摘要

β-内酰胺抗生素替莫西林（6-α-甲氧基 - 替卡西林）对大多数扩展谱β-内酰胺酶显示出稳定性，但被认为对铜绿假单胞菌无活性。 MexAB-OprM外排系统中的突变，天然存在于囊性纤维化（CF）分离株中，先前已显示可逆转这种内在抗性。在本研究中，我们测量了铜绿假单胞菌CF分离物的大型集合（n = 333）中的替莫西林活性。 29％的菌株MICs≤16 mg / L（建议替莫唑林的临床断点）。对于替莫西林MIC≤512mg / L（核苷酸插入或缺失，提前终止，串联重复，不停止和错义突变）的mexA或mexB，观察到突变。观察到替莫西林MIC与N-苯基-1-萘胺（MexAB-OprM荧光底物）的流出速率和胞外外多糖丰度（有助于粘液表型）之间的相关性。 OpdK或OpdF阴离子特异性孔蛋白表达将替莫西林MIC仅减少约1倍两倍稀释。与替莫西林对铜绿假单胞菌无活性的常见假设相反，我们在此展示临床上可利用的MIC，其中不可忽视比例的CF分离物由mexA和/或mexB中的多种突变解释。在更广泛的背景下，这项工作有助于增加我们对MexAB-OprM功能的理解，并帮助描述抗生素如何与MexA和MexB相互作用。

The β-lactam antibiotic temocillin (6-α-methoxy-ticarcillin) shows stability to most extended spectrum β-lactamases, but is considered inactive against Pseudomonas aeruginosa. Mutations in the MexAB-OprM efflux system, naturally occurring in cystic fibrosis (CF) isolates, have been previously shown to reverse this intrinsic resistance. In the present study, we measured temocillin activity in a large collection (n = 333) of P. aeruginosa CF isolates. 29% of the isolates had MICs ≤ 16 mg/L (proposed clinical breakpoint for temocillin). Mutations were observed in mexA or mexB in isolates for which temocillin MIC was ≤512 mg/L (nucleotide insertions or deletions, premature termination, tandem repeat, nonstop, and missense mutations). A correlation was observed between temocillin MICs and efflux rate of N-phenyl-1-naphthylamine (MexAB-OprM fluorescent substrate) and extracellular exopolysaccharide abundance (contributing to a mucoid phenotype). OpdK or OpdF anion-specific porins expression decreased temocillin MIC by ~1 two-fold dilution only. Contrarily to the common assumption that temocillin is inactive on P. aeruginosa, we show here clinically-exploitable MICs on a non-negligible proportion of CF isolates, explained by a wide diversity of mutations in mexA and/or mexB. In a broader context, this work contributes to increase our understanding of MexAB-OprM functionality and help delineating how antibiotics interact with MexA and MexB.

https://www.nature.com/articles/srep40208