耐碳青霉烯类铜绿假单胞菌的耐药机制及同源性研究

The investigation of drug resistance mechanism and homology for carbapenem-resistant Pseudomonas aeruginosa

目的分析柳州市4所医院耐碳青霉烯类铜绿假单胞菌的耐药机制及同源性,为指导临床合理用药、控制感染提供依据。方法 69株耐碳青霉烯类铜绿假单胞菌分离自柳州市工人医院(甲医院),中医院(乙医院)、妇幼保健院(丙医院)和柳铁中心医院(丁医院)。采用纸片扩散法及微量肉汤稀释法进行药敏试验,采用脉冲场凝胶电泳(PFGE)分析菌株同源性,聚合酶链反应(PCR)检测β-内酰胺酶基因和OprD2基因,使用氰氯苯腙(CCCP)进行主动外排泵筛选试验。结果 69株铜绿假单胞菌对多黏菌素B和氨基糖苷类的耐药率较低,对头孢他定、头孢哌酮/舒巴坦和哌拉西林/他唑巴坦的耐药率分别为40.6%、40.6%、44.9%,其他抗菌药物的耐药率均>50%;β-内酰胺酶基因IMP阳性率为4.3%,OprD2基因阳性率为63.8%,VIM、KPC、GIM、OXA和SPM基因均未检出;主动外排表型阳性率为82.6%。PFGE分型分为A^J共10个基因型,其中A型为主要基因型,占44.1%;4所医院均存在克隆传播,甲、乙、丙、丁医院分别以A型、H型、D型、C型克隆株传播为主,甲、乙医院有共同的B型克隆株存在,甲、丁医院有共同的A型克隆株存在。结论克隆株在医院内、医院间播散已成为柳州市耐碳青霉烯类铜绿假单胞菌分离率不断上升、菌株不断增加的主要原因,主动外排泵高表达、OprD2蛋白表达缺失和产IMP型金属酶是该菌对碳青霉烯类药物耐药的主要机制。

Objective To analyze drug resistance and homology of carbapenem-resistant Pseudomonas aeruginosa of four hospitals in Liuzhou city for providing the basis of the clinical rational use of drugs and controling infection. Methods 69 strains of carbapenem-resistant Pseudomonas aeruginosa were isolated from Worker＇s Hospital （A Hospi- tal）,Hospital of Traditional Chinese Medicine （B Hospital）, Maternal and Child Health Hospital （C Hospital） and Liu Railway Central Hospital （D Hospital）.Drug sensitive test was conducted by by disk diffusion and broth micro dilution, the homology of bacterial strain was analyzed by pulsed-field gel electrophoresis （PFGE）,gene of β-1actamase and OprD2 was detected by PCR,active efflux pump screening test was conducted by CCCP. Results In 69 strains of Pseu- domonas aeruginosa,drug resistance rate of polymyxin B and aminoglycoside was lower, cefiazidime,cefoperazone / shu- batan and piperacillin/tazobactam was 40.6%,40.6%,44.9% respectively,other antibiotic＇s resistance rate was 〉50%;the positive rate of β-lactamase gene IMP was 4.3% and the positive rate of OprD2 was 63.8%,strains positive for gene VIM,KPC,GIM,OXA and SPM were not detected;the positive rate of active efflux phenotype was 82.6%.There were 10 genotype by PFGE named A-J,the major A type was 44.1%;four hospitals （ A,B,C.and D Hospital） all contained clone transmission and clone strain transmission of A,B,C and D Hospital was mainly type A,type H,type D,type C respec- tively,both A and B Hospital exsited type B clone strain,both A and D Hospital existed type A clone strain. Conclu- sion Clone strain spread in the hospital and among hospitals has become the main reason of the isolated rate of car- bapenem-resistant Pseudomonas aeruginosa rising,and strain increasing in Liuzhou city,the main mechanism of drug resistance of Pseudomonas aeruginosa to carbapenem drugs was high expression of active efflux pump,OprD2 protein expression loss and production IMP type metal enzyme.