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  • In an endemic country like India it is important to characterize M. tuberculosis population structure to understand the epidemiology of the disease. This study was undertaken in Ghatampur, Kanpur region of North India where the DOTS program is being closely monitored, by using two strains-specific molecular markers, namely spoligotyping and Mycobacterial Interspersed Repetitive Units-Variable number of Tandem Repeats (MIRU-VNTRs). A total of 335 isolates were genotyped by standard spoligotyping and the strains were compared with those in the SITVIT2 proprietary database of Institut Pasteur de la Guadeloupe. The Central Asian (CAS) sub-lineage was the most prevalent lineage comprising 59.1% (n =198) of all isolates, with SIT26/CAS1-Delhi being the most dominant genotype comprising, 27.16% of all isolates. Other lineages observed were: East-African Indian (EAI) (19.10%, n = 64), T (5.07%, n=17), Beijing (3.28%, n= 11), Manu (2.98%, n=10), X(2.68%, n=9), S(0.89%, n=3),H3,Ural (0.59%, n=2) each, LAM 9 (0.29%,n=1) and unknown (5.37%, n=18). This data was compared with published reports from India and data available in SITVIT2 database about genotypes of M.tuberculosis in neighboring countries.   This analysis confirms that CAS lineage has a pan India presence but EAI lineage is confined to southern parts of India as reported earlier. Comparison with prevalent genotypes in neighboring countries shows that China is unique in having a predominance of Beijing lineage,  Iran is  having an almost equal proportion of Ural and CAS lineages; while the rest of the Middle-East and Indian subcontinent are characterized by a gradient of CAS lineage predominating in the north of tropic of cancer, and the ancestral EAI lineage in the South and South-East Asia. Additionally, 12 loci MIRU-VNTR typing was performed in a subset of 13 spoligotyping defined clusters. MIRU–VNTR typing detected ninety two patterns, fifty three isolates showed >70% (similarity at nine loci or more than nine loci) homology. It was also observed that Beijing lineage strains were more associated with MDR strains (p-value=0.001). As MIRU-VNTR typing has been confirmed to have better discriminatory power and  multi-step application of combination of spoligotyping and MIRU-VNTR typing for analyzing the molecular epidemiology of TB may provide a better means of fingerprinting and studying transmission dynamics.