可疑产碳青霉烯酶肠杆菌科细菌的耐药基因研究

Study of Resistant Gene on Suspicious Carbapenemase Producing Enterobacteriaceae

目的 分析临床分离的可疑产碳青霉烯酶肠杆菌科细菌所携带的耐药基因.方法 采用全自动细菌鉴定仪进行药敏试验,用改良Hodge试验进行碳青霉烯酶筛选,用纸片扩散试验进行ESBLs表型检查,用头孢西丁三维试验进行AmpC β-内酰胺酶的表型检查,用聚合酶链反应检测β内酰胺酶基因、外膜蛋白（OmpK35和OmpK36）编码基因、转座子tpuA和tpuU基因,并进行产物测序.结果 ①62株可疑产碳青霉烯酶肠杆菌科细菌对厄他培南和氨苄西林100％耐药,对氨苄西林/舒巴坦、头孢唑啉、头孢他啶、头孢曲松、氨曲南的耐药率达87％以上,对亚胺培南、美洛培南、丁胺卡那霉素耐药率为35％以下;②β内酰胺酶的总检出率为75.81％,主要由同时产ESBLs和AmpC引起,检出率为33.87％,其次为单产ESBLs引起,检出率为30.65％;③检出KPC,DHA,CIT,TEM,CTX M,SHV和NDM1β内酰胺酶基因的阳性率分别为8.06％,14.51％,24.19％,25.81％,27.42％,40.32％和6.45％; Ompk35,Ompk36基因缺失率为38.70％和62.90％;tnpA和tnpU检出率分别为11.29％和32.25％.结论 产碳青霉烯酶并非菌株耐碳青霉烯类药物的主要原因,可能由产ESBLs或AmpC酶引起,合并外膜蛋白缺失,尚存在其它降低碳青霉烯类敏感性的耐药机制.

Objective To investigate the resistant gene on suspicious carbapenemase producing Enterobacteriaceae of clinical isolates.Methods Drug sensitivity tests were done by automatic bacterium identification instrument.Carbapenemase was de tected by Modified Hodge tests.Extended spectrum β lactamase was detected by disc diffusion test.AmpC beta lactamase was tested by cefoxitin three dimension test,and the polymerase chain reaction （PCR） was carried out to amplity the gene of β lactamase,outer membrane protein genes K35 and K36,transposon A and U,then DNA product were sequenced.Results ①Among 62 strains of suspicious carbapenemase producing Enterobacteriaceae,the drug resistance rates to Ertapenem and ampicillin were 100％,the resistance rates to Ampicillin/sulbactam,cefazolin,ceftazidime,ceftriaxone,aztreonam were as high as 87 ％ above,the resistance rates to amikacin,imipenem,meropenem were less than 35 ％.②The positive rate of β lac tamase phenotype was 75.81 ％ in 62 strains,which was caused by carrying ESBLs and AmpC at the same time,and the posi tive rate was 33.87 ％ ;secondly by carrying ESBLs and the positive rate was 30.65 ％.③The positive rates of KPC,DHA,CIT,TEM,CTXM,SHV and NDM 1gene were 8.06％,14.51％,24.19％,25.81％,27.42％,40.32％,6.45％,respective ly.The positive rates of deficiency of Ompk35 and Ompk36 were 38.70％ and 62.90％.The positive rates of tnpA and tnpU were 11.29％ and 32.25％.Conclusion Producing carbapenemase would not be the main reason of carbapenem resistant,which could be caused by carrying ESBLs or AmpC,combining deficiency of outer membrane protein genes,and there may be other resistant mechanisms to reduced susceptibility of carbapenem.