庆大霉素诱导粪肠球菌耐药及其对氯霉素协同敏感相关特性研究  

Study on the characteristics of collateral sensitivity to chloramphenicol of evolved gentamicin-resistant Enterococcus faecalis

作　　者：孙瑶[1] 徐雯雅 周翠[1] 吴庆[1] 陈栎江[1] 徐春泉[1] 叶建中[1] 周铁丽[1] Sun Yao;Xu Wen-ya;Zhou Cui;Wu Qing;Chen Li-jiang;Xu Chun-quan;Ye Jian-zhong;Zhou Tie-li(Laboratory Medicine Center,the First Affiliated Hospital of Wenzhou Medical University,Wenzhou 325000)

机构地区：[1]温州医科大学附属第一医院医学检验中心,温州325000

出　　处：《中国抗生素杂志》2023年第2期228-234,共7页Chinese Journal of Antibiotics

基　　金：国家自然科学基金青年科学基金项目(No.81802069)。

摘　　要：目的通过研究庆大霉素诱导粪肠球菌耐药突变株对氯霉素协同敏感(collateral sensitivity,CS)的相关特性,为基于协同敏感治疗策略的临床应用提供依据。方法采用庆大霉素通过梯度浓度平板对3株庆大霉素敏感的粪肠球菌临床分离株和1株粪肠球菌标准菌株ATCC29212进行体外实验诱导耐药(experimental evolution);通过微量肉汤稀释法测定庆大霉素诱导后耐药突变株对常见抗菌药物的最低抑菌浓度(minimum inhibitory concentrations,MICs),分析抗菌药物间交叉敏感性;PCR检测诱导耐药菌株氨基糖苷类修饰酶编码基因携带情况;通过琼脂平板稀释法测定庆大霉素诱导耐药(evolved gentamicin-resistant)菌株和亲本菌株对其协同敏感药物(氯霉素)的防耐药突变浓度(mutant prevention concentration,MPC),最后,测定氯霉素对庆大霉素诱导耐药菌株和亲本菌株的时间杀菌曲线。结果结果显示4株粪肠球菌亲本菌株经过庆大霉素诱导后,对庆大霉素MICs升高了16~64倍,并对阿米卡星表现出交叉耐药;诱导耐药菌株及其亲本菌株均未检出aac(6’)-Ie-aph(2")-Ia。此外,研究发现诱导后菌株对氯霉素表现出协同敏感。氯霉素对庆大霉素诱导耐药粪肠球菌及其亲本菌株的MPC均大于1024μg/mL,突变选择窗较宽;时间杀菌曲线显示当氯霉素浓度为亲本菌株的1/2×MIC或1/4×MIC时,庆大霉素诱导耐药菌株的生长被有效抑制,而亲本菌株则不能被抑制。结论基于协同敏感性制定抗菌药物治疗策略可能有助于有效清除耐药病原菌并延长抗菌药物的使用寿命。Objective To study the characteristics of collateral sensitivity to chloramphenicol of evolved gentamicin-resistant Enterococcus faecalis,and to provide foundation for clinical application of treatment strategies based on collateral sensitivity(CS).Methods Three clinically isolated antimicrobial susceptible Enterococcus faecalis and E.faecalis standard strain ATCC29212 were used to adaptation laboratory evolution in vitro with gentamicin gradient concentration plates.The minimum inhibitory concentrations(MICs)of evolved gentamicin-resistant strains were determined by the microbroth dilution method,and the cross susceptibilities between antimicrobials were analyzed.The prevalence of aminoglycoside modifying enzyme encoding gene was detected by PCR in the evolved resistant strains.The agar plate dilution method was performed to determine the mutant prevention concentration(MPC)of evolved gentamicin-resistant strains and parental strains to the collateral sensitivity drug(chloramphenicol).Finally,the time-kill curve of chloramphenicol on the strains before and after evolution of gentamicin was determined.Results The results showed that the MICs of four strains of E.faecalis were increased 16~64 fold after evolution to gentamicin,and they showed cross-resistance to Amikacin.aac(6’)-Ie-aph(2")-Ia had not been detected in all evolved resistant strains and their parental strains.Interestingly,the evolved resistant strains exhibited collateral sensitivity to chloramphenicol.The MPC of chloramphenicol in E.faecalis before and after evolution to gentamicin was higer than 1,024μg/mL,indicating a wide mutant selection window(MSW).The timekill curve showed that when the concentration of chloramphenicol was 1/2×MIC or 1/4×MIC of the ancestral strain,the evolved resistant strains were effectively inhibited,while the parental strains could not be effectively inhibited.Conclusion Antimicrobials therapy strategies based on collateral sensitivity possibly help to effectively eliminate drug-resistant pathogens and prolong the l

关 键 词：粪肠球菌 协同敏感 庆大霉素 氯霉素 诱导耐药 

分 类 号：R978.1[医药卫生—药品]