Background. Considering that Hormozgan province in Iran (southern part of Iran on the Persian Gulf) is among the areas with high prevalence of MDR-MTB and attracts so many sailors and tourists, genetic diversity of MTB isolates circulating in this part of Iran was evaluated. Pattern of TB transmission was also examined. Methods and Material. A total of 38 isolates of MTB were cultured from TB patients from Hormozgan province of Iran and standard MIRU-VNTR typing and spoligotyping were applied to genotype these isolates. Drug susceptibility testing was performed using proportion method. Results. There were 28 VNTR profiles comprising 5 clusters and 23 unique isolates compared to 21 spoligotyping profiles, which contained 9 clusters and 12 unique isolates. Latin American-Mediterranean (n = 9, 23.6%) was found to be the most predominant lineage. MIRU-VNTR analysis, with an HGDI of 0.975, was more discriminating than spoligotyping, which had an HGDI of 0.955. The estimated proportion of TB cases due to recent transmission was 26.3% and 44.7% by MIRU-VNTR and spoligotyping, respectively. The rates of monodrug resistance and MDR were 15.8% and 7.9%, respectively. Two of 3 MDR strains were found to be related to MIRU-VNTR and belonged to the same spoligotyping cluster characterized with T1/SIT53 genotype. Conclusions. The high genetic diversity among MTB isolates suggests that transmission occurred from different sources to this area. Reactivation of a priori, latent MTB infection was found to contribute mainly to TB cases in this geographic region.