环丙沙星联合妥布霉素对兔铜绿假单胞菌环丙沙星耐药突变选择窗的影响

Combined application of ciprofloxacin and tobramycin on mutant selective windows of ciprofloxacin against Pseudomonas aeruginosa

目的 以兔组织笼感染模型观察环丙沙星联合妥布霉素,能否缩小环丙沙星对铜绿假单胞菌的耐药突变选择窗.方法 体外测定环丙沙星与妥布霉素对铜绿假单胞菌标准菌株ATCC27853的最低抑菌浓度（MIC）、防耐药突变浓度（MPC）、突变选择窗（MSW,MPC～MIC）、突变选择指数（SI,MPC/M1C）,同时棋盘格法测定联合后的MIC.建立兔组织笼模型6只,高效液相色谱法测定环丙沙星在兔组织液中药代动力学参数.兔组织笼感染模型55只,随机平均分为11组,1组为牛理盐水组（对照组）,5组单用环丙沙星（环丙沙星组）,5组为环丙沙星联合妥布霉素（联合组）.环丙沙星10次/d给药,每次给药间隔2 h,使兔组织液环丙沙星稳态浓度分别达到0.25、0.5、1.0、2.0和4.0 mg/L,即5个亚组;妥布霉素2.0 mg·kg^-1～·d^-1,其组织液峰浓度达到2.0 mg/L左右.3 d后抽取组织液,涂布于含有0.25 ms/L浓度环丙沙星的琼脂平板,有细菌生长者为耐药突变体.结果 体外环丙沙星与妥布霉素对铜绿假单胞菌的MIC、MPC、S1分别为0.25 mg/L、4.0 mg/L、16与0.25 mg/L、8.0 mg/L、32.环丙沙星组中环丙沙星浓度0.25、0.5、1.0及2.0 mg/L亚组组织液中均有耐药突变体生长,4.0 mg/L亚组无耐药突变体牛长;即体内环丙沙星MPC与体外相同,SI为16.联合组中环丙沙星浓度0.25 mg/L业组有耐药突变体生长,0.5、1.O、2.0及4.0 mg/L亚组无耐药突变体生长;即联合妥布霉素后环丙沙星的MPC为0.5 mg/L,MIC为0.125 mg/L,因此SI为4.结论 环丙沙星联合妥布霉素可缩小环丙沙星对铜绿假单胞菌的耐药突变选择窗,减少耐药突变体的产生,有利于减少细菌耐药.

Objective To observe whether the mutant selective windows（MSW）of ciprofloxacin would be reduced after its combination against Pseudomonas aeruginosa in rabbits. Methods Firstly the minimal inhibitory concentration（MIC）,mutant prevention concentration（MPC）,mutant selective windows （MSW,MPC-MIC）and selective indices（SI,MPC/MIC）of ciprofloxacin and tobramycin were measured in vitro respectively with standard strain ATCC27853. And the MIC was detected for the combination of ciprofloxacin and tobramycin. The rabbit tissue cage model was constructed to determine the pharmacokinetic parameters of ciprofloxacin by HPLC（high performance liquid chromatography）. Fifty-five rabbits were randomly divided by a random number table into 11 groups：physiological saline in 1 group,ciprofloxacin alone in 5 groups and ciprofloxacin plus tobramycin in another 5 groups. The rabbits received ciprofloxacin 10 times a day at a 2-hour dosing interval. In 2 dosing groups. the steady state concentrations of ciprofloxacin reached to 0. 25,0. 5,1. 0,2. 0 and 4. 0 me,/L respectively. The dose of tobramycin was 2. 0 mg·kg^-1·d^-1and its peak concentration reached around 2.0 mg/L. At Day 3,the tissue juice was extracted, diluted and coated on agar plates with ciprofloxacin at a concentration of 0. 25 ms/L so as toobserve the growing condition of mutants. Results Against Pseudomonas aeruginosa,the values of MIC,MPC and SI of ciprofloxacin were 0. 25 mr,/L,4. 0 mg/L and 16 while 0. 25 mg/L,8. 0 mg/L and 32 for tobramycin respectively. Single groups：the mutants were found in 0. 25,0. 5,1. 0 and 2. 0 mg/L groups,but none in 4. 0 mg/L group. The MPC of cipmfloxacin was the same for in vivo and in vitro. Both were at 16. Combination groups： the mutants were only found in the group with a concentration of ciprofloxacin at 0. 25 mg/L while no mutants in the other groups. And MPC was 0. 5 mg/L and MIC 0. 125 mg/L for ciprofloxacin plus tobramycin. And the value of SI was 4. Conclusion The combined use of ciprofloxacin and tobramycin may reduce the mutant selective windows of ciprofloxacin against P. aeruginosa in rabbits so as to reduce the occurrence of mutants to control its drug resistance.