【摘要】：目的研究銅綠假單胞菌的臨床分布和抗菌藥物的耐藥性,從分子水平探討其產(chǎn)生多重耐藥的可能機(jī)制和感染的同源性,從而為有效地預(yù)防和控制多重耐藥銅綠假單胞菌感染提供科學(xué)的參考依據(jù)。方法1.收集并保存寧夏醫(yī)科大學(xué)附屬醫(yī)院自2011年9月至2012年9月期間臨床分離的非重復(fù)PA菌株285株,采用法國(guó)梅里埃公司全自動(dòng)微生物分析儀(VITEK 2 COMPACT)鑒定細(xì)菌;采用K-B法檢測(cè)PA臨床分離株對(duì)14種常用抗生素的藥物敏感試驗(yàn),應(yīng)用WHONET5.6軟件對(duì)上述藥物敏感結(jié)果進(jìn)行統(tǒng)計(jì)分析,篩選出多重耐藥銅綠假單胞菌。2.將篩選出的124株MDRP菌株通過(guò)外排泵抑制劑利血平對(duì)羧芐西林、紅霉素、亞胺培南及慶大霉素的瓊脂稀釋法抑制試驗(yàn)分別檢測(cè)銅綠假單胞菌的Mex AB-Opr M、Mex CD-Opr J、Mex EF-Opr N、Mex XY-Opr M四種外排泵的表型,同時(shí)檢測(cè)加入利血平后四種藥物對(duì)銅綠假單胞菌MIC值的變化;應(yīng)用聚合酶鏈反應(yīng)(PCR)技術(shù)檢測(cè)外排泵基因。3.將篩選出的124株MDRP菌株,通過(guò)PCR檢測(cè)整合酶基因,將IntⅠ檢測(cè)陽(yáng)性菌株繼續(xù)采用PCR方法擴(kuò)增可變區(qū);采用PCR-RFLP分析并確定菌株可變區(qū)類型。4.應(yīng)用脈沖場(chǎng)凝膠電泳(PFGE)技術(shù)對(duì)多重耐藥銅綠假單胞菌進(jìn)行同源性分析。結(jié)果1.2011年9月至2012年9月我院共分離出285株銅綠假單胞菌,標(biāo)本的來(lái)源主要是痰液(65.5%),科室的分布主要集中在ICU(34.7%),耐藥結(jié)果顯示:臨床分離的銅綠假單胞菌對(duì)多粘菌素B敏感,對(duì)亞胺培南、頭孢他啶等耐藥率低,在臨床治療中可以選擇聯(lián)合用藥;篩選出的124株MDRP亦主要來(lái)自痰液(62.9%),主要分布在ICU(22.6%)。2.外排泵表型陽(yáng)性結(jié)果為:Mex AB-Opr M74株、Mex XY-Opr M28株、Mex EF-Opr N12株和Mex CD-Opr J10株;124株MDRP菌株中74株Mex AB-Opr M外排泵表型陽(yáng)性。3.檢測(cè)124株MDRP其中Ⅰ類整合子陽(yáng)性菌株有87株,未檢測(cè)出Ⅱ類整合子;87株IntⅠ基因陽(yáng)性的菌株中有53株有效的擴(kuò)增出Ⅰ類整合子可變區(qū),通過(guò)測(cè)序發(fā)現(xiàn)6種形式的耐藥基因盒。4.對(duì)124株MDRP進(jìn)行脈沖場(chǎng)凝膠電泳(PFGE)分析得到8個(gè)基因型,其中主要的有5個(gè)基因型。結(jié)論1.在臨床常用抗生素中,銅綠假單胞菌臨床分離株對(duì)多粘菌素B最敏感,對(duì)復(fù)方新諾明的耐藥率最高。標(biāo)本來(lái)源主要是痰液,PA主要分布在ICU。2.四種外排泵表型中最常見(jiàn)的是Mex AB-Opr M,推測(cè)其可能是銅綠假單胞菌多重耐藥的重要機(jī)制。3.多重耐藥銅綠假單胞菌整合子的分布類型以Ⅰ類整合子為主,整合子中的氨基糖甙類修飾酶基因和β-內(nèi)酰胺類基因的存在與多藥耐藥銅綠假單胞菌的多重耐藥密切相關(guān)。4.在基因水平上通過(guò)PFGE對(duì)MDRP進(jìn)行分析,得出MDRP感染來(lái)源、傳播途徑及分布規(guī)律,對(duì)感染流行的監(jiān)測(cè)提供可靠依據(jù)。
[Abstract]:Objective to study the clinical distribution and antimicrobial resistance of Pseudomonas aeruginosa and to explore the possible mechanism of multidrug resistance and the homology of infection at molecular level. It provides a scientific reference for the prevention and control of multidrug resistant Pseudomonas aeruginosa infection. Method 1. From September 2011 to September 2012, 285 strains of non-repetitive PA strains were collected and preserved in the affiliated Hospital of Ningxia Medical University. The bacteria were identified by VITEK 2 COMPACT. K-B method was used to detect the drug sensitivity of clinical isolates of PA to 14 common antibiotics. The results of drug sensitivity were statistically analyzed by WHONET5.6 software, and the multidrug resistant Pseudomonas aeruginosa was screened out. Carbenicillin and erythromycin were separated from 124 strains of MDRP by reserpine, an efflux pump inhibitor. Imipenem and gentamicin Agar dilution inhibition tests were used to detect the phenotypes of four efflux pumps of Pseudomonas aeruginosa (Mex AB-Opr MMex CD-Opr JnMex EF-Opr NMex XY-Opr M), and the changes of MIC value of four drugs on Pseudomonas aeruginosa after adding reserpine. Polymerase chain reaction (PCR) technique was used to detect the efflux pump gene. 124 strains of MDRP were screened, the integrase gene was detected by PCR, the Int 鈪，

本文編號(hào)：2225629