Characterization of predominant Mycobacterium tuberculosis strains from different subpopulations of India Jyoti Arora a , Urvashi Balbir Singh a, *, Naga Suresh a , Tanu Rana a , Chhavi Porwal a , Amit Kaushik a , Jitendra Nath Pande b a Department of Microbiology, All India Institute of Medical Sciences, New Delhi, India b Sitaram Bharatiya Institute of Science and Research, New Delhi, India 1. Introduction The resurgence of tuberculosis (TB) has renewed interest in understanding the epidemiology and pathogenesis of the disease. Genotyping of clinical isolates in different parts of the world has shown that global epidemiology of TB is propagated by thousands of different genotypes (van Soolingen et al., 1999; Warren et al., 1999). The strains occur at different frequencies, and the relative frequencies in different areas vary between districts, cities, countries and continents (Brudey et al., 2006; Filliol et al., 2002; Filliol et al., 2003). The dynamics of TB epidemic in a given area and time frame may therefore be a factor of the different strains circulating in that region. The knowledge of circulating strains can be used for molecular evolutionary and population genetics studies. IS6110-based typing is the most widely applied genotyping method in the molecular epidemiology of Mycobacterium tubercu- losis. The IS6110 insertion sequence is specific to the M. tuberculosis, are present in different copy numbers (0–25) and their positions in the genome are highly variable among different isolates (van Embden et al., 1993). Spoligotyping targeting the DNA polymorph- ism at the direct repeat locus (DR locus) of the genome of M. tuberculosis complex, allows simultaneous detection and differ- entiation of M. tuberculosis complex strains (Kamerbeek et al., 1997). The DR locus is well conserved and stable enough rendering it specific for detecting M. tuberculosis complex strains. SNPs in the katG and gyrA genes classify M. tuberculosis isolates into three principal genetic groups (PGGs). TbD1 is specifically present in a subset of PGG1 strains, but absent in other strains of PGG1, and in PGG2 and PGG3 strains. Based on the presence or absence of an M. tuberculosis specific deletion (TbD1), M. tuberculosis strains can be segregated into ‘‘ancestral’’ versus ‘‘modern’’ lineages (Brosch et al., 2002). Though the known ‘‘modern’’ M. tuberculosis families are very widely prevalent worldwide (Filliol et al., 2002; Filliol et al., 2003; Brudey et al., 2006), ancient principal genetic group 1 (PGG1) clones are responsible for TB in India (Kulkarni et al., 2005; Gutierrez et al., 2006; Singh et al., 2007). Studies in India lack reference to preferential localization of certain strain families in different Infection, Genetics and Evolution 9 (2009) 832–839 ARTICLE INFO Article history: Received 31 October 2008 Received in revised form 27 April 2009 Accepted 3 May 2009 Available online 20 May 2009 Keywords: Tuberculosis Spoligotyping Restriction fragment length polymorphism Shared type Clade ABSTRACT The predominant strains from India belong to Central-Asian (CAS) and the East-African-Indian (EAI) clade of Mycobacterium tuberculosis. The two clades have also been shown to be geographically partitioned. The study of such strains may help to understand the characteristics that make M. tuberculosis an effective pathogen and its overrepresentation in certain populations. M. tuberculosis isolates characterized by spoligotyping under a population based tuberculosis study covering different regions from the North and South India were further analyzed by restriction fragment length polymorphism (RFLP) and by deletion analysis of M. tuberculosis specific deletion region 1 (TbD1). The genetic relationship of the two clades inferred using different genetic markers showed good correlation. In the North where the CAS clade predominates the isolates are characterized by presence of high IS6110 copy number and absence of TbD1 region whereas in the South where the EAI clade predominates the isolates are characterized by low copy number of IS6110 and presence of TbD1 region. The ancestral EAI strains were found to be less often associated with drug resistance or young age as compared to the CAS clade. The study highlights that the EAI lineage is well established in India and that CAS may be emerging or more recently introduced to India. The results depict a distinction in the lineage of strains from the North versus South India indicating a need to study if the pathogen has adapted to specific human populations. ß 2009 Elsevier B.V. All rights reserved. * Corresponding author. Tel.: +91 11 26588500/700x6436; fax: +91 11 26588641. E-mail address: drurvashi@gmail.com (U.B. Singh). Contents lists available at ScienceDirect Infection, Genetics and Evolution journal homepage: www.elsevier.com/locate/meegid 1567-1348/$ – see front matter ß 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.meegid.2009.05.008 Exhibit 3F