耐碳青霉烯类鲍曼不动杆菌耐药机制研究

目的了解碳青霉烯类耐药鲍曼不动杆菌的耐药性、同源性及其耐药机制。方法收集碳青霉烯类耐药鲍曼不动杆菌20株,用E试验行药敏测定;脉冲场凝胶电泳(PFGE)行同源性分析;通过等电点聚热电泳(IEF)、三维试验及抑制试验、PCR、克隆测序、十二烷基硫酸钠-聚丙酰胺凝胶电泳(SDS-PAGE)等方法分析碳青霉烯酶型及外膜蛋白(OMP)。结果20株碳青霉烯类耐药鲍曼不动杆菌对多黏菌素B仍保持良好的敏感性(90%),莫西沙星对约40%菌株的MIC在0.125mg/L以下;根据PFGE图谱将其分为5型,A1型是主要流行株(9株),主要集中在我校附属二院ICU和附属一院呼吸科;所有菌株均产等电点6.7的碳青霉烯酶,PCR方法及测序证实为OXA-23型碳青霉烯酶,未发现金属酶,且都含有Ⅰ类整合子,但未发现质粒;OMP分析发现部分克隆有31ku的条带丢失。结论亚胺培南耐药不动杆菌存在医院间克隆传播,值得关注;我校鲍曼不动杆菌主要产生OXA-23型碳青霉烯酶;OMP31ku的条带丢失是耐药机制之一;我校耐碳青霉烯类鲍曼不动杆菌耐药机制可能是产酶和OMP丢失共同作用。

Study on the mechanism of carbapenem resistance in Acinetobacter baumannii

Objective To investigate the resistance pattern, homology and mechanism of carbapenem resistance in Acinetobacter baumannii. Methods Twenty carbapenem-resistant A. baumannii (CRAB) isolates collected from 2 teaching hospitals of China Medical University were evaluated for antibiotic sensitivity by E-test, homology by pulsed-field gel electrophoresis (PFGE), and carbapenemases by isoelectric focusing electrophoresis (IEF), three dimensional tests, polymerase chain reaction (PCR), cloning and sequencing methods. Outer membrane proteins were extracted by ultrasound physical method and analyzed by SDS-PAGE. Results CRAB were highly susceptible to polymyxin B. Only 5% of the isolates were resistant. The MIC of moxifloxacin was lower than 0.125 mg/L for 40% of the CRAB isolates. According to PFGE, 20 CRAB isolates were typed as A, B, C, D and E, of which 9 belonged to type A1, which were from ICU and Department of Respiratory Medicine in the 2 teaching hospitals. All isolates produced OXA-23 carbapenemases with pI 6.7. Metallo-beta-lactamase was not detected. OMP analysis showed that a protein of 31 ku was missing in some isolates. Conclusions There was an endemic of CRAB in CMU teaching hospitals. OXA-23 is the major OXA-type carbapenemase of CRAB. Depletion of 31 ku OMP might be one of the mechanisms underlying carbapenems resistance. Both OXA-23 and OMP deletion might contribute to the development of CRAB.