广东鹅场大肠杆菌耐药状况及blaCTX-M基因的传播特征

doi:10.16431/j.cnki.1671-7236.2020.05.032

摘要/Abstract

摘要： 为揭示广东地区鹅场动物和环境源大肠杆菌的耐药情况及超广谱β-内酰胺酶CTX-M的流行与传播特征，本研究从广东省江门及阳江市共10处鹅场采集鹅及环境样品199份，采用MALDI-TOF-MS法分离鉴定大肠杆菌。采用琼脂稀释法对菌株进行耐药性分析，采用PCR法检测头孢噻肟耐药菌中bla_CTX-M基因及其基因环境，采用脉冲场凝胶电泳（PFGE）、接合转移和质粒复制子分型等方法探究bla_CTX-M基因的传播特征。结果显示，共获得196株大肠杆菌，对氨苄西林、多西环素、氟苯尼考和链霉素耐药率均超过50%，第三代头孢菌素耐药率为10%~25%，其中头孢噻肟耐药菌有49株（24.6%）。阳江地区大肠杆菌对受试药物的耐药率高于江门，且动物源高于环境源，尤其是头孢噻肟和头孢噻呋均存在显著差异（P<0.05）。头孢噻肟耐药菌中共检出19株携带bla_CTX-M基因，包括bla_CTX-M-55（n=17）、bla_CTX-M-27（n=1）和bla_CTX-M-65（n=1），且bla_CTX-M基因阳性菌均可对5~11种药物耐药，呈现多重耐药的表型。bla_CTX-M-55基因环境均为ISEcp1-bla_CTX-M-55-orf477，且在ISEcp1与bla_CTX-M基因之间有3种长度的间隔序列；而bla_CTX-M-27和bla_CTX-M-65的基因环境均为ISEcp1-bla_CTX-M-27/65-IS903。19株bla_CTX-M基因阳性菌呈现10种PFGE谱型，存在一种主要流行的谱型（47.4%），其包括多种来源菌株，暗示存在克隆传播现象。12株（63.2%）bla_CTX-M基因阳性大肠杆菌中bla_CTX-M基因转移成功，bla_CTX-M基因阳性接合子携带的复制子型为IncFⅡ（n=10）和IncHⅠ2（n=2），且存在多西环素和氟苯尼考耐药表型与bla_CTX-M基因共转移现象。研究发现，阳江鹅场大肠杆菌耐药情况较为严重，bla_CTX-M基因存在一定的流行性且以bla_CTX-M-55亚型为主，bla_CTX-M基因阳性菌的克隆传播和质粒及插入序列ISEcp1介导的水平传播是导致该基因在鹅场大肠杆菌中扩散的主要原因，应引起高度重视。

关键词: 鹅场; 大肠杆菌; 耐药性; bla_CTX-M基因; 第三代头孢菌素

Abstract: To investigate the antimicrobial resistance and the prevalence and tranmission of extended-spectrum β-lactamase CTX-M,a total of 199 samples were collected from geese and the environment from 10 geese farms in Jiangmen and Yangjiang of Guangdong.Isolation and identification of the Escherichia coli (E.coli) were performed by MALDI-TOF-MS.Antibiotic susceptibility test of E.coli strains was characterized using agar dilution method.The bla_CTX-M gene and its genetic environment were detected by PCR analysis among cefotaxime-resistant E.coli isolates.Pulsed-field gel electrophoresis (PFGE),conjugation experiments and plasmid replicon types were conducted to determine the transmission of bla_CTX-M gene.A total of 196 E.coli strains were obtained and antimicrobial susceptibility testing showed that over half of these isolates were resistant to ampicilin,florfenicol,doxycycline and streptomycin.Meanwhile,10% to 25% were third-generation cephalosporins-resistant,and 49 (24.6%) isolates showed resistance to cefotaxime.For comparison,resistance rates of all the tested drugs among E.coli strains from Yangjiang and geese were generally higher than the ones from Jiangmen and environment,respectively,especially for cefotaxime and ceftiofur (P<0.05).Among the cefotaxime-resistant E.coli strains,19 possessed bla_CTX-Mgene,including bla_CTX-M-55(n=17),bla_CTX-M-27(n=1) and bla_CTX-M-65(n=1).All of the bla_CTX-M-carrying strains showed multidrug-resistant phenotype against 5 to 11 kinds of antimicrobial agents.The ISEcp1-bla_CTX-M-55-orf477 structure was found in all of the 17 bla_CTX-M-55-carrying E.coli strains with three different spacer sequences between ISEcp1 and bla_CTX-M-55,while the ISEcp1-bla_CTX-M-27/65-IS903 structure was identified in 2 isolates harboring bla_CTX-M-9G.Ten pulsed-field gel electrophoresis (PFGE) patterns were found in 19 bla_CTX-M-positive E.coli strains,with a dominant profile (47.4%),including isolates from various sources,indicated the clonal transmission of bla_CTX-M-positive isolates in local regions.The bla_CTX-Mgene was successfully transferred among 12 isolates (63.2%) carrying bla_CTX-M,and IncFⅡ plasmid (n=10) was the most prevalent replicon identified among these bla_CTX-M-carrying transconjugants,followed by IncHⅠ2 (n=2).The doxycycline- and florfenicol-resistant phenotypes were co-transferred with bla_CTX-M gene.In conclusion,our study revealed that it was serious for the antimicrobial resistance of E.coli strains in Yangjiang and a relatively high prevalence of bla_CTX-M gene,mainly bla_CTX-M-55 gene,was found.The clonal spread and horizontal transfer via plasmids and ISEcp1 element were major factors contributing to the dissemination of bla_CTX-M gene in the geese farms in Guangdong.It should be further monitored and attached more attention.

Key words: geese farm; Escherichia coli; antimicrobial resistance; bla_CTX-M gene; the third-generation cephalosporins

中图分类号:

S852.61

岑道机, 麦嘉琳, 周毓源, 耿昊宇, 刘昌海, 邓国辉, 姜琪, 方亮星. 广东鹅场大肠杆菌耐药状况及bla_CTX-M基因的传播特征[J]. 中国畜牧兽医, 2020, 47(5): 1571-1582.

CEN Daoji, MAI Jialin, ZHOU Yuyuan, GENG Haoyu, LIU Changhai, DENG Guohui, JIANG Qi, FANG Liangxing. Characteristics of the Antimicrobial Resistance and Dissemination of bla_CTX-M Genes Among E.coli from Geese Farms in Guangdong[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(5): 1571-1582.

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