Service de Microbiologie, Hôpital Beaujon, AP-HP, Clichy, France Institut Pasteur, Microbial Evolutionary Genomics, Paris, France CNRS, UMR3525, Paris, France.
INSERM U1047, Université Montpellier 1, UFR de Médecine, Nîmes, France Laboratoire de Bactériologie, CHU Caremeau, Nîmes, France.
J Antimicrob Chemother. 2015 Jan;70(1):81-8. doi: 10.1093/jac/dku340. Epub 2014 Sep 4.
In Klebsiella pneumoniae, overexpression of the AcrAB efflux pump and the more recently described OqxAB efflux pump has been linked to an antibiotic cross-resistance phenotype, but the mechanisms of regulation are largely unknown. Moreover, while AcrAB has been shown to participate in K. pneumoniae virulence, the contribution of OqxAB has not yet been assessed.
In the present study we investigated a K. pneumoniae clinical isolate (KPBj1 E+), displaying cross-resistance to quinolones, chloramphenicol and cefoxitin, and its phenotypic revertant (KPBj1 Rev, susceptible to antibiotics) by using whole-genome sequencing, RT-PCR, complementation and a Caenorhabditis elegans virulence model.
We detected a point mutation in the oqxR repressor gene of KPBj1 E+, which overexpressed genes rarA, encoding a transcriptional regulator, and oqxB, but not acrB. Complementation with wild-type oqxR restored antibiotic susceptibility and normalized rarA and oqxB expression levels. Whole-genome sequencing showed that KPBj1 Rev had lost the entire rarA-oqxABR locus, situated close to an integration hot spot of phage P4. This large deletion seemed responsible for the significantly lower virulence potential of strain KPBj1 Rev compared with KPBj1 E+. Moreover, we found that KPBj1 E+ ΔacrB was significantly less virulent than its parental strain.
This work demonstrates the role of the overexpression of efflux pump OqxAB, due to a mutation in gene oqxR, in the antibiotic resistance phenotype of a clinical isolate, and suggests that the presence of AcrAB, associated with overexpression of OqxAB, is required for high virulence potential.
在肺炎克雷伯菌中，AcrAB 外排泵和最近描述的 OqxAB 外排泵的过度表达与抗生素交叉耐药表型有关，但调节机制在很大程度上尚不清楚。此外，虽然已经表明 AcrAB 参与了肺炎克雷伯菌的毒力，但尚未评估 OqxAB 的贡献。
本研究通过全基因组测序、RT-PCR、互补和秀丽隐杆线虫毒力模型，研究了一株具有喹诺酮类、氯霉素和头孢西丁交叉耐药性的肺炎克雷伯菌临床分离株（KPBj1 E+）及其表型回复突变株（KPBj1 Rev，对抗生素敏感）。
我们在 KPBj1 E+中检测到 oqxR 抑制剂基因的点突变，导致 rarA（编码转录调节剂）和 oqxB 过度表达，但 acrB 没有。用野生型 oqxR 互补恢复了抗生素敏感性，并使 rarA 和 oqxB 的表达水平正常化。全基因组测序显示，KPBj1 Rev 失去了整个 rarA-oqxABR 基因座，该基因座靠近噬菌体 P4 的整合热点。这种大片段缺失似乎导致菌株 KPBj1 Rev 的毒力潜力明显低于 KPBj1 E+。此外，我们发现 KPBj1 E+ΔacrB 的毒力明显低于其亲本菌株。
这项工作证明了由于基因 oqxR 的突变导致外排泵 OqxAB 的过度表达在临床分离株的抗生素耐药表型中的作用，并表明与 OqxAB 过度表达相关的 AcrAB 的存在是高毒力潜力所必需的。
