NICU肺炎克雷伯菌质粒介导的耐药基因研究
发布时间：2018-12-17 03:43
【摘要】：目的了解肺炎克雷伯菌(Klebsiella pneumoniae,KP)在神经重症加强治疗病房(Neurological intensive care unit,NICU)环境中的存在情况,检测NICU环境中及感染患者标本中肺炎克雷伯菌质粒上携带耐药基因的具体情况,并分析肺炎克雷伯菌质粒上携带的耐药基因与菌株耐药表型的关系,指导临床抗菌药物的使用和NICU环境的消毒灭菌工作,从而降低NICU中患者肺炎克雷伯菌的感染发生率。方法对2015年1月-2015年12月在医院NICU内入住的患者进行感染监测,并对患者居住的常规环境及感染环境进行病原学监测,收集住院感染患者标本中及在环境中定植的肺炎克雷伯菌,检测其临床耐药表型并保留菌株。将收集到的肺炎克雷伯菌接种于LB肉汤培养基中进行培养,然后使用质粒小提试剂盒提取肺炎克雷伯菌内的质粒DNA,再以质粒DNA为模版通过聚合酶链反应方法对肺炎克雷伯菌菌株质粒上携带的β-内酰胺类、氨基糖苷类和喹诺酮类耐药基因进行检测。结果2015年1月-2015年12月期间,在NICU病房环境中,革兰氏阳性菌占主导地位,其次为少量的革兰氏阴性菌和真菌,分布在环境中的病原菌与NICU内感染患者送检标本中检出的病原菌大致一致。其中,于常规环境中检测到1株肺炎克雷伯菌,并于感染患者居住的感染环境中检测出5株肺炎克雷伯菌。研究期间NICU入住患者共259人,其中发生感染179例,在其送检的病原学标本中共检出肺炎克雷伯菌31株。将NICU感染患者送检标本中分离出的31株肺炎克雷伯菌及病房环境中分离出的6株肺炎克雷伯菌接种于LB肉汤培养基中培养并进行细菌质粒DNA的提取,发现37株肺炎克雷伯菌中均存在质粒。同时对每株肺炎克雷伯菌质粒上是否携带β-内酰胺类、氨基糖苷类和喹诺酮类21种耐药基因进行检测。感染患者标本中分离出的31株肺炎克雷伯菌内质粒中均检出SHV和rmt B耐药基因(100%),而且质粒携带的耐药基因均在两种以上,其中有8株肺炎克雷伯菌的质粒同时携带β-内酰胺类和氨基糖苷类耐药基因,其余23株均同时携带β-内酰胺类、氨基糖苷类和喹诺酮类耐药基因。在病房环境标本中分离出的6株肺炎克雷伯菌的质粒中也均检出SHV和rmt B耐药基因(100%),而且质粒携带的耐药基因也均在两种以上,其中有5株肺炎克雷伯菌的质粒同时携带β-内酰胺类和氨基糖苷类耐药基因,仅有1株肺炎克雷伯菌的质粒上同时携带β-内酰胺类、氨基糖苷类和喹诺酮类耐药基因。结论NICU内患者感染和环境中检出的肺炎克雷伯菌内均存在质粒,同一质粒上可同时携带有SHV、TEM、IMP、rmt B、qnr B、qnr S和acc(6)-Ib-cr等多个β-内酰胺类、氨基糖苷类和喹诺酮类耐药基因,且每株肺炎克雷伯菌质粒上均携带有SHV和rmt B类基因,但是肺炎克雷伯菌内质粒所携带的耐药基因并不与其临床耐药表型完全成正比关系。
[Abstract]:Objective to investigate the presence of Klebsiella pneumoniae (Klebsiella pneumoniae,KP) in (Neurological intensive care unit,NICU, a neurointensive intensive care unit. To detect the drug resistance genes of Klebsiella pneumoniae in NICU environment and infected patients, and to analyze the relationship between drug resistance genes and phenotypes of Klebsiella pneumoniae. In order to reduce the incidence of Klebsiella pneumoniae infection in patients with NICU, we should guide the use of clinical antibiotics and sterilize the environment of NICU. Methods the patients admitted to hospital NICU from January 2015 to December 2015 were monitored for infection, and the routine environment and infection environment were monitored by etiology. Klebsiella pneumoniae (Klebsiella pneumoniae) was collected from inpatients with infection and in the environment to detect the phenotype of clinical drug resistance and to retain the strain. The collected Klebsiella pneumoniae was cultured in LB broth medium, and then the plasmid DNA, was extracted by plasmid extraction kit. The 尾 -lactam, aminoglycoside and quinolone resistance genes carried on the plasmid of Klebsiella pneumoniae were detected by polymerase chain reaction using plasmid DNA as template. Results from January 2015 to December 2015, Gram-positive bacteria dominated the NICU ward environment, followed by a small number of Gram-negative bacteria and fungi. The pathogens distributed in the environment were approximately the same as those detected in the specimens of patients with NICU infection. One strain of Klebsiella pneumoniae was detected in the normal environment and 5 strains of Klebsiella pneumoniae were detected in the infected environment. A total of 259 patients were admitted to NICU during the study, of which 179 cases were infected. A total of 31 strains of Klebsiella pneumoniae were detected in the pathogenic specimens. 31 strains of Klebsiella pneumoniae isolated from NICU infected patients and 6 strains of Klebsiella pneumoniae isolated from ward environment were inoculated in LB broth culture medium to extract bacterial plasmid DNA. Plasmids were found in all 37 strains of Klebsiella pneumoniae. At the same time, 21 resistant genes of 尾-lactams, aminoglycosides and quinolones were detected on the plasmids of Klebsiella pneumoniae. SHV and rmt B resistance genes were detected in 31 strains of Klebsiella pneumoniae isolated from infected patients (100%). Among them, 8 strains of Klebsiella pneumoniae carried 尾 -lactam and aminoglycoside resistance genes simultaneously, and the other 23 strains all carried 尾 -lactam, aminoglycoside and quinolone resistance genes simultaneously. SHV and rmt B resistance genes (100%) were also detected in 6 strains of Klebsiella pneumoniae isolated from ward environmental samples, and the plasmids carried more than two kinds of drug resistance genes. Among them, 5 strains of Klebsiella pneumoniae contained 尾 -lactam and aminoglycoside resistance genes simultaneously, and only one strain of Klebsiella pneumoniae contained 尾 -lactams, aminoglycosides and quinolones on the plasmids. Conclusion there are plasmids in Klebsiella pneumoniae detected in patients with NICU and in the environment. The same plasmid can carry several 尾 -lactams, such as SHV,TEM,IMP,rmt Bnr, acc (6)-Ib-cr and so on. Aminoglycosides and quinolones were resistant genes, and each strain of Klebsiella pneumoniae plasmid carried SHV and rmt B genes, but the drug resistance genes carried in the plasmids of Klebsiella pneumoniae were not directly proportional to the clinical phenotype of drug resistance.
【学位授予单位】：华北理工大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R446.5
[Abstract]:Objective to investigate the presence of Klebsiella pneumoniae (Klebsiella pneumoniae,KP) in (Neurological intensive care unit,NICU, a neurointensive intensive care unit. To detect the drug resistance genes of Klebsiella pneumoniae in NICU environment and infected patients, and to analyze the relationship between drug resistance genes and phenotypes of Klebsiella pneumoniae. In order to reduce the incidence of Klebsiella pneumoniae infection in patients with NICU, we should guide the use of clinical antibiotics and sterilize the environment of NICU. Methods the patients admitted to hospital NICU from January 2015 to December 2015 were monitored for infection, and the routine environment and infection environment were monitored by etiology. Klebsiella pneumoniae (Klebsiella pneumoniae) was collected from inpatients with infection and in the environment to detect the phenotype of clinical drug resistance and to retain the strain. The collected Klebsiella pneumoniae was cultured in LB broth medium, and then the plasmid DNA, was extracted by plasmid extraction kit. The 尾 -lactam, aminoglycoside and quinolone resistance genes carried on the plasmid of Klebsiella pneumoniae were detected by polymerase chain reaction using plasmid DNA as template. Results from January 2015 to December 2015, Gram-positive bacteria dominated the NICU ward environment, followed by a small number of Gram-negative bacteria and fungi. The pathogens distributed in the environment were approximately the same as those detected in the specimens of patients with NICU infection. One strain of Klebsiella pneumoniae was detected in the normal environment and 5 strains of Klebsiella pneumoniae were detected in the infected environment. A total of 259 patients were admitted to NICU during the study, of which 179 cases were infected. A total of 31 strains of Klebsiella pneumoniae were detected in the pathogenic specimens. 31 strains of Klebsiella pneumoniae isolated from NICU infected patients and 6 strains of Klebsiella pneumoniae isolated from ward environment were inoculated in LB broth culture medium to extract bacterial plasmid DNA. Plasmids were found in all 37 strains of Klebsiella pneumoniae. At the same time, 21 resistant genes of 尾-lactams, aminoglycosides and quinolones were detected on the plasmids of Klebsiella pneumoniae. SHV and rmt B resistance genes were detected in 31 strains of Klebsiella pneumoniae isolated from infected patients (100%). Among them, 8 strains of Klebsiella pneumoniae carried 尾 -lactam and aminoglycoside resistance genes simultaneously, and the other 23 strains all carried 尾 -lactam, aminoglycoside and quinolone resistance genes simultaneously. SHV and rmt B resistance genes (100%) were also detected in 6 strains of Klebsiella pneumoniae isolated from ward environmental samples, and the plasmids carried more than two kinds of drug resistance genes. Among them, 5 strains of Klebsiella pneumoniae contained 尾 -lactam and aminoglycoside resistance genes simultaneously, and only one strain of Klebsiella pneumoniae contained 尾 -lactams, aminoglycosides and quinolones on the plasmids. Conclusion there are plasmids in Klebsiella pneumoniae detected in patients with NICU and in the environment. The same plasmid can carry several 尾 -lactams, such as SHV,TEM,IMP,rmt Bnr, acc (6)-Ib-cr and so on. Aminoglycosides and quinolones were resistant genes, and each strain of Klebsiella pneumoniae plasmid carried SHV and rmt B genes, but the drug resistance genes carried in the plasmids of Klebsiella pneumoniae were not directly proportional to the clinical phenotype of drug resistance.
【学位授予单位】：华北理工大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R446.5
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