上海交通大学学报（医学版）

上海交通大学学报（医学版） >

2022 , Vol. 42 >Issue 7: 839 - 845

DOI: https://doi.org/10.3969/j.issn.1674-8115.2022.07.001

论著 · 基础研究

铜绿假单胞菌对环丙沙星异质性耐药的研究

  • 李聪聪 ,
  • 姚玉峰 ,
  • 张传珍
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  • 上海交通大学基础医学院免疫学与微生物学系，上海 200025
李聪聪（1998—），男，硕士生；电子信箱：m15295528299@163.com。
张传珍，电子信箱：1049119959@qq.com。

收稿日期: 2022-04-02

  录用日期: 2022-05-22

  网络出版日期: 2022-09-04

基金资助

国家自然科学基金(81830068);国家自然科学基金青年科学基金项目(82102407)

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Emergence of ciprofloxacin heteroresistance in clinical Pseudomonas aeruginosa

  • Congcong LI ,
  • Yufeng YAO ,
  • Chuanzhen ZHANG
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  • Department of Immunology and Microbiology, Shanghai Jiao Tong University College of Basic Medical Sciences, Shanghai 200025, China
ZHANG Chuanzhen，E-mail: 1049119959@qq.com.

Received date: 2022-04-02

  Accepted date: 2022-05-22

  Online published: 2022-09-04

Supported by

National Natural Science Foundation of China(81830068);National Natural Sciences Fund Youth Fund Project(82102407)

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摘要

目的·探究铜绿假单胞菌对环丙沙星的异质性耐药机制。方法·通过圆盘扩散法对临床分离鉴定的铜绿假单胞菌（n=227）的环丙沙星异质性耐药进行初筛，同时根据抑菌圈直径对这些菌株的耐药性进行划分。通过无抗生素条件下连续传代后测定敏感性来判断异质性耐药菌株的稳定性。利用群体谱型分析法（population analysis profiling，PAP）对初筛结果进行验证。通过全基因测序分析异质性耐药的产生机制，最后测定比较不同突变类型的耐药亚群对不同抗生素敏感性的影响。结果·在圆盘扩散法中，227株临床菌株中有142株（62.6%）对环丙沙星表现为敏感，26株（11.5%）表现为中介，59株（26.0%）表现为耐药。该方法筛选得到18株潜在异质性耐药菌株，其中11株经PAP及稳定性检测后确定为稳定异质性耐药菌。随机挑选的18株初筛鉴定为非异质性耐药的菌株中，有3株经PAP鉴定为异质性耐药菌株。通过全基因组测序发现11株耐药亚群中，有8株发生mexS突变，2株发生gyrA突变；除此之外，基因fleQ、PA2632和PAKAF_02255均分别在1株菌株中发生突变。mexS与gyrA这2类突变型均导致莫西沙星耐药，部分mexS突变株同时对氯霉素与亚胺培南耐药。结论·铜绿假单胞菌对环丙沙星的异质性耐药率为4.8%（11/227）。圆盘扩散法作为铜绿假单胞菌异质性耐药筛选方法灵敏度较低，存在一定假阴性率。mexS突变是耐药亚群中最常见的突变类型，该突变可能通过使外排泵MexEF-OprN过表达来介导对环丙沙星在内的多种抗生素耐药。

关键词： 铜绿假单胞菌; 环丙沙星; 异质性耐药; 圆盘扩散法; 群体谱型分析

本文引用格式

李聪聪 , 姚玉峰 , 张传珍 . 铜绿假单胞菌对环丙沙星异质性耐药的研究[J]. 上海交通大学学报（医学版）, 2022 , 42(7) : 839 -845 . DOI: 10.3969/j.issn.1674-8115.2022.07.001

Abstract

Objective

·To investigate ciprofloxacin heteroresistance in Pseudomonas aeruginosa (PA).

Methods

·Ciprofloxacin heteroresistance in 227 clinical PA strains was initially identified by disk diffusion. Meanwhile, susceptibilities of these strains was classified according to the diameter of the inhibition zone. The stability of the identified heteroresistant strains was tested by measuring the sensitivity after serial passage under antibiotic-free conditions. Mechanisms mediating heteroresistance were analyzed by whole-genome sequencing. Susceptibilities of resistant subpopulations to different antibiotics were also detected.

Results

·Based on disk diffusion, 142 (62.6%), 26 (11.5%), and 59 (26.0%) of 227 isolates were classified as susceptible, intermediate and resistant to ciprofloxacin respectively. Eighteen putative heteroresistant strains were detected through primary disk diffusion selection, and 11 among them were further verified as stable heteroresistant strains by population analysis profiling (PAP). By whole-genome sequencing, 8 and 2 among 11 resistant subpopulations carried mutations in mexS and mutations in gyrA individually. In addition, mutations in fleQ, PA2632 or PAKAF_02255 were detected respectively. Both mexS and gyrA mutants led to moxifloxacin resistance, and some mexS mutants also showed resistance to chloramphenicol and imipenem.

Conclusion

·Ciprofloxacin heteroresistance rate inPA is 4.8% (11/227). As a screening method for heteroresistance of PA, the disk diffusion has low sensitivity and has a certain false negative rate. The mutation mexS is the predominant mutation type in resistant subpopulations and it might enhance the overexpression of efflux pump MexEF-OprN, thus mediating resistance to different antibiotics including ciprofloxacin.

Key words： Pseudomonas aeruginosa; ciprofloxacin; heteroresistance; disk diffusion; population analysis profiling (PAP)

参考文献

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