Laboratory of Infection and Immunity, West China Center of Medical Sciences, Sichuan University, Chengdu 610041, China.
J Antimicrob Chemother. 2017 Jul 1;72(7):1893-1900. doi: 10.1093/jac/dkx110.
Ofloxacin and moxifloxacin are the most commonly used fluoroquinolones (FQs) for the treatment of tuberculosis. As a new generation FQ, moxifloxacin has been recommended for the treatment of ofloxacin-resistant TB. However, the mechanism by which ofloxacin-resistant Mycobacterium tuberculosis further gains resistance to moxifloxacin remains unclear.
We used Mycobacterium smegmatis as a model for studying FQ resistance in M. tuberculosis . Moxifloxacin-resistant M. smegmatis was selected in vitro based on strains with primary ofloxacin resistance. The gyrA and gyrB genes of the resistant strains were sequenced to identify resistance-associated mutations. An in vitro competition assay was applied to explore the influence of gyrA / gyrB mutations on bacterial fitness. Finally, we evaluated the clinical relevance of our findings by analysing the WGS data of 1984 globally collected M. tuberculosis strains.
A total of 57 moxifloxacin-resistant M. smegmatis strains based on five ofloxacin-resistant strains were obtained. Sequencing results revealed that all moxifloxacin-resistant strains harboured second-step mutations in gyrA or gyrB . The relative fitnesses of the double-mutation strains varied from 0.65 to 0.93 and were mostly lower than those of their mono-mutation parents. From the genomic data, we identified 37 clinical M. tuberculosis strains harbouring double mutations in gyrA and/or gyrB and 36 of them carried at least one low-level FQ-resistance mutation.
Double mutation in DNA gyrase leads to moxifloxacin resistance and decreased fitness in M. smegmatis . Under current dosing of moxifloxacin, double mutations mainly happened in M. tuberculosis strains with primary low-level resistance mutations.
氧氟沙星和莫西沙星是治疗结核病最常用的氟喹诺酮类药物（FQ）。莫西沙星作为新一代 FQ，已被推荐用于治疗氧氟沙星耐药结核病。然而，耐氧氟沙星结核分枝杆菌进一步获得莫西沙星耐药的机制尚不清楚。
我们使用耻垢分枝杆菌作为结核分枝杆菌 FQ 耐药性的模型。根据初始对氧氟沙星耐药的菌株，在体外选择莫西沙星耐药的耻垢分枝杆菌。对耐药株的 gyrA 和 gyrB 基因进行测序，以鉴定耐药相关突变。应用体外竞争试验探讨 gyrA/gyrB 突变对细菌适应性的影响。最后，通过分析全球收集的 1984 株结核分枝杆菌的 WGS 数据，评估我们发现的临床相关性。
共获得基于 5 株氧氟沙星耐药株的 57 株莫西沙星耐药耻垢分枝杆菌株。测序结果显示，所有莫西沙星耐药株均在 gyrA 或 gyrB 中存在第二步突变。双突变株的相对适应性从 0.65 到 0.93 不等，大多低于其单突变亲本。从基因组数据中，我们鉴定出 37 株临床结核分枝杆菌株在 gyrA 和/或 gyrB 中存在双突变，其中 36 株至少携带一个低水平 FQ 耐药突变。
DNA 回旋酶的双突变导致莫西沙星耐药和耻垢分枝杆菌适应性降低。在目前莫西沙星的给药剂量下，双突变主要发生在具有原发性低水平耐药突变的结核分枝杆菌菌株中。
