A third group of isolates (n = 36 [16.0% of the tested isolates] segregated into 29 newly identified

spoligotype patterns (not reported by SpolDB4). The strain families that could be grouped by SpolDB4 included: LAM (46.4%, n = 104), Haarlem (16.0%, n = 36), T (14.3%, n = 32), X (6.2%, n = 14), S (4.5%, n = 10), U (4.9%, 11), W/Beijing (1.8%, n = 4), MANU2 (0.4%, n = 1). Twelve (4.8%) isolates had an unclassified spoligopattern. Five isolates were included as Haarlem because of their spoligotype signature but did not match any of the patterns in SpolDB4 [21]. Table 2 Frequency of 27 shared spoligotypes (SITs) according to Brudey et al. [21] identified in 158 INH resistant M. tuberculosis strains isolated from South America. SIT Octal Strains in this n Strains in n Lineag APO866 mouse 1 0000000000037 3 1.3 5610 13.2 Beijing 47 7777777740207 6 2.6 1021 2.4 Haarlem 602 7777777700007 2 0.9 48 0.1 U 50 7777777777207 19 8.5 2128 5.0

Haarlem 49 7777777777207 3 1.3 115 0.3 Haarlem3 20 6777776077607 9 4.0 588 1.4 LAM 17 6777376077607 6 2.4 473 1.1 LAM 33 7761776077607 8 3.6 770 1.8 LAM 4 0000000077607 3 1.3 220 0.5 LAM3/S 211 5761776077607 2 0.9 63 0.1 LAM 828 3777776077607 3 1.3 20 0.0 LAM 93 7777376077607 10 4.5 267 0.6 LAM 64 7777776075607 9 4.0 157 0.4 LAM 435 7637776077607 3 1.3 4 0.0 LAM 177 3777776077607 3 1.3 50 0.1 LAM 388

7377776077607 2 0.9 15 0.0 LAM 42 7777776077607 22 9.9 1926 4.5 LAM 1938 7763777777607 7 3.1 3 0.0 S 53 7777777777607 17 7.6 3738 MK0683 mouse 8.8 T1 397 7777776000007 2 0.9 13 0.0 U 402 7777776000000 3 1.3 14 0.0 U 1241 7777776077007 3 1.3 28 0.0 U 119 7777767777607 2 0.9 659 1.8 X1 137 7777767777606 MycoClean Mycoplasma Removal Kit 3 1.3 720 2.0 X2 92 7000767777607 3 1.3 328 0.8 X3 91 7000367777607 2 0.9 143 0.4 X3 60 7777776077607 3 1.3 83 0.2 LAM Association between MIC levels, characterized mutations and spoligotype strain families Higher level INH resistance (≥2 μg/mL) was significantly associated with the S315T katG mutation, as shown by a greater odds ratio of 1.97 (Table 3). Of note, in isolates with MIC ≥16 μg/mL (83.0%, n = 38) a mutation was found one or more of the studied genes. We next evaluated for potential the relationship between MIC levels and mutations and strain families. The S315T katG mutation was found in LAM isolates (77.9%, n = 81), Haarlem isolates (94.4%, n = 34), and in T isolates (68.7%, n = 22). Of the Beijing strains (n = 4), 3 presented with the S315T katG mutation. We noted a statistical association between Haarlem strain family with the S315T katG mutation (p = 0.01) (Table 3). When the specific S315T katG mutation was considered, the Haarlem genotype occurred more frequently among those M.