Abstract: To investigate the genetic variation among Mycobacterium tuberculosis isolates from East and West Azarbaijan provinces of Iran and to evaluate the manner of recent transmission of tuberculosis (TB), we performed IS6110-based restriction fragment length polymorphism analysis of isolates. Restriction fragment length polymorphism (RFLP) typing performed on 165 culture-positive specimens from East and West Azarbaijan. Using IS6110 as a probe, Mycobacterium tuberculosis strains assigned to clusters based on identical DNA fingerprints. Rates of patients have clustered were 27.68% in East and 30.19% in West Azarbaijan. There was not statistically significant differences in clustering of patients in two provinces (p = 0.4533) but infection with Mycobacterium tuberculosis in males and females in two provinces were different (p = 0.0048). In East Azarbaijan there was not difference in transmission of tuberculosis between males and females, also in males and females belonged to clusters we couldn’t find statistical difference (p = 0.1833). The rate of active transmission of TB in West Azarbaijan was slightly more than East Azarbaijan. It can be due to different factors such as poor economic and less developed condition in West Azarbaijan.