山东某院耐碳青霉烯类肠杆菌科细菌临床感染特征及耐药基因分析

doi:10.3969/j.issn.1673-8640.2020.08.004

摘要/Abstract

摘要：

目的了解济宁医学院附属医院耐碳青霉烯类肠杆菌科细菌（CRE）临床感染特征、药物敏感性及耐药基因分型,为预防和治疗CRE感染及医院多重耐药菌管理提供参考。方法收集CRE临床分离株93株,采用全自动微生物鉴定系统进行细菌鉴定及体外药物敏感性试验,并通过全自动快速微生物质谱检测系统确证菌株。采用简易碳青霉烯类灭活法（sCIM）试验进行碳青霉烯酶表型确证,采用Xpert Carba-R检测并鉴别碳青霉烯酶分型,通过聚合酶链反应（PCR）扩增碳青霉烯酶耐药基因并进行测序分型。结果济宁医学院附属医院CRE主要分离自痰液样本,其次是血液和尿液样本,科室来源主要为重症监护病房。CRE对临床常用的24种抗菌药物均耐药,对替加环素均敏感。sCIM试验检出79株产碳青霉烯酶;Xpert Carba-R检出KPC基因阳性38株、NDM基因阳性31株、IMP基因阳性6株、NDM+KPC基因阳性4株。PCR扩增结果显示,有38株携带KPC耐药基因,31株携带NDM耐药基因,6株携带IMP耐药基因,4株同时携带KPC和NDM耐药基因,未检出VIM及OXA-48耐药基因。结论济宁医学院附属医院CRE临床分离株对多种临床常用抗菌药物耐药,但对替加环素具有良好的体外敏感性;肺炎克雷伯菌碳青霉烯类耐药基因主要为KPC型,大肠埃希菌主要为NDM型;应加强对CRE的监控,防范其在院内的暴发流行。

关键词: 耐碳青霉烯类肠杆菌科细菌, 临床感染特征, 耐药基因

Abstract:

Objective To investigate the clinical infection characteristics,drug susceptibility and drug resistance genotypes of carbapenem-resistant Enterobacteriaceae（CRE） in the Affiliated Hospital of Jining Medical University,and to provide a reference for the prevention and treatment of CRE infection and the management of multi-drug resistant bacteria. Methods Totally,93 isolates of CRE were collected,bacterial identification and drug susceptibility tests were carried out by VITEK2 Compact automatic microbial identification and drug susceptibility system,and the isolates were confirmed by automatic rapid microbial mass spectrometry. Carbapenemase was confirmed by simplified carbapenem inactivation method（sCIM） phenotype confirmation test. Carbapenemase models were determined and identified by Xpert Carba-R. Carbapenemase resistance gene was amplified by polymerase chain reaction（PCR） and sequenced. Results CRE isolates were mainly isolated from sputum,blood and urine samples. CRE isolates were mainly distributed in intensive care unit. CRE isolates were resistant to 24 commonly used antibiotics and sensitive to tigecycline. Totally,79 isolates were positive for carbapenemase phenotype by sCIM phenotype confirmation test. Xpert Carba-R showed that there were 38 isolates of KPC gene positive,31 isolates of NDM gene positive,6 isolates of IMP gene positive and 4 isolates of NDM+KPC gene positive. A total of 38 isolates carrying KPC resistance genes,31 isolates carrying NDM resistance genes,6 isolates carrying IMP resistance genes and 4 isolates carrying KPC + NDM resistance genes. VIM and OXA-48 resistance genes were not determined. Conclusions CRE in the Affiliated Hospital of Jining Medical University is resistant to many commonly used antibiotics,but it has good drug susceptibility to tigecycline in vitro. The carbapenem-resistant gene of Klebsiella pneumoniae is mainly KPC type,and that of Escherichia coli is mainly NDM type. The surveillance should be strengthened to prevent its outbreak and nosocomial epidemic.

Key words: Carbapenem-resistant Enterobacteriaceae, Clinical infection characteristic, Drug resistance gene

中图分类号:

R446.1

陆书华, 李晓哲, 刘凌云, 金呈强, 戈宁宁, 董海新. 山东某院耐碳青霉烯类肠杆菌科细菌临床感染特征及耐药基因分析[J]. 检验医学, 2020, 35(8): 757-762.

LU Shuhua, LI Xiaozhe, LIU Lingyun, JIN Chengqiang, GE Ningning, DONG Haixin. Clinical infection characteristics and their drug resistance genes of carbapenem-resistant Enterobacteriaceae in a hospital of Shandong[J]. Laboratory Medicine, 2020, 35(8): 757-762.

图/表 4

参考文献 13

[1]	JING X,ZHOU H,MIN X,et al. The simplified carbapenem inactivation method(sCIM) for simple and accurate detection of carbapenemase-producing Gram-negative bacilli[J]. Front Microbiol,2018,9:2391.
[2]	VAN DER ZEE A,ROORDA L,BOSMAN G,et al. Multi-centre evaluation of real-time multiplex PCR for detection of carbapenemase genes OXA-48,VIM,IMP,NDM and KPC[J]. BMC Infect Dis,2014,14:27.
[3]	POIREL L,HÉRITIER C,TOLÜN V,et al. Emergence of oxacillinase-mediated resistance to imipenem in Klebsiella pneumoniae[J]. Antimicrob Agents Chemother,2004,48(1):15-22.
[4]	LOGAN L K,WEINSTEIN R A. The epidemiology of carbapenem-resistant Enterobacteriaceae:the impact and evolution of a global menace[J]. J Infect Dis,2017,215(suppl 1):S28-S36.
[5]	IOVLEVA A,DOI Y. Carbapenem-resistant Enterobacteriaceae[J]. Clin Lab Med,2017,37(37):303-315.
[6]	DUIN D V,DOI Y. The global epidemiology of carbapenemase producing Enterobacteriaceae[J]. Virulence,2016,13(3):3-8.
[7]	ZHANG R,CHAN E W,ZHOU H,et al. Prevalence and genetic characteristics of carbapenem-resistant Enterobacteriaceae strains in China[J]. Lancet Infect Dis,2017,17(3):256-257.
[8]	潘璐,朱凤雪,李纾,等. 重症监护病房中耐碳青霉烯类肠杆菌感染危险因素分析[J]. 中国医药,2016,11(8):1200-1203.
[9]	POTTER R F,SOUZA A W,DANTAS G. The rapid spread of carbapenem-resistant Enterobacteriaceae[J]. Drug Resist Updat,2016,29:30-46.
[10]	ZHANG Y,WANG Q,YIN Y,et al. Epidemiology of carbapenem-resistant Enterobacteriaceae infections:report from the China CRE Network[J]. Antimicrob Agents Chemother,2018,62(2):e01882-17.
[11]	WANG Q,WANG X,WANG J,et al. Phenotypic and genotypic characterization of carbapenem-resistant Enterobacteriaceae:data from a longitudinal large-scale CRE study in China(2012-2016)[J]. Clin Infect Dis,2018,67(suppl 2):S196-S205.
[12]	RAFAILIDIS P I,FALAGAS M E. Options for treating carbapenem-resistant Enterobacteriaceae[J]. Curr Opin Infect Dis,2014,27(6):479-483.
[13]	WANG X,WANG Q,CAO B,et al. Retrospective observational study from a Chinese network of the impact of combination therapy versus monotherapy on mortality from carbapenem-resistant Enterobacteriaceae bacteremia[J]. Antimicrob Agents Chemother,2018,63(1):e01511-18.

基因名称	引物序列（3'~5'）	退火温度/℃	产物长度/bp
KPC	F：TCGCTAAACTCGAACAGG	63	785
	R：TTACTGCCCGTTGACGCCCAATCC	63	785
NDM	F：TTGGCCTTGCTGTCCTTG	60	89
	R：ACACCAGTGACAATATCACCG	60	89
VIM	F：AAGTCCGTTAGCCCATTCCG	59	114
	R：GCGATATGCGACCAAACACC	59	114
IPM	F：GGGCGTTGTTCCTAAACATGG	61	185
	R：TAAGCCACTCTATTCCGCCC	61	185
OXA	F：TTGGTGGCATCGATTATCGG	60	743
	R：GAGCACTTCTTTTGTGATGGC	60	743

基因名称	引物序列（3'~5'）	退火温度/℃	产物长度/bp
KPC	F：TCGCTAAACTCGAACAGG	63	785
	R：TTACTGCCCGTTGACGCCCAATCC	63	785
NDM	F：TTGGCCTTGCTGTCCTTG	60	89
	R：ACACCAGTGACAATATCACCG	60	89
VIM	F：AAGTCCGTTAGCCCATTCCG	59	114
	R：GCGATATGCGACCAAACACC	59	114
IPM	F：GGGCGTTGTTCCTAAACATGG	61	185
	R：TAAGCCACTCTATTCCGCCC	61	185
OXA	F：TTGGTGGCATCGATTATCGG	60	743
	R：GAGCACTTCTTTTGTGATGGC	60	743

抗菌药物	敏感	中介	耐药
替加环素	100	0	0
头孢唑啉	0	0	100
氨苄西林	0	0	100
氨苄西林-舒巴坦	0	0	100
替卡西林-克拉维酸	0	0	100
阿米卡星	0	0	100
头孢曲松	0	0	100
亚胺培南	0	0	100
美洛培南	0	0	100
头孢噻肟	0	0	100
哌拉西林	0	0	100
哌拉西林-他唑巴坦	0	0	100
厄他培南	0	0	100
氨曲南	11.83	6.45	81.72
复方磺胺甲噁唑	47.31	0	52.69
环丙沙星	25.81	4.30	69.89
左氧氟沙星	45.16	0	54.84
庆大霉素	25.81	0	74.19
头孢吡肟	0	13.41	88.17
头孢哌酮-舒巴坦（纸片扩散法）	0	13.41	88.17
头孢他啶	0	3.23	96.77
头孢替坦	34.41	0	65.59
妥布霉素	18.28	3.23	78.49
四环素	11.83	0	88.17

抗菌药物	敏感	中介	耐药
替加环素	100	0	0
头孢唑啉	0	0	100
氨苄西林	0	0	100
氨苄西林-舒巴坦	0	0	100
替卡西林-克拉维酸	0	0	100
阿米卡星	0	0	100
头孢曲松	0	0	100
亚胺培南	0	0	100
美洛培南	0	0	100
头孢噻肟	0	0	100
哌拉西林	0	0	100
哌拉西林-他唑巴坦	0	0	100
厄他培南	0	0	100
氨曲南	11.83	6.45	81.72
复方磺胺甲噁唑	47.31	0	52.69
环丙沙星	25.81	4.30	69.89
左氧氟沙星	45.16	0	54.84
庆大霉素	25.81	0	74.19
头孢吡肟	0	13.41	88.17
头孢哌酮-舒巴坦（纸片扩散法）	0	13.41	88.17
头孢他啶	0	3.23	96.77
头孢替坦	34.41	0	65.59
妥布霉素	18.28	3.23	78.49
四环素	11.83	0	88.17

[1]	朱昱蓉, 张丹, 贺雅星, 李晶晶, 黄晶, 李婷, 刘鹏, 刘荣华. 肺炎克雷伯菌临床分离株分子分群和流行病学特征[J]. 检验医学, 2023, 38(5): 435-440.
[2]	方咏梅, 章燕, 徐雪莹, 钟峰. 金黄色葡萄球菌耐药性与耐药基因和毒力基因的相关性[J]. 检验医学, 2023, 38(4): 357-361.
[3]	王佳玮, 朱威南, 陈颖盈, 季萍, 王颖. 血培养分离碳青霉烯类耐药肺炎克雷伯菌临床分布和耐药基因分析[J]. 检验医学, 2023, 38(4): 362-367.
[4]	杨思敏, 轩乾坤, 羽晓瑜, 温冬华, 李广波. 国产某品牌大肠埃希菌和肺炎克雷伯菌及3种碳青霉烯耐药基因试剂性能验证[J]. 检验医学, 2023, 38(1): 56-59.
[5]	宋皓月, 舒文. 肉芽肿性乳腺炎患者结核硬脂酸棒状杆菌毒力因子和耐药性分析[J]. 检验医学, 2022, 37(11): 1075-1078.
[6]	姚锂凤, 郑巧平, 杨蒙, 罗清琼, 陈旭, 陈福祥. 胶体金免疫层析法快速检测CRE碳青霉烯酶效果评价[J]. 检验医学, 2022, 37(1): 68-71.
[7]	张冠怡, 姚冬婷, 王健, 胡晓波. 光滑念珠菌耐药机制及中西药治疗光滑念珠菌感染研究进展[J]. 检验医学, 2021, 36(7): 761-765.
[8]	薛玉芹, 黄韵, 张正银, 王亚婷, 金英, 桂芸, 余彬. 致伤口感染犬巴斯德菌耐药基因全基因测序分析[J]. 检验医学, 2020, 35(7): 699-701.
[9]	杜金龙, 冯燕, 李恒涛. 实时荧光定量PCR结合探针检测MP的临床价值及MP耐药情况分析[J]. 检验医学, 2020, 35(6): 535-539.
[10]	池宏波, 杨英梅. 台州地区结核分枝杆菌分离及耐药特征分析[J]. 检验医学, 2020, 35(5): 455-457.
[11]	黄春兰, 徐秋红, 余洪立, 马丽梅, 廖致红. 1株血液感染大肠弯曲菌生物学特性及全基因组序列相关性分析[J]. 检验医学, 2019, 34(12): 1091-1096.
[12]	苏思婷, 温洪宇, 袁振亚, 王秀颖, 王国振, 俞晨洁. 使用抗菌药物对婴幼儿肠道微生态的影响[J]. 检验医学, 2018, 33(8): 764-767.
[13]	张景皓, 杨峰, 方毅, 杨丽华, 刘文健, 赵虎, 张艳梅. 急性感染性腹泻沙门菌分离株的耐药谱及其相关基因分析[J]. 检验医学, 2018, 33(12): 1098-1102.
[14]	吴永琴, 应春妹. 白念珠菌唑类药物相关耐药基因及其调控机制的研究进展[J]. 检验医学, 2016, 31(9): 739-743.
[15]	刘立荣, 李向阳, 瞿玲娜, 唐玉霞, 曹景宏, 颜小萍. 替加环素对不同耐药基因型耐碳青霉烯类肠杆菌科细菌的敏感性研究[J]. 检验医学, 2016, 31(5): 383-386.