乙酰唑胺对万古霉素耐药肠球菌体内外抗菌活性作用和机制  

Study on the antibacterial activity and mechanism of acetazolamide againstEnterococcus in vitro and in vivo

作　　者：庞茜茜 刘云宁[1] 张艺 姜爱雯[1] 刘斐[2] 汤建华[1] PANG Xiqian;LIU Yunning;ZHANG Yi;JIANG Aiwen;LIU Fei;TANG Jianhua(Pharmacy Department,The First Affiliated Hospital of Hebei North University,Zhangjiakou 075000,Hebei,China;Critical Care Medicine,The First Affiliated Hospital of Hebei North University,Zhangjiakou 075000,Hebei,China)

机构地区：[1]河北北方学院附属第一医院药学部,河北张家口075000 [2]河北北方学院附属第一医院危重病医学,河北张家口075000

出　　处：《西部医学》2024年第7期968-973,共6页Medical Journal of West China

基　　金：张家口市市级科技计划自筹经费项目(2221070D)。

摘　　要：目的探讨乙酰唑胺对万古霉素耐药肠球菌(VRE)体内外抗菌活性作用和可能的作用机制。方法实验菌株为医院临床分离的肠球菌菌株,经鉴定为VRE菌株,分别进行体外和体内抗菌活性研究。采用琼脂平板稀释法测定乙酰唑胺对VRE的最小抑菌浓度(MIC)和最小杀菌浓度(MBC),杀菌曲线实验测定待测菌株的时间杀菌变化曲线,检测胞外碱性磷酸酶(AKP)含量评估对细胞壁的破坏作用,计算相对电导率测定评估细胞膜渗透性。使用临床分离的VRE菌株建立腹膜炎小鼠模型,分为对照组(正常小鼠),模型组(腹膜炎小鼠)和乙酰唑胺组(30 mg/kg/5 h干预),每组10只。干预30 h后统计各组小鼠死亡情况,进行腹膜液、脾脏、肝脏菌落计数,HE染色观察脾脏、肝脏病理改变。结果乙酰唑胺对VRE的抑菌活性检测显示,MIC和MBC分别为1.45 mg/L和4.62 mg/L。Time-Kill曲线分析乙酰唑胺杀菌动力学显示,乙酰唑胺杀菌活性与时间和浓度呈正相关,乙酰唑胺浓度越大,细菌死亡率越高,时间越长,杀死的细菌越多。当乙酰唑胺浓度达到16×MIC,作用时间达到12 h时,VRE细菌生长被完全抑制。干预30 h后,对照组小鼠均存活,存活率100%;模型组小鼠7只陆续死亡,存活率30%;乙酰唑胺组小鼠2只出现死亡,存活率80%,存活率明显高于模型组(P<0.05)。相较于对照组,模型组小鼠腹膜液、肝脏、脾脏菌载量均明显提高(P<0.05);相较于模型组,乙酰唑胺组小鼠腹膜液、肝脏、脾脏菌载量均明显降低(P<0.05)。与对照组比较,1×MIC组和4×MIC组AKP含量显著升高(P<0.05);与1×MIC组比较,4×MIC组AKP含量显著升高(P<0.05),乙酰唑胺对VRE细胞壁的破坏作用呈现剂量依赖。与对照组比较,1×MIC组和4×MIC组电导率显著升高(P<0.05);与1×MIC组比较,4×MIC组相对电导率显著升高(P<0.05),乙酰唑胺对VRE细胞渗透性的提高作用呈剂量依赖。结论乙酰唑胺对VRE具有较好的体内、体�Objective To explore the in vitro and in vivo antibacterial activity of acetazolamide against vancomycin resistant enterococci,and investigate its possible mechanism of action.Methods The study object was Enterococcus(N1520),which was clinically isolated from the hospital and identified as vancomycin resistant Enterococcus(VRE).The antibacterial activity in vitro and in vivo were studied respectively.Determination of minimum inhibitory concentration(MIC)and minimum bactericidal concentration(MBC)of acetazolamide on VRE using agar plate dilution method.Determination of the Time Sterilization Change Curve of the Strain to be Tested through the Sterilization curve experiment.Detection of extracellular alkaline phosphatase(AKP)content to evaluate the destructive effect on cell walls,calculation of relative conductivity measurement to evaluate cell membrane permeability.Establishing a mouse model of peritonitis using clinically isolated VRE strains,divided into a control group(normal mice),a model group(peritonitis mice),and an acetazolamide group(30mg/kg/5h intervention).After 30 hours of intervention,the death status of each group of mice was counted,and bacterial colonies in peritoneal fluid,spleen,and liver were counted.HE staining was used to observe pathological changes in the spleen and liver.Results The antibacterial activity test of acetazolamide on VRE showed that MIC and MBC were 1.45 mg/L and 4.62 mg/L,respectively.Time Kill curve analysis showed that the bactericidal activity of acetazolamide was positively correlated with time and concentration.The higher the concentration of acetazolamide,the higher the bacterial mortality rate.The longer the time,the more bacteria were killed.When the concentration of acetazolamide reaches 16×MIC,when the action time reaches 12 hours,the growth of VRE bacteria was completely inhibited.After 30 hours of intervention,the survival rate of the blank control group was 100%,the model group was 30%,and the acetazolamide group was 80%.The survival rate of mice in the acetazol

关 键 词：乙酰唑胺 万古霉素耐药 肠球菌 抗菌活性 作用机制 

分 类 号：R915[医药卫生—微生物与生化药学] R965[医药卫生—药学]