Department of Infectious Diseases, Wenzhou Central Hospital, The Dingli Clinical College of Wenzhou Medical University, The Second Affiliated Hospital of Shanghai University, Wenzhou, China.
Laboratory of Infectious Diseases, Wenzhou Central Hospital, The Dingli Clinical College of Wenzhou Medical University, The Second Affiliated Hospital of Shanghai University, Wenzhou, China.
Microbiol Spectr. 2024 Jun 4;12(6):e0385923. doi: 10.1128/spectrum.03859-23. Epub 2024 May 13.
This study aimed to assess the possible causes of discordant results between Xpert MTB/RIF (Xpert) and Bactec MGIT 960 Culture System (MGIT960) regarding rifampicin (RIF) susceptibility in . Patients with previous RIF-resistant tuberculosis who were admitted to Wenzhou Central Hospital from January 2020 to December 2022 were enrolled. The isolates obtained from these patients were subjected to RIF susceptibility tests using Xpert and MGIT960, and the minimum inhibitory concentration (MIC) of RIF was determined by the MYCOTB MIC plate test. Additionally, molecular docking and molecular dynamics (MD) simulations were performed to evaluate the binding efficacy of rpoB and RIF based on rpoB mutations detected in the isolates with discordant RIF susceptibility results. A total of 28 isolates with discordant RIF susceptibility test results were detected, 15 of them were RIF susceptible with MICs ≤ 0.5 µg/mL. Twelve out of 15 isolates contained borderline RIF resistance-associated mutations [L430P ( = 6), H445N ( = 6)], 1 isolate had D435Y and Q429H double mutation, and the remaining 2 isolates had a silent (Q432Q) mutation. Compared with the affinity of RIF toward the wild type (WT) (-45.83 kcal/mol) by MD, its affinity toward L452P (-55.52 kcal/mol), D435Y (-47.39 kcal/mol), L430P (approximately -69.72 kcal/mol), H445N (-49.53 kcal/mol), and Q429H (-55.67 kcal/mol) increased. Borderline RIF resistance-associated mutations were the main cause for the discordant RIF susceptibility results between Xpert and MGIT960, and the mechanisms of the resistance need further investigated.IMPORTANCEThis study is aimed at assessing discordant results between Xpert MTB/RIF (Xpert) assay and Bactec MGIT 960 Culture System (MGIT960) regarding the detection of rifampicin (RIF)-resistant isolates in Wenzhou, China. The discordant results of RIF between these two assays were mainly caused by borderline RIF resistance-associated mutations, subsequently by silent mutations of rpoB. Borderline RIF resistance- associated mutations detected in our study were demonstrated to not be affected by the affinity of rpoB and RIF by molecular dynamics, and the mechanism of resistance was needed to be clarified. For the discordant results of RIF by Xpert and MGIT960 that occurred, DNA sequencing was recommended to investigate its association with resistance to RIF.
这项研究旨在评估 Xpert MTB/RIF（Xpert）和 Bactec MGIT 960 培养系统（MGIT960）在检测中国温州耐利福平（RIF）结核分枝杆菌分离株时，结果不一致的可能原因。该研究纳入了 2020 年 1 月至 2022 年 12 月期间因先前耐 RIF 结核病而入住温州市中心医院的患者。从这些患者中获得的分离株分别用 Xpert 和 MGIT960 进行 RIF 药敏试验，并用 MYCOTB MIC 板试验测定 RIF 的最小抑菌浓度（MIC）。此外，还进行了分子对接和分子动力学（MD）模拟，以根据分离株中检测到的 rpoB 突变评估 rpoB 和 RIF 的结合效果，这些分离株的 RIF 药敏试验结果不一致。共检测到 28 株 RIF 药敏试验结果不一致的分离株，其中 15 株的 MIC 值≤0.5μg/ml，为 RIF 敏感。在 15 株 RIF 敏感的分离株中，有 12 株含有边界 RIF 耐药相关突变[L430P（=6），H445N（=6）]，1 株同时存在 D435Y 和 Q429H 双重突变，其余 2 株存在无义突变（Q432Q）。与野生型（WT）相比，RIF 通过 MD 与 rpoB 的亲和力（-45.83 kcal/mol），其与 L452P（-55.52 kcal/mol）、D435Y（-47.39 kcal/mol）、L430P（约-69.72 kcal/mol）、H445N（-49.53 kcal/mol）和 Q429H（-55.67 kcal/mol）的亲和力增加。边界 RIF 耐药相关突变是导致 Xpert 和 MGIT960 之间 RIF 药敏结果不一致的主要原因，其耐药机制仍需进一步研究。
重要性：
本研究旨在评估中国温州地区 Xpert MTB/RIF（Xpert）检测与 Bactec MGIT 960 培养系统（MGIT960）检测耐利福平（RIF）结核分枝杆菌分离株时，RIF 检测结果不一致的原因。这两种检测方法 RIF 结果不一致的主要原因是边界 RIF 耐药相关突变，其次是 rpoB 的沉默突变。我们的研究表明，检测到的边界 RIF 耐药相关突变并不受 rpoB 和 RIF 亲和力的影响，需要阐明其耐药机制。对于 Xpert 和 MGIT960 检测到的 RIF 结果不一致，建议进行 DNA 测序以研究其与 RIF 耐药的关系。
