Division of Bioresources, Research Center for Zoonosis Control, Hokkaido University, Japan.
Faculty of Veterinary Medicine, Kasetsart University, Thailand.
J Infect Chemother. 2020 Nov;26(11):1139-1145. doi: 10.1016/j.jiac.2020.06.002. Epub 2020 Jul 12.
Plasmid-encoded quinolone resistance protein Qnr is an important factor in bacterial resistance to quinolones. Qnr interacts with DNA gyrase and reduces susceptibility to quinolones. The gene qnr likely spreads rapidly among Enterobacteriaceae via horizontal gene transfer. Though the vast amounts of epidemiological data are available, molecular details of the contribution of QnrB19, the predominant Qnr in Salmonella spp., to the acquisition of quinolone resistance has not yet been understood well.
We aimed to examine the role of QnrB19 in quinolone resistance acquisition using recombinant Salmonella Typhimurium DNA gyrases and QnrB19.
Recombinant QnrB19 was expressed in E. coli and purified by Ni-NTA agarose column chromatography. DNA supercoiling activities of recombinant Salmonella Typhimurium DNA gyrase were assessed with or without QnrB19 under the existence of three quinolones to measure ICs, the concentration of each quinolone required for 50% inhibition in vitro.
The ICs of norfloxacin, ciprofloxacin and nalidixic acid against DNA gyrases were measured to be 0.30, 0.16 and 17.7 μg/mL, respectively. The addition of QnrB19 increased the ICs of norfloxacin and ciprofloxacin to be 0.81 and 0.48 μg/mL, respectively, where no effect of QnrB19 was observed on the IC of nalidixic acid.
QnrB19 was shown for the first time in vitro to have ability to grant non-classical quinolone resistance to S. Typhimurium DNA gyrase. Structural insight on quinolones in this study may contribute to investigate drugs useful for preventing the spread of plasmid carrying PMQR along with other factors associating with antimicrobial resistance in S. Typhimurium and other bacteria.
质粒编码的喹诺酮类药物耐药蛋白 Qnr 是细菌对喹诺酮类药物耐药的重要因素。Qnr 与 DNA 回旋酶相互作用，降低了对喹诺酮类药物的敏感性。该基因 qnr 可能通过水平基因转移在肠杆菌科中迅速传播。尽管已经有大量的流行病学数据，但目前还不完全了解主要存在于沙门氏菌属中的 QnrB19 对获得喹诺酮类药物耐药性的贡献的分子细节。
我们旨在使用重组鼠伤寒沙门氏菌 DNA 回旋酶和 QnrB19 来研究 QnrB19 在喹诺酮类药物耐药性获得中的作用。
在大肠杆菌中表达重组 QnrB19 并通过 Ni-NTA 琼脂糖柱层析纯化。在存在三种喹诺酮类药物的情况下，评估重组鼠伤寒沙门氏菌 DNA 回旋酶的 DNA 超螺旋活性，以测量 IC 值，即每种喹诺酮类药物在体外抑制 50%时的浓度。
分别测量诺氟沙星、环丙沙星和萘啶酸对 DNA 回旋酶的 IC 值，结果分别为 0.30、0.16 和 17.7μg/mL。加入 QnrB19 后，诺氟沙星和环丙沙星的 IC 值分别增加至 0.81 和 0.48μg/mL，而 QnrB19 对萘啶酸的 IC 值没有影响。
首次在体外证明 QnrB19 具有赋予鼠伤寒沙门氏菌 DNA 回旋酶非经典喹诺酮类耐药性的能力。本研究中对喹诺酮类药物的结构研究可能有助于研究预防携带 PMQR 的质粒与其他与鼠伤寒沙门氏菌和其他细菌的抗菌耐药性相关的因素一起传播的有用药物。
