Cycloserine resistance among drug-resistant tuberculosis cases in Taiwan

Cycloserine (CS) is a widely used drug for drug-resistant tuberculosis (DR-TB) treatment. The World Health Organization (WHO) recently suggested a critical concentration (CC) for CS using the MGIT 960 system (MGIT). To strengthen our DR-TB management program, we performed CS resistance analyses to provide comprehensive drug susceptibility testing (DST). This retrospective study included Mycobacterium tuberculosis complex (MTBC) isolates obtained from 114 rifampin-resistant (RR) and multidrug-resistant (MDR) TB cases. We compared the results of phenotypic DST (pDST) and genotypic DST (gDST) and evaluated the minimum inhibitory concentration (MIC) using both MGIT and the Sensititre MYCOTB MIC Plate (Sensititre). Sanger sequencing and whole-genome sequencing were conducted to analyze the mutations of CS-resistant-associated ald and alr genes. Our results indicated that the optimal consistency with gDST was achieved with the CC of 16 µg/mL for MGIT, which aligns with the WHO recommendations, and the CC of 8 µg/mL for Sensititre. Of the 114 MTBC isolates, we found 5 (4.4%) CS-MGIT-resistant isolates, which all harbored mutations in the alr gene, including three previously known mutations M343T, T20M, L113R, and a novel mutation R243S, whereas seven low-MIC isolates harboring alr Q30R mutations might not be associated with CS resistance. Notably, M1I, E118K, and A184T in the ald gene and L113R, R243S, S261N, and M343T in the alr gene were predicted to have a destabilizing effect, which could interfere with protein functions and induce drug resistance. For accurate routine diagnosis of CS susceptibility, we adopted the CC of 16 µg/mL and suggested an interim 8 µg/mL using MGIT and Sensititre, respectively.IMPORTANCETo strengthen the DR-TB management program in Taiwan, we performed cycloserine (CS) resistance analyses to enhance treatment outcomes. Of the 114 drug-resistant tuberculosis (DR-TB) isolates, we found 5 (4.4%) CS-MGIT-resistant isolates, with four isolates classified as multidrug-resistant (MDR)-TB and one isolate as Pre-XDR-TB. In addition, we observed all CS-MGIT-resistant isolates harbored mutations in the alr gene, including three previously known high-confidence mutations M343T, T20M, and L113R, as well as the novel R243S mutation. We also found that mutations could lead to CS resistance by disrupting protein stability.