本文選題：醫(yī)院感染 + 鮑曼不動(dòng)桿菌�。� 參考：《安徽醫(yī)科大學(xué)》2012年碩士論文

【摘要】：研究目的和內(nèi)容 1.了解廈門部分地區(qū)醫(yī)院感染病原體鮑曼不動(dòng)桿菌(BA)的耐藥情況,掌握耐藥譜隨時(shí)間變化趨勢(shì),為臨床選擇適當(dāng)?shù)目股靥峁┮罁?jù)。 2.探索鮑曼不動(dòng)桿菌的脈沖場(chǎng)凝膠電泳條件,了解醫(yī)院感染病原體以及分離株的基因多態(tài)性,閘明其菌株間的遺傳親緣關(guān)系。 3.結(jié)合流行病學(xué)資料分析鮑曼不動(dòng)桿菌基因圖譜的聚類分析結(jié)果,鑒別優(yōu)勢(shì)克隆菌株的時(shí)間、地區(qū)以及人群的三間分布,判斷爆發(fā)菌株和散發(fā)菌株。 4.分析廣泛耐藥鮑曼不動(dòng)桿菌常見(jiàn)耐藥基因的攜帶情況。 研究方法 1.菌株的收集方法：2009年1月-2011年12月,收集廈門部分地區(qū)三級(jí)醫(yī)院住院的醫(yī)院感染患者分離的鮑曼不動(dòng)桿菌,標(biāo)本來(lái)源于醫(yī)院住院患者的痰、血液、尿液、糞便和膿液等樣本,所有菌株經(jīng)西門子Walkaway96鑒定定藥敏儀(美國(guó),德靈)鑒定,并剔除同一患者相同樣本重復(fù)菌株。 2.藥物敏感性測(cè)定方法：按美國(guó)臨床及實(shí)驗(yàn)室標(biāo)準(zhǔn)協(xié)會(huì)推薦標(biāo)準(zhǔn),采用紙片擴(kuò)散法對(duì)所有收集的菌株進(jìn)行藥物敏感性試驗(yàn)。 3.基因分型方法：建立適合鮑曼不動(dòng)桿菌的脈沖場(chǎng)凝膠電泳分型的實(shí)驗(yàn)室條件,對(duì)所有收集的菌株進(jìn)行PFGE基因分型,采用BionumericSV4.0軟件中的非加權(quán)配對(duì)算數(shù)平均法進(jìn)行聚類分析,分析菌株間的親緣關(guān)系。 4.廣泛耐藥菌株耐藥基因的檢測(cè)：PCR擴(kuò)增法檢測(cè)多重耐藥鮑曼不動(dòng)桿菌分離株中的超廣譜β-內(nèi)酰胺酶、金屬β-內(nèi)酰胺酶等常見(jiàn)耐藥基因。 研究結(jié)果 1.廈門部分地區(qū)2009-2011年醫(yī)院感染BA的藥物敏感性結(jié)果 1.1BA藥敏的總體情況本研究共收集到386株醫(yī)院感染鮑曼不動(dòng)桿菌,對(duì)20種常用抗生素(青霉素類及加酶抑制劑類、頭孢菌素類及加酶抑制劑類、頭霉素類、氨基糖苷類、氟喹諾酮類、磺胺類、多粘菌素類、四環(huán)素類)均表現(xiàn)出不同程度的耐藥。從高到低依次如下：氨曲南(87.05%)、哌拉西林(77.72%)、頭孢吡肟(77.72%)、頭孢噻肟(72.28%)、厄他培南(71.50%)、替卡西林/克拉維酸(69.17%)、復(fù)方新諾明(68.91%)、左氧氟沙星(67.62%)、妥布霉素(67.62%)、環(huán)丙沙星(67.36%)、頭孢他啶(66.32%)、慶大霉素(65.54%)、阿莫西林/克拉維酸(64.51%)、美羅培南(63.73%)、啞胺培南(63.47%)、阿米卡星(57.77%)、頭孢哌酮/舒巴坦(52.85%)、多粘菌素(41.19%),米諾環(huán)素(0.00%)；沒(méi)有發(fā)現(xiàn)泛耐藥及全敏感菌株；分離菌株對(duì)常見(jiàn)p-內(nèi)酰胺類抗生素及加酶抑制劑、氨基糖苷類、喹諾酮類抗生素、磺胺類耐藥較為嚴(yán)重,均超過(guò)50%,有的接近90%；對(duì)碳青霉烯類抗生素也呈現(xiàn)較高的耐藥性,耐藥率均超過(guò)60%；多粘菌素耐藥率低于50%,米諾環(huán)素耐藥率為0.00%。 1.2BA藥敏的時(shí)間變化趨勢(shì) 廈門部分地區(qū)分離的BA耐藥情況較為嚴(yán)重,3年間,廈門部分地區(qū)分離的鮑曼不動(dòng)桿菌對(duì)于多數(shù)常見(jiàn)的抗生素的耐藥性呈現(xiàn)逐年升高的趨勢(shì),尤其以亞胺培南、美羅培南、厄他培南和多粘菌素增長(zhǎng)較為明顯。 1.3多重耐藥及泛耐藥鮑曼不動(dòng)桿菌分離株的檢出情況 386株鮑曼不動(dòng)桿菌中檢出多重耐藥菌株118株,廣泛耐藥菌株97株,檢出率分別為30.57%和25.13%。 2.醫(yī)院感染鮑曼不動(dòng)桿菌基因分型結(jié)果 2.1PFGE應(yīng)用于BA的幾個(gè)技術(shù)關(guān)鍵點(diǎn)：0D值為4.5, Spa I20U酶切3h以上,脈沖角度為120度,電場(chǎng)強(qiáng)度為6V/cm,電泳20h。在此條件下90%以上的鮑曼不動(dòng)桿菌的圖潛條帶數(shù)目、大小、亮度及長(zhǎng)度均非常合適。 2.2在使用相同的試劑、儀器和參數(shù)的前提下,386株菌經(jīng)限制性內(nèi)切酶酶切后進(jìn)行PFGE電泳,每株約出現(xiàn)21-33個(gè)電泳條帶,分子量在30kb-800kb之間。經(jīng)BionumericSV4.0軟件進(jìn)行聚類分析,相似值在45.9%-100%之間,可分為242個(gè)型(相似值為100%視為同一PFGE型別)； 2.3最常見(jiàn)的基因型有40株菌,20,09年12株,2010年15株,2011年13株。 2.497株廣泛耐藥鮑曼不動(dòng)桿菌的基因分型 97株廣泛耐藥的鮑曼不動(dòng)桿菌桿菌中經(jīng)BionumericSV4.0軟件進(jìn)行聚類分析,相似值在67.9%-100%之間,可分為67個(gè)型。 3.廣泛耐藥鮑曼不動(dòng)桿菌分離株的耐藥基因的研究 97株泛耐藥鮑曼不動(dòng)桿菌檢出TEM基因陽(yáng)性23例,檢出率為23.71%；CTX-M-13基因陽(yáng)性檢出17例,檢出率為17.53%。97株泛耐藥鮑曼不動(dòng)桿菌檢出VIM-2陽(yáng)性菌株30例,檢出率為30.93%；IMP-1陽(yáng)性菌株39例,檢出率為40.20%；OXA-58陽(yáng)性菌株10例,檢出率為10.31%；OXA-51陽(yáng)性菌株80例,檢出率為82.47%；OXA-23陽(yáng)性菌株85例,檢出率為87.63%；OXA-24陽(yáng)性11株,檢出率為11.34%。沒(méi)有檢出NDM-1基因。 研究結(jié)論 1、廈門部分地區(qū)2009年到2011年鮑曼不動(dòng)桿菌主要分離自呼吸系統(tǒng)標(biāo)本,科室分布主要是重癥監(jiān)護(hù)病房和呼吸內(nèi)科,分離患者人群集中在45歲以上人群。 2、對(duì)廈門部分地區(qū)2009年到2011年分離的醫(yī)院感染鮑曼不動(dòng)桿菌的耐藥性調(diào)查表明：386株分離菌的對(duì)20種常用抗生素均表現(xiàn)出不同程度的耐藥性,對(duì)常見(jiàn)的p-內(nèi)酰胺類抗生素及加酶抑制劑、氨基糖苷類、喹諾酮類抗生素、磺胺類耐藥較為嚴(yán)重,均超過(guò)50%,有的接近90%；對(duì)碳青霉烯類抗生素也呈現(xiàn)較高的耐藥性,耐藥率均超過(guò)60%；多粘菌素耐藥率低于50%,米諾環(huán)素耐藥率為0.00%。3年間,廈門部分地區(qū)分離的鮑曼不動(dòng)桿菌對(duì)于多數(shù)常見(jiàn)的抗生素的耐藥性呈現(xiàn)逐年升高的趨勢(shì),尤其以亞胺培南、美羅培南、厄他培南和多粘菌素增長(zhǎng)較為明顯。 3、PFGE是一種穩(wěn)定、精確的基因分型方法,適合作為院內(nèi)感染的同源性分析。 4、349株的PFGE進(jìn)行基因分型,結(jié)果顯示廈門部分地區(qū)存在242種基因型的流行,在多家醫(yī)院病房中不存在優(yōu)勢(shì)克隆株；廈門部分地區(qū)2009年到2011年未出現(xiàn)過(guò)大規(guī)模的爆發(fā)流行,ICU病房出現(xiàn)頻率高,交叉感染出現(xiàn)于不同醫(yī)院之間；另外,不同醫(yī)院之間出現(xiàn)基因型完全一致菌株,提示及時(shí)的流行病學(xué)調(diào)查的重要性。 5、廈門部分地區(qū)泛耐藥鮑曼不動(dòng)桿菌攜帶的耐藥基因包括ESBLs耐藥基因組中的TEM和CTX-M-13亞型和碳青霉烯酶中的OXA-23, OXA-24, OXA-51, OXA-58和VIM-2、IMP-1,主要以O(shè)XA-23型和OXA-51型碳青霉烯酶為主。
[Abstract]:Purpose and content of the study

1 . To understand the drug resistance of acinetobacter baumanii ( BA ) infected by nosocomial infection in some parts of Xiamen , and to grasp the trend of drug resistance spectrum over time and provide the basis for clinical selection of appropriate antibiotics .

2 . To explore the genetic polymorphism of the pathogen of nosocomial infection and the genetic polymorphism of isolated strain , and to explore the genetic relationship among the isolates .

3 . Based on the results of clustering analysis , the time , region and three - compartment distribution of the dominant clonal strains were identified , and the outbreak strains and sporadic strains were identified .

4 . To analyze the carrying situation of the common drug resistant gene of broad - resistant acinetobacter baumanii .

Research Methods

1 . Collection method of bacterial strain : From January 2009 to December 2011 , the patient was collected from hospital infected patients at three levels of hospital in Xiamen , and the samples were collected from sputum , blood , urine , feces and pus samples from the hospital in hospital . All the strains were identified by Siemens Walkaway96 ( United States , Germany ) and the same sample repeated strains of the same patient were excluded .

2 . Method for measuring drug sensitivity : According to the recommendations of the American Society for Clinical and Laboratory Standards , all collected strains were tested for drug sensitivity using sheet diffusion method .

3 . genotyping method : establishing a laboratory condition suitable for a pulsed field gel electrophoresis typing of baumanii , performing PFGE genotyping on all collected strains , performing cluster analysis by adopting a non - weighted pair arithmetic mean method in the BionumericSV4.0 software , and analyzing the relationship among strains .

4 . Detection of resistance gene of broad - resistant strains : PCR amplification method is used to detect the common drug - resistant genes such as ultra - wide spectrum 尾 - lactamase , 尾 - lactamase and other common drug resistance genes .

Results of the study

1 . Results of drug sensitivity of BA for nosocomial infection in some parts of Xiamen from 2009 to 2011

1 . A total of 386 nosocomial infections were collected from a total of 386 hospitals in this study . There were 20 common antibiotics ( penicillin and enzyme inhibitors , cephem , aminoglycosides , fluoroquinolone , sulfanime , polymyxin and tetracycline ) . The results showed that there were no significant differences in resistance to antibiotics ( 87.05 % ) , piperacillin ( 77.72 % ) , cefpime ( 66.32 % ) , gentamicin ( 65.54 % ) , amoxicillin / clavianic acid ( 64.51 % ) , meropenem ( 63.73 % ) , dumb - amine penem ( 63.47 % ) , amikaine ( 57.77 % ) , cefperazone / Sulptan ( 52.85 % ) . Polymyxin ( 41.19 % ) , Minoryclines ( 0.00 % ) ;
No drug resistance and all - sensitive strains were found .
The isolates were more resistant to common p - lactam antibiotics and enzyme inhibitors , aminoglycosides , quinolinone antibiotics and sulfanilamide , more than 50 % and nearly 90 % .
The drug resistance of enem antibiotics was higher than that of 60 % .
The resistance rate of polymyxin was lower than 50 % , and the drug resistance rate was 0.00 % .

1.2 Time variation trend of drug sensitivity of 2BA

5 . The drug - resistant genes carried by Pan - drug - resistant acinetobacter baumanii in some parts of Xiamen include TEM and CTX - M - 13 subtype and OXA - 23 , OXA - 24 , OXA - 51 , OXA - 58 and vim - 2 , IMP - 1 , which are mainly OXA - 23 and OXA - 51 .

1.3 Detection of multi - drug resistant and pan - resistant acinetobacter baumanii isolates

Among 386 strains of acinetobacter baumanii , 118 strains of resistant strains and 97 strains of resistant strains were detected , and the detection rates were 30.57 % and 25.13 % , respectively .

2 . Results of the genotyping results of M . baumanii in nosocomial infection

2.1 PFGE is applied to several technical key points of BA : the D value is 4.5 , the Spa I20U enzyme is cut for more than 3h , the pulse angle is 120 degrees , the electric field intensity is 6 V / cm , and the electrophoresis is 20 hours . Under this condition , the number , the size , the brightness and the length of the gram latent bands of the acinetobacter baumanii are very suitable .

2.2 On the premise of using the same reagents , instruments and parameters , 386 strains were subjected to restriction enzyme digestion and PFGE electrophoresis was carried out . The molecular weight was between 30 kb and 800 kb . Cluster analysis was performed by BionumericSV4.0 software . The similarity values were between 45.9 % -100 % , and the similarity values were between 45.9 % -100 % .


2.3 The most common genotypes were 40 strains , 12 strains in 20 and 09 , 15 in 2010 and 13 in 2011 .

Gene typing of 2.497 strains of broad - resistant acinetobacter baumanii

Cluster analysis was performed by BionumericSV4.0 software in 97 broad - resistant strains of Escherichia coli , and the similarity values were between 67.9 % and 100 % , which could be divided into 67 types .

3 . Study on the resistance genes of widely resistant isolates of acinetobacter baumanii

There were 23 cases of TEM gene positive in 97 pan - resistant acinetobacter baumandii , and the detection rate was 23.71 % .
The positive rate of CTX - M - 13 gene was 17.53 % . The positive rate was 30.93 % .
The positive rate of IMP - 1 was 40.20 % .
The positive rate of OXA - 58 was 10.31 % .
The positive rate of OXA - 51 was 82.47 % .
The positive rate of OXA - 23 was 87.63 % .
The positive rate of OXA - 24 was 11.34 % . NDM - 1 gene was not detected .

Conclusions of the study

1 . The main isolated self - respiratory system samples from 2009 to 2011 in Xiamen area were mainly from intensive care unit and respiratory department , and the population of isolated patients was more than 45 years old .

2 . The drug resistance of M . baumanii isolated from 2009 to 2011 in some parts of Xiamen showed that the resistance of 386 isolates showed different levels of resistance to 20 common antibiotics , and more than 50 % of the common p - lactam antibiotics and enzyme inhibitors , aminoglycosides , quinolinone antibiotics and sulfanilamide were more severe , with nearly 90 % of them .
The drug resistance of enem antibiotics was higher than that of 60 % .
The drug resistance rate of polymyxin was lower than 50 % , and the resistance rate of Minoryclines was 0.00 % . In the past three years , the drug resistance of acinetobacter baumanii isolated from some parts of Xiamen was increasing year by year for most common antibiotics , especially in Imipenem , meropenem , UNMEE penem and polymyxin .

3 . PFGE is a stable and accurate genotyping method , which is suitable for the homology analysis of nosocomial infection .

The results showed 242 genotypes were prevalent in some parts of Xiamen , and there were no dominant clones in many hospitals .
In some parts of Xiamen , there were no large - scale outbreaks in 2009 - 2011 , the frequency of ICU wards was high , and the cross - infection occurred between different hospitals ;
In addition , there was a completely identical genotype among different hospitals , which suggested the importance of timely epidemiological investigation .

The resistant condition of isolated BA in some parts of Xiamen was more serious . In the past three years , the drug resistance of acinetobacter baumanii isolated from some parts of Xiamen was increasing year by year for most common antibiotics , especially the growth of Imipenem , meropenem , UNMEE penem and polymyxin .
【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R446.5;R3416

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本文編號(hào)：2079355