Because the INH resistance-conferring mutations observed here, i.e. katG S315T and inhA promoter C15T, are known to be associated with a low fitness cost [11], they might not require compensation. All RIF resistant isolates harbored mutations in rpoB at codons D516F, D516Y or S531L except one, which did Rapamycin not have any mutation in the 600pb rpoB fragment sequenced. DST was repeated for this

case, confirming the MDR phenotype. Furthermore, common rpoB katG and inhA promoter mutations were excluded by Genotype MTBDRplus. Nevertheless, it has been estimated that mutations in the RIF resistance determining region (81-bp region in rpoB) account only for 95% of RIF resistance [6] and therefore other mechanisms cannot be excluded.

Mutation S531L has been linked to high-level RIF resistance [12], whereas D516Y was associated with low-level resistance [13–15]. Mutation D516F has only been reported in Kazakhstan [16] and may also cause low-level resistance. Low-level RIF resistance has been little considered, but could influence treatment, especially knowing that phenotypic DST outcomes may differ from the actual efficacy of the anti-TB drugs in patients [17]. STR resistant isolates harbored mutations in rpsL (codons K43R, K88Q, K88R) and rrs (nucleotide A514C), as previously reported [18, 19]. One isolate was mutated at codon V77G in gidB, a mutation which was not reported before. One STR resistant isolate did not present any mutation in any of these genes. Mutations in gidB have been associated with low-level STR resistance [20, VX 809 21], but were also reported in sensitive strains [22]. In this study, gidB mutations A10P, L16R, E92D, and A205A were observed among strains resistant to other drugs than STR. We further explored gidB mutations in whole genomes of 21 pan-susceptible strains representative of the

six defined M. tuberculosis lineages [23]. Mutation gidB V77G, which we observed in one STR resistant isolate from PNG, could not be found in any of the 21 pan-susceptible strains. This mutation could therefore indeed be involved in drug triclocarban resistance or could be a transitory polymorphism in the population. The mutation A10P observed in one STR sensitive isolate was not found in any of the 21 pan-susceptible genomes. Mutations L16R was observed in genome sequences from Lineage 4 strains (Euro-American lineage) and E92D in Lineage 2 strains (East-Asian lineage). This supports the recent observation that gidB L16R occurred in LAM strains (i.e. Lineage 4), whereas gidB E92D occurred in Beijing strains [24]. A205A appeared mutated in all strains not belonging to Lineage 4, therefore indicating that this mutation, identified by comparison to H37Rv, is a Lineage 4 mutation. Observations from the 21 pan-susceptible genomes suggest that most gidB mutations rather reflect M. tuberculosis lineage evolution than drug resistance.