引起呼吸机相关性肺炎铜绿假单胞菌的耐药机制研究

发布时间：2018-06-25 05:43

本文选题：铜绿假单胞菌 + 多药耐药　；参考：《中国人民解放军军医进修学院》2012年硕士论文

【摘要】：目的：1、研究高龄患者呼吸机相关性肺炎的病原菌分布及耐药情况。2、探讨引起呼吸机相关性肺炎的铜绿假单胞菌耐药机制。3、对多药耐药铜绿假单胞菌进行体外联合药敏试验，，为临床合理用药提供依据。方法： 1、收集2007年1月～2011年6月诊断为VAP的高龄患者123例，研究其病原菌分布及耐药情况。 2、将两周之内，同一患者且对某一类抗菌药物药敏有变化的两株铜绿假单胞菌配为一对，其中碳青霉烯类20对，喹诺酮类22对，氨基糖苷类15对，多粘菌素B10对。收集来自30个高龄VAP患者的PA122株。 3、琼脂稀释法检测PA对14种抗生素的MIC、外排泵表型。PFGE法检测基因同源性。双纸片协同法检测金属酶表型。PCR法检测耐药基因。结晶紫染色法检测细菌生物被膜形成能力。扫描电镜观察生物被膜。棋盘法设计，琼脂稀释法行体外联合药敏。结果： 1、123例高龄老年VAP患者共检出细菌409株，其中革兰氏阴性菌355株，革兰氏阳性菌54株，革兰阴性菌中铜绿假单胞菌24.44%、鲍曼不动杆菌17.11%、嗜麦芽窄食单胞14.67%、洋葱伯克霍尔德菌7.58%;革兰阳性菌中以金黄色葡萄球菌9.05%为主。铜绿假单胞菌对常用抗菌药物的耐药性较高。 2、所收集的配对PA菌株共来自30个患者，其中70%的患者痰标本分离的配对菌株经PFGE证明为同一基因来源，30%的患者痰标本分离的配对菌株经PFGE证明为不同基因来源。 3、4个患者共8株菌扩增出OXA-1基因，3个患者共6株菌扩增出OXA-10基因，仅一个患者1株菌扩增出GES基因，13个患者共33株菌扩增出OprD2基因，余菌株为OprD2基因缺失或部分缺失菌株。未扩增出KPC、GIM、OXA-2、VIM及IMP耐药基因。同一患者来源且经PFGE证明为同一基因来源的菌株中，PCR法检测其耐药基因型如OXA-1、OXA-10、GES、OprD2并不完全一致。仅有一个患者分离的共4株菌金属酶表型阳性。5个患者共9株菌外排泵表型阳性。所有菌株均有不同程度的生物膜形成能力。 4、各组药物联用后，除TZP和CIP联合主要表现为无关作用外，其他组合均主要表现为协同和相加作用，浓度-累积抑菌百分率曲线均左移。结论：（1）PA在高龄VAP患者中分离率占首位，且高度耐药。（2）70%老年PA-VAP患者自身不同时间点所分离的PA为同一基因来源。（3）PA的高度耐药为以产生物膜为主的多种机制共同作用结果。（4）MDRP可选联合用药方案。
[Abstract]:Objective to study the distribution and drug resistance of pathogenic bacteria in ventilator-associated pneumonia (VAP) in elderly patients, and to explore the mechanism of resistance of Pseudomonas aeruginosa to multidrug resistant Pseudomonas aeruginosa in vitro, and to investigate the mechanism of drug resistance of Pseudomonas aeruginosa, and to study the drug resistance of Pseudomonas aeruginosa in vitro. To provide the basis for clinical rational use of drugs. Methods: 1. 123 elderly patients diagnosed as VAP from January 2007 to June 2011 were collected to study the distribution of pathogenic bacteria and drug resistance. Two strains of Pseudomonas aeruginosa were matched in the same patient, including 20 pairs of carbapenems, 22 pairs of quinolones, 15 pairs of aminoglycosides and 10 pairs of polymyxin B _ (10). PA122 strains from 30 old patients with VAP were collected. The MICs of PA to 14 antibiotics were detected by Agar dilution method. The homology of gene was detected by efflux pump phenotype. PFGE method. Detection of metallozyme phenotypes by double disk synergy. PCR was used to detect drug resistance genes. The ability of bacterial biofilm formation was detected by crystal violet staining. The biofilm was observed by scanning electron microscope. Chessboard design, Agar dilution method combined with in vitro drug sensitivity. Results: a total of 409 strains of bacteria were detected in 1123 elderly patients with VAP, of which 355 were Gram-negative bacteria and 54 were Gram-positive bacteria. Pseudomonas aeruginosa 24.44m, Acinetobacter baumannii 17.11m, maltophilia 14.677m, Bacillus cepacia 7.58m, Gram-positive bacteria 9.05%. Pseudomonas aeruginosa has higher resistance to common antimicrobial agents. 2. Pairing PA strains were collected from 30 patients. Among them, 70% of the matched strains isolated from sputum samples from patients were identified as the same gene source by PFGE, and 30% of them were identified as different gene sources by PFGE. OXA-1 was amplified from 8 strains from 34 patients. OXA-10 gene was amplified from 6 strains of 3 patients. GES gene was amplified from one strain of one patient and OprD2 gene was amplified from 33 strains of 13 patients. The remaining strains were OprD2 gene deletion or partial deletion. KPC-GIMA OXA-2 vim and IMP resistance genes were not amplified. The genotypes of OXA-1, OXA-10, GESS-OprD2, which were identified by PFGE as the same gene source, were not identical with each other by polymerase chain reaction (PCR). Only 4 strains were positive for metallozyme phenotype and 9 strains were positive for efflux pump phenotype in 5 patients. All the strains had different biofilm forming ability. 4. After combined use of TZP and CIP, the combination of TZP and CIP were mainly synergistic and additive. The curves of concentration-cumulative bacteriostatic percentage were all shifted to the left. Conclusion: (1) the isolation rate of PA in the elderly patients with VAP is the highest. And high drug resistance. (2) PA isolated from 70% old PA-VAP patients at different time points was the same gene source. (3) the high resistance of PA was the result of multiple mechanisms of biofilm production. (4) MDRP could be used in combination.
【学位授予单位】：中国人民解放军军医进修学院
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R563.1

【参考文献】