 to 14.55% in 2018. The average prevalence of MDR-TB in our study is approximately 14.18%, which is much higher than the global average MDR-TB incidence of 3.5%, and this is very alarming. Conclusion: Given the findings of the present study on non-tuberculous mycobacteria and drug sensitivity that have been documented and conducted over the past 8 years, we suggest that the identification, diagnosis and sensitivity tests for NTMs should be standardized and integrated into clinical systems.