L strategies for their high levels of TB [21]. Consistent with our data, previous studies in Colombia have reported a high proportion of M. tuberculosis groupings using genetic methods for different regions; in particular, grouping proportions between 20 and 74 have been reported [22?4]. Each region must expand the number of isolates characterized to determine the own genotypes. One strength of this work is the characterization of a large number of isolates (741) from 31 of the 33 departments in Colombia over a long study period.PLOS ONE | DOI:10.1371/journal.pone.0124308 June 11,9 /Mycobacterium tuberculosis Grazoprevir web genotypes in ColombiaFig 2. Distribution of the genotypic families according to the isolation period. doi:10.1371/journal.pone.0124308.gA comparison of the two seven-year periods emphasizes that the status of TB transmission has not changed; this observation agrees with reports of classical epidemiology in which no variation was observed in the number of new cases diagnosed over time in Colombia [1]. There were no significant TSA site differences in the MDR or sensitive isolates in the two periods studied with respect to the groupings, which may indicate that the level of active transmission is not decreasing in Colombia. The Haarlem family was associated with grouped isolates, whereas the T, X and Orphan families were associated with the non-grouped isolates (p <0.001), which most likely represented endogenous reactivation or latent tuberculosis. In this study, a high genetic diversity of M. tuberculosis was reported, with 170 different genotypes present that were mainly represented by four families: LAM (39.9 ), Haarlem (19 ), Orphan (17 ) and T (9 ). The isolates of type SIT42 were the most common isolates belonging to the LAM9 family, which was found in all the departments included in this study. The SIT62 (H1) was the second most common type of isolate Fig 1. The LAM family has been described as prevalent in other countries including Paraguay and Venezuela and in countries in the Americas, Europe and the Caribbean [25, 26]. Previous studies that genotyped isolates in Colombia using spoligotyping reported that the LAM family was the most frequent family in regional circulation, followed by the Haarlem family [24?7]. Moreover, in this study the MANU family is first report in Colombian isolates from individuals coinfected with HIV. In total, 83 genotypes were found that had not been previously reported (orphans). These genotypes were likely native from Colombia, and 72.2 of the newly discovered genotypes were resistant to one or more drugs Fig 3A and 3B. The high proportion of patterns and orphan spoligotypes detected in this study, particularly those belonging to new cases, indicates that these genotypes should be monitored and investigated further because they may have been generated by recent developments in pre-existing genotypes. Regarding the association of families with phenotypes susceptible to first-line drugs, it was shown that an isolate of the Beijing family was a predictor of drug-resistant insulation; the frequency of these isolates increased significantly during the second period of the study. Previous studies showed that some isolates of the Beijing family are sensitive to drugs [28] and that inPLOS ONE | DOI:10.1371/journal.pone.0124308 June 11,10 /Mycobacterium tuberculosis Genotypes in ColombiaFig 3. Transmission dynamics of genotypes in isolates resistant (A) and sensitive (B) to first-line drugs. doi:10.1371/journal.L strategies for their high levels of TB [21]. Consistent with our data, previous studies in Colombia have reported a high proportion of M. tuberculosis groupings using genetic methods for different regions; in particular, grouping proportions between 20 and 74 have been reported [22?4]. Each region must expand the number of isolates characterized to determine the own genotypes. One strength of this work is the characterization of a large number of isolates (741) from 31 of the 33 departments in Colombia over a long study period.PLOS ONE | DOI:10.1371/journal.pone.0124308 June 11,9 /Mycobacterium tuberculosis Genotypes in ColombiaFig 2. Distribution of the genotypic families according to the isolation period. doi:10.1371/journal.pone.0124308.gA comparison of the two seven-year periods emphasizes that the status of TB transmission has not changed; this observation agrees with reports of classical epidemiology in which no variation was observed in the number of new cases diagnosed over time in Colombia [1]. There were no significant differences in the MDR or sensitive isolates in the two periods studied with respect to the groupings, which may indicate that the level of active transmission is not decreasing in Colombia. The Haarlem family was associated with grouped isolates, whereas the T, X and Orphan families were associated with the non-grouped isolates (p <0.001), which most likely represented endogenous reactivation or latent tuberculosis. In this study, a high genetic diversity of M. tuberculosis was reported, with 170 different genotypes present that were mainly represented by four families: LAM (39.9 ), Haarlem (19 ), Orphan (17 ) and T (9 ). The isolates of type SIT42 were the most common isolates belonging to the LAM9 family, which was found in all the departments included in this study. The SIT62 (H1) was the second most common type of isolate Fig 1. The LAM family has been described as prevalent in other countries including Paraguay and Venezuela and in countries in the Americas, Europe and the Caribbean [25, 26]. Previous studies that genotyped isolates in Colombia using spoligotyping reported that the LAM family was the most frequent family in regional circulation, followed by the Haarlem family [24?7]. Moreover, in this study the MANU family is first report in Colombian isolates from individuals coinfected with HIV. In total, 83 genotypes were found that had not been previously reported (orphans). These genotypes were likely native from Colombia, and 72.2 of the newly discovered genotypes were resistant to one or more drugs Fig 3A and 3B. The high proportion of patterns and orphan spoligotypes detected in this study, particularly those belonging to new cases, indicates that these genotypes should be monitored and investigated further because they may have been generated by recent developments in pre-existing genotypes. Regarding the association of families with phenotypes susceptible to first-line drugs, it was shown that an isolate of the Beijing family was a predictor of drug-resistant insulation; the frequency of these isolates increased significantly during the second period of the study. Previous studies showed that some isolates of the Beijing family are sensitive to drugs [28] and that inPLOS ONE | DOI:10.1371/journal.pone.0124308 June 11,10 /Mycobacterium tuberculosis Genotypes in ColombiaFig 3. Transmission dynamics of genotypes in isolates resistant (A) and sensitive (B) to first-line drugs. doi:10.1371/journal.
