亚抑菌浓度氟苯尼考作用下质粒pSD11对大肠杆菌适应性的影响

Effect of plasmid pSD11 on the fitness of Escherichia coli in sub-minimum inhibitory concentration of florfenicol

通过分析cfr阳性质粒pSD11对大肠杆菌的适应性影响,并测定氟苯尼考对携带质粒pSD11的大肠杆菌MG1655(Escherichia coli MG1655)的最低筛选浓度(MSC),评价亚抑菌浓度氟苯尼考作用下多重耐药基因cfr在大肠杆菌中的持续存在和扩散传播的风险.通过电转化的方法将质粒pSD11导入模式菌E.coli MG1655中,获得耐药菌株E.coli MG1655/pSD11.通过测定生长曲线,比较无抗生素作用下耐药菌株E.coli MG1655/pSD11与同源敏感菌株E.coli MG1655的生长动力学参数,评价质粒pSD11对宿主菌适应性的影响.并通过比较分析不同亚抑菌浓度(0.031 25,0.062 5,0.125,0.25,0.5,1.0和2.0μg/mL)氟苯尼考作用下的生长速率,测定氟苯尼考对携带质粒pSD11的宿主菌最小筛选浓度(MSC).携带质粒pSD11的耐药菌E.coli MG1655/pSD11的延滞时间(2.853 3±0.031 8 h/λ)相对于同源敏感菌E.coli MG1655极显著延长(2.825 3±0.024 4 h/λ),且世代时间也显著延长(分别为:1.705 7±0.006 4 h/G和1.526 9±0.007 8 h/G).氟苯尼考对耐药菌E.coli MG1655/pSD11的MSC为0.042 5μg/mL约为相应敏感菌的MICsus的1/100.本研究表明携带cfr基因的质粒pSD11在无抗生素选择下对宿主菌有明显的适应性影响,但是在极低的氟苯尼考浓度的作用下存在选择优势,能够平衡适应性成本使宿主菌得到富集与传播,以期为对该质粒在环境中的传播风险评估及氟苯尼考的临床应用提供的参考.

The sustainability and spread of the multi-resistant gene cfr in Escherichia coli was evaluated by analyzing the fitness effect of the cfr-carrying plasmid pSDll in E.coli,and determining the minimum selectable concentration(MSC) of florfenicol for pSD11-harboring E.coli.Plasmid pSDll was transferred into E.coli MG1655 by electroporation to obtain the drug-resistant strain E.coli MG1655/pSDll.The growth kinetics parameters of this strain and the homologous sensitive strain E.coli MG1655 were compared by measuring the growth curve without antibiotics.The MSC for strain E.coli MG1655/pSDll was determined by analyzing and comparing the growth rates of strain E.coli MG1655/pSDll and E.coli MG1655 at different sub-minimum inhibitory concentrations(0.031 25,0.062 5,0.125,0.25,0.5,1.0,and 2.0 μg/mL) of florfenicol.The lag time of the drug-resistant with plasmid pSDll strain E.coli MG1655/pSDll(2.853 3±0.031 8 h/λ) is significantly longer than that the homologous sensitive E.coli MG1655(2.825 3±0.024 4 h/λ),and the generation time was also significantly longer(1.705 7±0.006 4 h/G and 1.526 9±0.007 8 h/G,respectively).The MSC of florfenicol for strain E.coli MG1655/pSD11 was 0.042 5μg/mL,which was 1/100 of the MICsus value of the homologous sensitive strain E.coli MG1655.The cfr-carrying plasmid pSDll has a significant adaptive effect on the host bacteria.However,it can also possess the host strain of selective advantage under extremely low concentration of florfenicol,which can balance the fitness cost of plasmid and benefit the enrichment and transmission of host bacteria.