大肠埃希菌异质性耐药的研究进展

doi:10.13560/j.cnki.biotech.bull.1985.2022-0854

摘要/Abstract

摘要：

大肠埃希菌（Escherichia coli）是人类和动物中常见细菌病原体,E.coli已对多种抗菌药物产生耐药,且由于抗菌药物选择压力越来越大,致使菌株的适应性和致病性更强,耐药的传播更快、定植率更高,给全球流行病学和临床治疗带来极大的挑战。近年来,E.coli的异质性耐药（heteroresistance,HR）相关研究相继被报道,异质性耐药是指细菌中的同源亚群对某种抗菌药物表现出不同的敏感性,是敏感菌进化为耐药菌的中间阶段,不同于完全耐药,异质性耐药在临床上不易及时检出,常导致临床抗菌药物治疗失败。因此,为有效防控E.coli感染及耐药的演变,深入剖析E.coli异质性耐药的流行特征及可能的机制尤为重要,本文就E.coli的异质性耐药研究进行了梳理,并对异质性耐药从表型特征、检测方法和可能的机制等方面进行了概述。旨在加强对E.coli异质性耐药的认识和研究,以期为全面了解E.coli耐药过程及发生机制、优化抗菌药物使用策略、减缓完全耐药菌种的出现提供参考。

关键词: 大肠埃希菌, 抗菌药物, 异质性耐药, 机制

Abstract:

Escherichia coli is a commonly pathogen in humans and animals,which has developed resistance to a variety of antibiotics. Due to the increasing selection pressure of antibiotics,the E. coli strains are more adaptable and pathogenic,the spread of antimicrobial resistance is faster,and the colonization rate is higher,that poses great challenges to epidemiology and clinical treatment. In recent years,researches related to heteroresistance（HR）of E. coli have been reported. HR refers to the fact that the homologous subpopulations in bacteria show different susceptibility to a certain antibiotic,which is the intermediate stage of sensitive bacteria evolving into drug-resistant bacteria. Different from full resistance,it is difficult for HR to be detected clinically in time,which often leads to the failure of clinical therapy. Therefore,in order to effectively prevent and control E. coli infection and the evolution of antimicrobial resistance,it is particularly important to deeply analyze the epidemiological characteristics and possible mechanisms of E. coli's HR. This paper sorts the studies on HR,and reviews the phenotypic characteristics,detection methods and possible mechanisms of HR. It is aimed to strengthen the understanding and research on the HR of E.coli,and to provide a comprehensive understanding of E. coli antimicrobial resistance process and mechanism,optimizing antibiotics use strategies,and slowing down the emergence of resistant strains.

Key words: Escherichia coli, antibiotics, heteroresistance, mechanism

赵艳坤, 刘慧敏, 孟璐, 王成, 王加启, 郑楠. 大肠埃希菌异质性耐药的研究进展[J]. 生物技术通报, 2022, 38(9): 59-71.

ZHAO Yan-kun, LIU Hui-min, MENG Lu, WANG Cheng, WANG Jia-qi, ZHENG Nan. Research Progress in Heteroresistance of Escherichia coli[J]. Biotechnology Bulletin, 2022, 38(9): 59-71.

图/表 7

参考文献 59

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方法 Methods	优点 Advantages	缺点 Disadvantages	参考文献 Reference
E-test法、K-B纸片扩散法 E-test and Kirby-Bauer test	（1）传统、经典的药敏试验方法; （2）可用于异质性耐药的初筛; （3）操作简便、成本较低、结果直观易判读。	（1）由于平板上的细胞密度低,不易检出低频率出现的耐药亚群; （2）可信度低,假阳性、假阴性检出率较高。	[14]
M-E-test法M-E-test method	（1）可用于异质性耐药的大规模筛选; （2）操作简单。	培养板上细胞密度越高,检测异质性耐药亚群的概率越高,但检测的准确率不稳定,易造成误判。	[11]
GRD E-test、琼脂筛选 GRD E-test and agar screen	（1）临床实验室常规使用的药敏试验方法; （2）用于观察某种程度的耐药性和异质性耐药性之间存在强烈相关性的简单筛选方法（hVISA）。	（1）无法检测异质性耐药; （2）缺乏有关亚群频率和MIC的信息。	[15-16]
微量肉汤稀释法 Broth microdilution（BMD）	临床实验室常规使用的药敏试验方法。	（1）在存在耐药亚群的情况下,MIC的读数高度依赖于耐药细菌的接种密度和生长速率; （2）没有关于耐药亚群频率的信息,其MIC信息较差; （3）操作繁琐。	[17]
琼脂稀释 Agar dilution	临床实验室常规使用的药敏试验方法。	（1）使用耐药细菌的单个小菌落有可能错过低频率的亚群; （2）如果检测到异质性耐药,则提供有关耐药亚群的频率和MIC的少许信息; （3）耗时、费力。	[14]
群体谱分析法 PAP test	（1）检测异质性耐药较可靠的方法;（2）能够提供有关耐药亚群频率和MIC的有效信息; （3）可定量分析、准确度高、稳定性好、可重复性好。	（1）操作繁琐且成本昂贵; （2）耗时耗力; （3）技术性强,临床推广应用困难。	[12]
微量稀释群体谱分析法 Microdilution PAP test	（1）比常规PAP测试更快、更便宜; （2）可得知耐药亚群的频率和MIC。	检测的效果取决于细胞密度,误差较大。	[11-12]
改良群体谱分析曲线法 PAP-AUC	（1）hVISA检测的黄金标准; （2）准确性高。	（1）步骤繁琐且成本昂贵; （2）参考hVISA菌株可能会对不同药物浓度具有高度依赖性的耐药亚群分离株进行错误分类,造成假阳性或假阴性结果。	[18]
自动肉汤系统（如VITEK 2、MicroScan、WalkAway） Automated broth systems（for example,VITEK 2,MicroScan,and WalkAway）	临床实验室常规使用药敏试验方法。	在检测耐药亚群方面效果不佳。	[12]
分子检测方法（如探针分析和GeneXpert-Xpert-MTB/RIF-Ultra） Molecular detection methods（for example,line probe assays,GeneXpert Xpert MTB/RIF Ultra）	（1）与传统或自动检测方法相比,更能快速检测异质性耐药; （2）能够准确检测细菌亚群的点突变。（3）有时可直接用于患者样本（如,结核分枝杆菌的痰样本）。	（1）并非所有探针分析系统都能检测耐药亚群,检测效率和稳定性不稳定; （2）依赖于对耐药基因的全面了解,会造成结果差异较大,不适用于所有细菌。	[19]

方法 Methods	优点 Advantages	缺点 Disadvantages	参考文献 Reference
E-test法、K-B纸片扩散法 E-test and Kirby-Bauer test	（1）传统、经典的药敏试验方法; （2）可用于异质性耐药的初筛; （3）操作简便、成本较低、结果直观易判读。	（1）由于平板上的细胞密度低,不易检出低频率出现的耐药亚群; （2）可信度低,假阳性、假阴性检出率较高。	[14]
M-E-test法M-E-test method	（1）可用于异质性耐药的大规模筛选; （2）操作简单。	培养板上细胞密度越高,检测异质性耐药亚群的概率越高,但检测的准确率不稳定,易造成误判。	[11]
GRD E-test、琼脂筛选 GRD E-test and agar screen	（1）临床实验室常规使用的药敏试验方法; （2）用于观察某种程度的耐药性和异质性耐药性之间存在强烈相关性的简单筛选方法（hVISA）。	（1）无法检测异质性耐药; （2）缺乏有关亚群频率和MIC的信息。	[15-16]
微量肉汤稀释法 Broth microdilution（BMD）	临床实验室常规使用的药敏试验方法。	（1）在存在耐药亚群的情况下,MIC的读数高度依赖于耐药细菌的接种密度和生长速率; （2）没有关于耐药亚群频率的信息,其MIC信息较差; （3）操作繁琐。	[17]
琼脂稀释 Agar dilution	临床实验室常规使用的药敏试验方法。	（1）使用耐药细菌的单个小菌落有可能错过低频率的亚群; （2）如果检测到异质性耐药,则提供有关耐药亚群的频率和MIC的少许信息; （3）耗时、费力。	[14]
群体谱分析法 PAP test	（1）检测异质性耐药较可靠的方法;（2）能够提供有关耐药亚群频率和MIC的有效信息; （3）可定量分析、准确度高、稳定性好、可重复性好。	（1）操作繁琐且成本昂贵; （2）耗时耗力; （3）技术性强,临床推广应用困难。	[12]
微量稀释群体谱分析法 Microdilution PAP test	（1）比常规PAP测试更快、更便宜; （2）可得知耐药亚群的频率和MIC。	检测的效果取决于细胞密度,误差较大。	[11-12]
改良群体谱分析曲线法 PAP-AUC	（1）hVISA检测的黄金标准; （2）准确性高。	（1）步骤繁琐且成本昂贵; （2）参考hVISA菌株可能会对不同药物浓度具有高度依赖性的耐药亚群分离株进行错误分类,造成假阳性或假阴性结果。	[18]
自动肉汤系统（如VITEK 2、MicroScan、WalkAway） Automated broth systems（for example,VITEK 2,MicroScan,and WalkAway）	临床实验室常规使用药敏试验方法。	在检测耐药亚群方面效果不佳。	[12]
分子检测方法（如探针分析和GeneXpert-Xpert-MTB/RIF-Ultra） Molecular detection methods（for example,line probe assays,GeneXpert Xpert MTB/RIF Ultra）	（1）与传统或自动检测方法相比,更能快速检测异质性耐药; （2）能够准确检测细菌亚群的点突变。（3）有时可直接用于患者样本（如,结核分枝杆菌的痰样本）。	（1）并非所有探针分析系统都能检测耐药亚群,检测效率和稳定性不稳定; （2）依赖于对耐药基因的全面了解,会造成结果差异较大,不适用于所有细菌。	[19]

抗菌药物名称 Antibiotic	样品（来源） Sample （Sources）	样本数 Number of samples	异质性耐药测定方法Method for determination of heteroresistance	异质性耐药率 Prevalence of heteroresistance/%	耐药率Prevalence of resistance/%	异质性耐药亚群的频次 Frequency of heteroresistant subpopulations	国家/地区Country /Region	文献 Reference
亚胺培南 Imipenem	医院分离株	11	E-test（抑菌圈内菌落生长初步判定异质性） PAP（异质性耐药以MIC/NIC≥8来确定）	0	0	ND	瑞典	[11]
厄他培南 Ertapenem		11		27.3	0	4.20×10^-7- 1.10×10^-5
美罗培南 Meropenem		11		0	0	ND
多利培南 Doripenem		11		0	0	ND
厄他培南 Ertapenem	医院分离株	140	改良PAP	35	2	ND	美国	[22]
亚胺培南 Imipenem		20		25	0
美罗培南 Meropenem		100		30	0
亚胺培南 Imipenem	医院分离株	332	纸片扩散,E-test（抑菌圈内菌落生长确定为异质性）	25	0	8.00×10^-8- 1.80×10^-7	中国重庆	[23]
厄他培南 Ertapenem				17.2	0.6
美罗培南 Meropenem				3.9	0
亚胺培南 Imipenem	医院分离株	200	E-test（抑菌圈内菌落生长确定为异质性） PAP（异质性耐药以MIC/NIC≥8来确定）	29	0	ND	中国广西	[24]
厄他培南 Ertapenem				20.5	2
美罗培南 Meropenem				6	0.5
亚胺培南 Imipenem	血液、尿液和痰液	850	K-B纸片法 E-test PAP（异质性耐药以MIC/NIC≥8来确定）	20.5	0	ND	中国广东	[25]
厄他培南 Ertapenem				17.5	1.5
美罗培南 Meropenem				7	0.5

抗菌药物名称 Antibiotic	样品（来源） Sample （Sources）	样本数 Number of samples	异质性耐药测定方法Method for determination of heteroresistance	异质性耐药率 Prevalence of heteroresistance/%	耐药率Prevalence of resistance/%	异质性耐药亚群的频次 Frequency of heteroresistant subpopulations	国家/地区Country /Region	文献 Reference
亚胺培南 Imipenem	医院分离株	11	E-test（抑菌圈内菌落生长初步判定异质性） PAP（异质性耐药以MIC/NIC≥8来确定）	0	0	ND	瑞典	[11]
厄他培南 Ertapenem		11		27.3	0	4.20×10^-7- 1.10×10^-5
美罗培南 Meropenem		11		0	0	ND
多利培南 Doripenem		11		0	0	ND
厄他培南 Ertapenem	医院分离株	140	改良PAP	35	2	ND	美国	[22]
亚胺培南 Imipenem		20		25	0
美罗培南 Meropenem		100		30	0
亚胺培南 Imipenem	医院分离株	332	纸片扩散,E-test（抑菌圈内菌落生长确定为异质性）	25	0	8.00×10^-8- 1.80×10^-7	中国重庆	[23]
厄他培南 Ertapenem				17.2	0.6
美罗培南 Meropenem				3.9	0
亚胺培南 Imipenem	医院分离株	200	E-test（抑菌圈内菌落生长确定为异质性） PAP（异质性耐药以MIC/NIC≥8来确定）	29	0	ND	中国广西	[24]
厄他培南 Ertapenem				20.5	2
美罗培南 Meropenem				6	0.5
亚胺培南 Imipenem	血液、尿液和痰液	850	K-B纸片法 E-test PAP（异质性耐药以MIC/NIC≥8来确定）	20.5	0	ND	中国广东	[25]
厄他培南 Ertapenem				17.5	1.5
美罗培南 Meropenem				7	0.5

抗菌药物名称 Antibiotic	样品（来源） Sample（Sources）	样本数 Number of samples	异质性耐药测定方法 Method for determination of heteroresistance	异质性耐药率 Prevalence of heteroresistance/%	耐药率 Prevalence of resistance/%	异质性耐药亚群的频次 Frequency of heteroresistant subpopulations	国家/地区Country / Region	文献 Reference
黏菌素 Colistin	临床分离株	11	E-test（抑菌圈内菌落生长初步判定异质性） PAP（异质性耐药以MIC/NIC≥8来确定）	0	0	ND	瑞典	[11]
多黏菌素B Polymyxin B	临床分离株	11	E-test（抑菌圈内菌落生长初步判定异质性） PAP（异质性耐药以MIC/NIC≥8来确定）	0	0	ND
黏菌素 Colistin	血液分离株	292	琼脂筛选[菌落生长在黏菌素（4 mg/L）确定为异质性耐药）]	3.4	0.3	1.25×10^-8-1.10×10^-5	匈牙利	[28]
黏菌素 Colistin	医院分离株	291	PAP（异质性耐药以MIC/NIC≥8来确定） PAP	1.37	0.69	4.00×10^-7-4.00×10^-6	中国温州	[29]
黏菌素 Colistin	猪病料样本、猪肛门拭子	177	PAP（异质性耐药以MIC/NIC≥4来确定）	91.67	0	6.61×10^-7-2.57×10^-6	中国郑州	[30]