利福平通过外排降低利福平耐药结核分枝杆菌对氧氟沙星的敏感性。
Rifampicin reduces susceptibility to ofloxacin in rifampicin-resistant Mycobacterium tuberculosis through efflux.
机构信息
Department of Biomedical Sciences, Faculty of Health Sciences, Stellenbosch University, Cape Town, South Africa.
出版信息
Am J Respir Crit Care Med. 2011 Jul 15;184(2):269-76. doi: 10.1164/rccm.201011-1924OC. Epub 2011 Apr 21.
RATIONALE
Central dogma suggests that rifampicin resistance in Mycobacterium tuberculosis develops solely through rpoB gene mutations.
OBJECTIVE
To determine whether rifampicin induces efflux pumps activation in rifampicin resistant M. tuberculosis strains thereby defining rifampicin resistance levels and reducing ofloxacin susceptibility.
METHODS
Rifampicin and/or ofloxacin minimum inhibitory concentrations (MICs) were determined in rifampicin resistant strains by culture in BACTEC 12B medium. Verapamil and reserpine were included to determine their effect on rifampicin and ofloxacin susceptibility. RT-qPCR was applied to assess expression of efflux pump/transporter genes after rifampicin exposure. To determine whether verapamil could restore susceptibility to first-line drugs, BALB/c mice were infected with a MDR-TB strain and treated with first-line drugs with/without verapamil.
MEASUREMENTS AND MAIN FINDINGS
Rifampicin MICs varied independently of rpoB mutation and genetic background. Addition reserpine and verapamil significantly restored rifampicin susceptibility (p = 0.0000). RT-qPCR demonstrated that rifampicin induced differential expression of efflux/transporter genes in MDR-TB isolates. Incubation of rifampicin mono-resistant strains in rifampicin (2 μg/ml) for 7 days induced ofloxacin resistance (MIC > 2 μg/ml) in strains with an rpoB531 mutation. Ofloxacin susceptibility was restored by exposure to efflux pump inhibitors. Studies in BALB/c mice showed that verapamil in combination with first-line drugs significantly reduced pulmonary CFUs after 1 and 2 months treatment (p < 0.05).
CONCLUSION
Exposure of rifampicin resistant M. tuberculosis strains to rifampicin can potentially compromise the efficacy of the second-line treatment regimens containing ofloxacin, thereby emphasising the need for rapid diagnostics to guide treatment. Efflux pump inhibitors have the potential to improve the efficacy of anti-tuberculosis drug treatment.
原理
中心法则表明，结核分枝杆菌的利福平耐药性仅通过 rpoB 基因突变发展。
目的
确定利福平是否诱导利福平耐药结核分枝杆菌菌株中流出泵的激活，从而定义利福平耐药水平并降低氧氟沙星敏感性。
方法
通过 BACTEC 12B 培养基培养确定利福平耐药株的利福平最小抑菌浓度（MIC）和氧氟沙星最小抑菌浓度（MIC）。加入维拉帕米和利血平以确定它们对利福平及氧氟沙星敏感性的影响。应用 RT-qPCR 评估利福平暴露后外排泵/转运体基因的表达。为了确定维拉帕米是否可以恢复对一线药物的敏感性，用 MDR-TB 菌株感染 BALB/c 小鼠并用/不用维拉帕米治疗一线药物。
测量和主要发现
利福平 MIC 独立于 rpoB 突变和遗传背景变化。添加利血平和维拉帕米显著恢复利福平敏感性（p = 0.0000）。RT-qPCR 表明利福平诱导 MDR-TB 分离株中外排/转运体基因的差异表达。在利福平（2μg/ml）中孵育利福平单耐药株 7 天可诱导 rpoB531 突变株中氧氟沙星耐药（MIC>2μg/ml）。用外排泵抑制剂暴露可恢复氧氟沙星敏感性。BALB/c 小鼠研究表明，维拉帕米与一线药物联合应用可显著降低治疗 1 个月和 2 个月后肺部 CFU（p<0.05）。
结论
利福平耐药结核分枝杆菌菌株暴露于利福平可能会影响包含氧氟沙星的二线治疗方案的疗效，从而强调需要快速诊断以指导治疗。外排泵抑制剂有可能提高抗结核药物治疗的疗效。
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