Resistance to the second-line injectable medicines (SLIDs) amikacin (AMK), capreomycin (CAP), and kanamycin (KAN) was known to be conferred by point mutations in the rrs gene and the eis promoter. While mutations in these classical genes impart the majority of SLID resistance, other routes of resistance were frequent. They posed a threat to accurate treatment decisions when utilizing traditional molecular diagnostics. 1,184 clinical Mycobacterium TB isolates from 7 countries were investigated for genetic markers linked to phenotypic resistance. Resistance to all 3 SLIDs was linked to the markers rrs: A1401G and rrs: G1484T, while kanamycin resistance (KAN-R) was linked to 3 known markers in the eis promoter (eis: G-10A, eis: C-12T, and eis: C-14T). Canonical mutations were found in 274 (84.3%), 250 (77.2%), and 249 (92.3%) of the 325, 324, and 270 AMK-R, CAP-R, and KAN-R isolates, respectively. About 13 of the isolates had several canonical mutations. Only 103 of the phenotypically resistant isolates had canonical mutations. 3 genes and promoters with mutations were related to unexplained resistance to at least 1 SLID in a genome-wide association investigation. The results linked whiB7 5′-untranslated-region mutations to KAN resistance, indicating that the previously discovered mechanism of KAN resistance had clinical implications. Researchers also found evidence for a new link between CAP resistance and the promoter of the Rv2680-Rv2681 operon, which encoded an exoribonuclease that might affect CAP’s ribosome binding.