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Abstract Background and objectives: Identification and monitoring of multidrug-resistant Mycobacterium tuberculosis strains (MDR) is highlighted by the high risk of their spreading in different areas. Prevalence of these strains was evaluated in Golestan province in northeast of Iran. Material and Methods: Drug susceptibility testing to Isoniazid and rifampin was carried out for 148 clinical samples that had grown in Mycobacteria growth indicator tube (MGIT) system, according to the manufacturer's instructions (Becton-Dickinson, USA). The association of drug resistance frequency with demographic characteristics and growth time were investigated. The appropriate statistical tests, X2 and student T- test were performed for comparison of these variants. A p value>0.05 was considered significant in all cases. Results: The turnaround time required for growth of Mycobacterium tuberculosis in MGIT system was between 2 to 55 days (mean 16.3±10.4 days). Of all samples studied, 17.6% and 3.4% were resistant to Isoniazid and rifampin, respectively, and 3.4% (5 samples) were MDR (CI 95% 1-6%). The turnaround time required for determining MDR cases was 9.6 days. No statistically significant association was found between the resistance to the drugs and none of the factors including sex, age, type of clinical sample, and positivity of the smear. Conclusion: The prevalence of MDR in the studied region was determined to be 3.4% which is similar to the country-wide evaluations. The turnaround time for Mycobacterium growth and anti drug susceptibility result can be shortened by MGIT method. Key words: Mycobacterium tuberculosis, Mycobacterium Growth Indicator Tube, Multidrug Resistant