耐环丙沙星鲍曼不动杆菌主动外排机制的研究及耐药逆转初步探讨

发布时间：2018-01-10 13:39

  本文关键词：耐环丙沙星鲍曼不动杆菌主动外排机制的研究及耐药逆转初步探讨　出处：《中南大学》2009年硕士论文　论文类型：学位论文

  更多相关文章： 鲍曼不动杆菌 环丙沙星 主动外排泵 抑制剂

【摘要】： 目的了解长沙地区鲍曼不动杆菌对环丙沙星的耐药现状,为临床治疗鲍曼不动杆菌感染、合理应用喹诺酮类抗生素提供指导;研究鲍曼不动杆菌对喹诺酮类药物的耐药性与主动外排机制的关系;初步探讨鲍曼不动杆菌对喹诺酮类药物的耐药逆转机制,寻求恢复其对喹诺酮类药物敏感性的方法。 方法(1)微量稀释法检测56株鲍曼不动杆菌对环丙沙星的敏感性。(2)采用直接荧光法筛选出胞内环丙沙星聚集量明显差异的菌株作为耐药组,并测定敏感组和耐药组在有无外排泵抑制剂情况下,鲍曼不动杆菌对环丙沙星的吸收和积累情况。(3)用逆转录-聚合酶链反应(RT-PCR)方法检测主动外排泵基因adeABC的表达水平。(4)扩增adeABC自身启动子区域片段基因并测序。(5)逆转实验观察三种抑制剂对外排泵的抑制作用。 结果(1)56株鲍曼不动杆菌中,对环丙沙星耐药的有48株(MIC＞4 mg/l);对环丙沙星中介有0株(1 mg/l≤MIC≤4 mg/l);对环丙沙星敏感的有8株(MIC＜1 mg/l)。(2)耐环丙沙星的所有菌株中,挑选胞内环丙沙星药物浓度累积差异最大的6株作为耐药组(R组),与敏感组(S组)相比较:无NaN_3时,S组的累积量(138.01±0.251 ng/ml)明显高于R组(62.15±0.743 ng/ml)(P＜0.01);加入NaN_3后,R组的累积量明显上升(114.78±0.415 ng/ml),与无NaN_3的S组(138.01±0.251 ng/ml)无差异(P＞0.05),但是与无NaN_3的R组(62.15±0.743 ng/ml)有明显差异(P＜0.05);加NaN_3后的S组(138.27±0.177 ng/ml)与未加NaN_3时(138.01±0.251 ng/ml)无明显差异(P＞0.05)。(3)耐药组主动外排基因adeB的表达水平(2.372±0.032)明显高于敏感组(1.654±0.040)(P＜0.01)。(4)adeABC自身启动子区域没有发现有意义突变。(5)分别在环丙沙星中加入外排泵抑制剂NaN_3、MC-207110或利血平后,临床分离敏感株MIC值变化不大;临床分离耐药株MIC均有所下降,且MIC值下降至原值1/4的株数(即外排阳性株)分别为6株、6株和2株。 结论:(1)长沙地区鲍曼不动杆菌对环丙沙星的耐药率高达85.7%。(2)adeABC基因的过度表达是鲍曼不动杆菌对环丙沙星耐药的重要原因之一。(3)在对环丙沙星耐药的鲍曼不动杆菌中,adeB自身启动子区域未见有意义突变。(4)使用外排泵抑制剂MC-207110和NaN_3后耐药株对环丙沙星的敏感性有显著的恢复;利血平对外排泵adeABC的抑制作用不如前两者明显。
[Abstract]:Objective to understand the Changsha area Bauman real status of resistance to ciprofloxacin for the treatment of Bauman coli, Acinetobacter infection, rational use of quinolones to provide guidance; the study of relationship between Bauman real resistance and active bacillus of quinolone efflux mechanism; preliminary study of Bauman Acinetobacter quinolone resistance reversal mechanism, seeking to restore the method of Quinolone drug sensitivity.
Methods (1) detection of trace dilution method in 56 strains of Acinetobacter strains to ciprofloxacin susceptibility to Bauman. (2) screened the intracellular accumulation of ciprofloxacin significantly different resistant strains as group by immunofluorescence assay and determination of sensitive group and resistant group in the presence of efflux pump inhibitors under Bauman real and accumulation the absorption of ciprofloxacin. Coli (3) by reverse transcription polymerase chain reaction (RT-PCR) pump gene expression adeABC method for detection of active efflux. (4) the amplification of adeABC promoter region gene fragment and sequencing. (5) observed the reversal of inhibition of three kinds of inhibitors efflux pump.
缁撴灉(1)56鏍�椴嶆浖涓嶅姩鏉嗚弻涓�,瀵圭幆涓欐矙鏄熻，

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