Background: Bedaquiline (BDQ), delamanid (DLM), and linezolid (LZD) are core drugs for multidrug-resistant tuberculosis (MDR-TB) treatment. Comprehensive regional data on phenotypic susceptibility and genotypic resistance-associated mutations are essential to guide optimized therapy. This study aimed to characterize phenotypic and genotypic resistance in treatment-naïve MTBC isolates by determining MICs of BDQ, DLM, and LZD and identifying associated genetic mutations. Methods: Minimum inhibitory concentrations (MICs) of BDQ, DLM, and LZD were determined for 71 Mycobacterium tuberculosis isolates (30 fully drug-susceptible and 41 MDR-TB) collected in the Cukurova region of Southern Türkiye between 2021 and 2023 using the MGIT 960/EpiCenter system. Sequencing of resistance-associated genes was analyzed: atpE and Rv0678 for BDQ, ddn and fbiA for DLM, and rrl and rplC for LZD. Results: MICs ranged from ≤0.025–6.4 µg/mL for BDQ, 0.005–0.32 µg/mL for DLM, and 0.125–2 µg/mL for LZD. According to WHO breakpoints, resistance rates were 7.0% for BDQ, 4.2% for DLM, and 7% for LZD, all confined to MDR-TB isolates. Nine MDR-TB isolates (12.7%) harbored atpE mutations, including both previously reported and putatively novel substitutions. A single ddn mutation was detected in a DLM-susceptible isolate. No mutations were observed in Rv0678, fbiA, rrl, or rplC. Conclusion: BDQ, DLM and LZD showed strong activity against MDR-TB isolates. However, the presence of atpE variants in both resistant and susceptible isolates demonstrated genotype-phenotype discordance of resistance. These results underline the necessity of the combination of genetic and phenotypic testing for detection of resistance and guidance for treatment.

Background: To determine the molecular relationship between pyrazinamide (PZA) and bedaquiline (BDQ) resistance mechanisms in multidrug-resistant tuberculosis (MDR-TB) and drug-susceptible Mycobacterium tuberculosis (M. tuberculosis) isolates to improve treatment strategies. Methods: Phenotypic drug susceptibility testing (DST), resistance gene amplification and sequencing (pncA, panD, hadC, and Rv0678), and efflux pump gene expression (mmpS5 and mmpL5) by qRT-PCR were performed on 71 clinical MTBC isolates from seven provinces in Southern Türkiye (2022-2023). Results: PZA resistance was closely associated with MDR-TB (χ² = 38.0, p < 0.0001), exclusively detected in MDR isolates (42% overall; 73% of MDR). pncA mutations were detected in 77% of PZA-resistant isolates with strong genotype-phenotype correlation (p = 0.009, r = 0.45), while panD mutations were present in 40% and showed less association (p = 0.14). No hadC or Rv0678 mutations were identified despite significant overexpression of efflux genes mmpS5 and mmpL5 in MDR strains (p < 0.001). Conclusions: PZA and BDQ resistance arise through distinct, non-overlapping mechanisms. PZA-resistant MDR-TB without Rv0678 mutations confirms bedaquiline's independent therapeutic value and success in treating PZA-resistant cases.