Cuella-Martin I, Runyambo D, De Bock S, Diels M, Niyompano H, Hakizayezu F, Keysers J, De Rijk W B, Habimana Y M, Gahamanyi N, Ardizzoni E, Muvunyi C M, de Jong B C, Ngabonziza J C S, Rigouts L
Unit of Mycobacteriology, Institute of Tropical Medicine, Antwerp, Belgium.
Department of Biomedical Sciences, Antwerp University, Antwerp, Belgium.
J Clin Microbiol. 2025 Apr 9;63(4):e0180624. doi: 10.1128/jcm.01806-24. Epub 2025 Mar 3.
This study evaluated thin-layer agar (TLA) as a faster alternative for both indirect minimum inhibitory concentration (MIC) determination of bedaquiline (BDQ) from culture isolates and direct drug-susceptibility testing (DST) from sputum samples. Indirect BDQ-MIC results from TLA were compared to the established 7H11 solid DST. Direct-TLA-DST performance was assessed using 143 baseline sputum samples from rifampicin-resistant TB cases. Direct-TLA tested susceptibility to rifampicin, isoniazid, levofloxacin, and BDQ, with results compared to Löwenstein-Jensen (LJ) and MGIT. For indirect BDQ-MIC determination, TLA accurately identified H37Rv MICs within the WHO control range (0.015-0.12μg/mL). Optimal results were obtained with standard incubation and day 7 reading of the plates. TLA correctly classified all wild-type clinical strains as BDQ-susceptible and detected 9 out of 10 BDQ-resistant strains with elevated MICs. Direct-TLA-DST detected MTB in 53.8% of samples, compared to 55.9% on LJ and 69.4% in MGIT. Uninterpretable results due to contamination or medium drying were low (4.9%). The median time to result was 15 days for smear-positive and 22 days for smear-negative samples, faster than WHO-endorsed methods. Sensitivity was 100% for rifampicin and 87.8% for isoniazid, with a specificity of 100% for all drugs except isoniazid (96.2%). No BDQ nor levofloxacin resistance was detected, thus direct TLA sensitivity could not be assessed. In conclusion, direct-TLA-DST offers a reliable and faster alternative to conventional DST methods for BDQ and other anti-TB drugs. Essentially, this technique can be operated at BioSafety Level 2, allowing decentralization of pDST for managing drug-resistant TB in settings with limited laboratory infrastructure.
This paper addresses the critical need for faster direct drug-susceptibility testing (DST) on sputum, especially for bedaquiline (BDQ), which is a key drug in treating drug-resistant TB. Currently, there is a lack of rapid, reliable methods for direct BDQ testing from sputum samples, limiting timely and accurate treatment decisions and monitoring. By demonstrating the potential of thin-layer agar (TLA) for direct BDQ-MIC determination, this study offers a promising solution that could significantly improve patient care.
本研究评估了薄层琼脂（TLA）作为一种更快的替代方法，用于从培养分离物中间接测定贝达喹啉（BDQ）的最低抑菌浓度（MIC）以及从痰标本中进行直接药敏试验（DST）。将TLA的间接BDQ-MIC结果与既定的7H11固体DST进行比较。使用来自耐利福平结核病病例的143份基线痰标本评估直接TLA-DST的性能。直接TLA检测对利福平、异烟肼、左氧氟沙星和BDQ的敏感性，结果与罗-琴（LJ）培养基和分枝杆菌生长指示管（MGIT）进行比较。对于间接BDQ-MIC测定，TLA在世界卫生组织控制范围内（0.015-0.12μg/mL）准确鉴定了H37Rv的MIC。在标准孵育和培养板第7天读数时获得了最佳结果。TLA将所有野生型临床菌株正确分类为对BDQ敏感，并检测出10株BDQ耐药菌株中的9株MIC升高。直接TLA-DST在53.8%的标本中检测到结核分枝杆菌，而LJ培养基上为55.9%，MGIT中为69.4%。由于污染或培养基干燥导致的无法解释的结果较低（占4.9%）。涂片阳性标本的中位报告时间为15天，涂片阴性标本为22天，比世界卫生组织认可的方法更快。利福平的敏感性为100%，异烟肼为87.8%，除异烟肼外所有药物的特异性均为100%（异烟肼为96.2%）。未检测到BDQ和左氧氟沙星耐药，因此无法评估直接TLA的敏感性。总之，对于BDQ和其他抗结核药物，直接TLA-DST为传统DST方法提供了一种可靠且更快的替代方法。从本质上讲，该技术可在生物安全2级操作，允许在实验室基础设施有限的环境中分散进行分子药敏试验以管理耐药结核病。
本文满足了对痰标本进行更快直接药敏试验（DST）的迫切需求，特别是对于贝达喹啉（BDQ），它是治疗耐药结核病的关键药物。目前，缺乏从痰标本中直接进行BDQ检测的快速、可靠方法，限制了及时、准确的治疗决策和监测。通过证明薄层琼脂（TLA）用于直接BDQ-MIC测定的潜力，本研究提供了一个有前景的解决方案，可显著改善患者护理。
