In Vivo Evolution to Colistin Resistance by PmrB Sensor Kinase Mutation in KPC-Producing Klebsiella pneumoniae Is Associated with Low-Dosage Colistin Treatment
In Vivo Evolution to Colistin Resistance by PmrB Sensor Kinase Mutation in KPC-Producing Klebsiella pneumoniae Is Associated with Low-Dosage Colistin Treatment
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DOI：
10.1128/aac.02555-14
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发表时间：
2014-08-01
影响因子：
4.9
通讯作者：
Rossolini, Gian Maria
中科院分区：
文献类型：
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作者：
Cannatelli, Antonio;Di Pilato, Vincenzo;Rossolini, Gian Maria
粘菌素是治疗由产生碳青霉烯酶的肠杆菌科广泛耐药菌株引起的感染的关键药物。然而，越来越多的报道出现了粘菌素耐药性，特别是在产生kpc型碳青霉烯酶（KPC-KP）的肺炎克雷伯菌菌株中。在这项工作中，我们分别从低剂量粘菌素治疗前和治疗后的KPC-KP感染患者中获得粘菌素敏感（KPB-1）和耐粘菌素（KPB-2）序列分离株。通过下一代测序方法和对两个分离株的比较基因组分析，我们在KPB-2中检测到编码PmrB传感器激酶基因的非同义核苷酸替换，导致82个氨基酸位置的亮氨酸到精氨酸替换。与KPB-1相比，KPB-2表现出pmrA和pmrK的转录上调，pmrK是pmrHFIJKLM操纵子的一部分，负责修饰粘菌素脂多糖靶点。KPB-2中与野生型pmrB的互补恢复了粘菌素敏感性，并将pmrA和pmrK的转录降低到基础水平，而pmrB （L82R）在KPB-1中的表达没有改变粘菌素敏感性或上调pmrA和pmrK的表达，证实了野生型pmrB相对于pmrB （L82R）突变体的优势。目前的结果表明，PmrB突变介导粘菌素耐药可能是在低剂量粘菌素治疗期间选择的。在没有选择压力的情况下，KPB-2的抗粘菌素表型在长达50代的时间里是稳定的，并且在竞争实验中与显著的适应成本无关。
Colistin is a key drug for the treatment of infections caused by extensively drug-resistant strains of Enterobacteriaceae producing carbapenemases. However, the emergence of colistin resistance is being increasingly reported, especially among Klebsiella pneumoniae strains producing KPC-type carbapenemases (KPC-KP). In this work, we investigated colistin-susceptible (KPB-1) and colistin-resistant (KPB-2) sequential isolates obtained from a patient with a KPC-KP infection before and after low-dosage colistin treatment, respectively. By using a next-generation sequencing approach and comparative genomic analysis of the two isolates, we detected in KPB-2 a nonsynonymous nucleotide substitution in the gene encoding the PmrB sensor kinase, resulting in a leucine-to-arginine substitution at amino acid position 82. Compared with KPB-1, KPB-2 exhibited upregulated transcription of pmrA and of pmrK, which is part of the pmrHFIJKLM operon responsible for modification of the colistin lipopolysaccharide target. Complementation with wild-type pmrB in KPB-2 restored colistin susceptibility and reduced the transcription of pmrA and pmrK to basal levels, while expression of PmrB(L82R) in KPB-1 did not alter colistin susceptibility or upregulate pmrA and pmrK expression, confirming the dominance of wild-type PmrB versus the PmrB(L82R) mutant. The present results indicated that PmrB mutations mediating colistin resistance may be selected during low-dosage colistin treatment. The colistin-resistant phenotype of KPB-2 was stable for up to 50 generations in the absence of selective pressure and was not associated with a significant fitness cost in a competition experiment.