【摘要】：氟喹諾酮是一類(lèi)人工合成藥物,抗菌譜廣,是臨床使用最廣泛的一類(lèi)抗菌藥物,其耐藥也尤為突出。自20世紀(jì)90年代以來(lái),我國(guó)呈現(xiàn)出大腸埃希菌高頻耐藥(64.5%-71.8%)現(xiàn)象,有關(guān)耐藥調(diào)查以及機(jī)制研究大多集中在中心城市三級(jí)醫(yī)院,縣級(jí)醫(yī)院及社區(qū)感染細(xì)菌耐藥研究極少,開(kāi)展縣級(jí)醫(yī)療機(jī)構(gòu)社區(qū)感染患者細(xì)菌對(duì)喹諾酮類(lèi)的耐藥研究,有助于指導(dǎo)這類(lèi)醫(yī)療機(jī)構(gòu)感染治療和抗菌藥物合理使用。 本研究第一部分收集來(lái)自全國(guó)七個(gè)地區(qū)30家縣級(jí)醫(yī)院社區(qū)感染非重復(fù)大腸埃希菌590株,肺炎克雷伯菌569株。細(xì)菌體外藥敏試驗(yàn)檢測(cè)其對(duì)17種抗菌藥物的敏感性；PCR方法檢測(cè)質(zhì)粒介導(dǎo)的喹諾酮耐藥(PMQR)基因：qnrA, qnrB, qnrC, qnrD, qnrS, aac(6')-Ib-cr, qepA和oqxAB及喹諾酮耐藥決定區(qū)(QRDR)DNA旋轉(zhuǎn)酶gyrA, gyrB亞基和拓?fù)洚悩?gòu)酶IV parC, parE亞基突變,采用DNASTAR軟件分析測(cè)序結(jié)果,突變比較參照E. coli k-12和Klebsiella pneumoniae MGH78578;對(duì)非QRDR介導(dǎo)耐藥菌株進(jìn)行外排泵抑制試驗(yàn)。 試驗(yàn)結(jié)果顯示590株大腸埃希菌對(duì)環(huán)丙沙星的耐藥率為51.2%,PMQR基因檢出率為37.3%,qnr類(lèi)的流行率為3.7%,aac(6')Ib-cr19.7%, qepA14.1%, oqxAB3.9%。24株同時(shí)攜帶2種以上PMQR基因。環(huán)丙沙星耐藥組100%存在QRDR突變,93.7%是3個(gè)以上位點(diǎn)突變；敏感組66.7%存在QRDR突變。569株肺炎克雷伯菌對(duì)環(huán)丙沙星耐藥率為8.3%,PMQR檢出率為70.0%,qnr類(lèi)流行率為36.6%,aac(6')Ib-cr9.1%, qepA43.8%。環(huán)丙沙星耐藥組73.9%存在QRDR突變,19.6%是3個(gè)以上位點(diǎn)突變。敏感組4%存在QRDR突變。12株(26.1%)肺炎克雷伯菌為非QRDR介導(dǎo)耐藥,環(huán)丙沙星最低抑菌濃度(MIC)可被外排泵抑制劑N-甲基吡咯烷酮(NMP)降為敏感水平。 分子流行病學(xué)研究發(fā)現(xiàn),縣級(jí)醫(yī)院社區(qū)感染大腸埃希菌和肺炎克雷伯菌氟喹諾酮耐藥率相差懸殊(51.2%versus8.3%),原因除肺炎克雷伯菌氟喹諾酮耐藥率相比大腸埃希菌長(zhǎng)期處于較低水平外,還與肺炎克雷伯菌24.5%(大腸埃希菌9.9%)來(lái)自0-14歲兒童有關(guān),該組環(huán)丙沙星的耐藥率僅為4.2%。大腸埃希菌在耐藥組(100%)甚至敏感組(66.7%)都有很高的QRDR突變率,對(duì)QRDR突變的積累明顯高于肺炎克雷伯菌(耐藥組73.9%,敏感組4%)。 本研究第二部分對(duì)302株環(huán)丙沙星耐藥大腸埃希菌和46株環(huán)丙沙星耐藥肺炎克雷伯菌遺傳特征及多PMQR菌株耐藥質(zhì)粒傳播特征進(jìn)行研究。大腸埃希菌MLST分型選用UCC數(shù)據(jù)庫(kù)(http://mlst.ucc.ie/mlst/dbs/Ecoli),肺炎克雷伯菌MLST分型選用pasteur數(shù)據(jù)庫(kù)(http://bigsdb.web.pasteur.fr/klebsiella/klebsiella.html)。測(cè)序結(jié)果在數(shù)據(jù)庫(kù)中在線(xiàn)比對(duì),獲得等位基因型和序列型(ST)。氟喹諾酮耐藥菌株遺傳進(jìn)化分析采用eBURST V3和商業(yè)化Bionumerics軟件。多PMQR耐藥質(zhì)粒復(fù)制子分型采用基于PCR的分型方法PBRT+ColE+IncR,可對(duì)腸桿菌科20種常見(jiàn)復(fù)制子進(jìn)行分型。并對(duì)這些多PMQR耐藥質(zhì)粒進(jìn)行接合傳播試驗(yàn)。 試驗(yàn)結(jié)果顯示302株環(huán)丙沙星耐藥大腸埃希菌共有60種ST型。常見(jiàn)流行型為ST131(14.6%), ST1193(7.9%), ST405(6.6%), ST69(6.3%), ST648(6.3%)。華東、中南、華北、東北、西北、華南和西南地區(qū)分別有15、26、28、11、17、22和13種。46株環(huán)丙沙星耐藥肺炎克雷伯菌共有23種ST型�？寺�(fù)合體CC11(ST11, ST340, ST258)占15.2%,其次為ST1(13.0%), ST15(10.9%), ST147(10.9%)和ST875(8.7%)。華東、中南、華北、東北、西北、華南和西南地區(qū)分別有3、2、5、4、8、5和7種。大腸埃希菌多PMQR耐藥質(zhì)粒復(fù)制子83.3%為IncF,大多與其他復(fù)制子共存,45.8%可接合轉(zhuǎn)移耐藥基因。肺炎克雷伯菌多PMQR耐藥質(zhì)粒復(fù)制子70%為ColE-like,75%可接合轉(zhuǎn)移耐藥基因。 本研究通過(guò)第二部分耐藥氟喹諾酮遺傳特征分析和多PMQR耐藥質(zhì)粒復(fù)制子分型和傳播特征研究發(fā)現(xiàn)氟喹諾酮耐藥大腸埃希菌序列型多樣,在各地區(qū)呈散在分布,沒(méi)有暴發(fā)流行克隆型。肺炎克雷伯菌雖然僅46株對(duì)環(huán)丙沙星耐藥,但序列型也很豐富,達(dá)23種。也呈散在分布,無(wú)克隆傳播趨勢(shì)。說(shuō)明大腸埃希菌氟喹諾酮耐藥菌株呈多點(diǎn)起源,散在分布。 本研究通過(guò)對(duì)中國(guó)七個(gè)地區(qū)30家縣級(jí)醫(yī)院社區(qū)感染大腸埃希菌和肺炎克雷伯菌氟喹諾酮耐藥分子流行及遺傳特征研究發(fā)現(xiàn)：大腸埃希菌對(duì)氟喹諾酮高頻耐藥,大腸埃希菌和肺炎克雷伯菌對(duì)QRDR突變積累存在差異可能是兩者耐藥率懸殊的一個(gè)重要影響因素。氟喹諾酮耐藥菌株呈多點(diǎn)起源,散在分布。高氟喹諾酮耐藥水平是長(zhǎng)期耐藥基因突變積累的結(jié)果,非優(yōu)勢(shì)克隆傳播引起。
[Abstract]:Flufenone is a kind of synthetic medicine, and its antibacterial spectrum is wide, it is the most widely used anti-bacterial drug in clinical use, and its drug resistance is especially prominent. Since the 1990 's, the high-frequency drug resistance (64.5%-71.8%) of E. coli is present in our country. The study on the drug-resistance of the bacteria of the community-infected patients in the county-level medical institution to the noonone class can help to guide the treatment of such medical institutions and the rational use of the anti-bacterial drugs. The first part of the study collected 590 strains of non-repeated E. coli from 30 county-level hospital communities in seven regions of the country, and K. pneumoniae 569 The susceptibility of the strain to the 17 antibacterial drugs was tested by the in vitro drug sensitivity test of the strain. The PCR method was used to detect the plasmid-mediated gene of the drug resistance (PQR): qnrA, qnrB, qnrC, qnrD, qnrS, aac (6 ')-Ib-cr, qeA and oqxAB, and the drug-resistance-determining region (QRDR), gyrA, gyrB subunit and topoisomerase IV par. C, pare subunit mutation, DNA STAR software was used to analyze the sequencing result, and the mutation was compared with E. coli k-12 and Klebsiella pneumonae MGH78578; and the non-QRDR-mediated drug-resistant strain was subjected to efflux pump inhibition test. The test results showed that the resistance rate of the 590 strains of E. coli to ciprofloxacin was 51.2%, the detection rate of PQR gene was 37.3%, the prevalence rate of qnr was 3.7%, aac (6 ') Ib-cr19.7%, qpA1.4%, oqxAB3.9%.24 strains carried 2 or more PMB simultaneously. The resistance of ciprofloxacin to ciprofloxacin was 8.3%, the detection rate of PQR was 70.0%, the prevalence of qnr was 36.6%, aac (6 ') Ib-cr9.1%, qpA43. The QRDR mutation was found in 73.9% of the ciprofloxacin-resistant group and 19.6% in the ciprofloxacin-resistant group. Point mutations.4% of the sensitive groups had a QRDR mutation.12 (26.1%) of Klebsiella pneumoniae were non-QRDR-mediated resistance, and the minimum inhibitory concentration (MIC) of ciprofloxacin was reduced to sensitivity by the efflux pump inhibitor N-methyl-tridecanone (NMP). The study of molecular epidemiology found that the drug-resistant rates of E. coli and Klebsiella pneumoniae in the community of the county-level hospital were significantly different (51.2% and 83.3%), because of the long-term relationship between E. coli and E. coli than that of Klebsiella pneumoniae. In addition to the lower level, it was also associated with 24.5% of klebsiella pneumoniae (9.9% of E. coli) from 0 to 14 years of age, and the rate of resistance to ciprofloxacin was only The mutation rate of QRDR was higher in the drug-resistant group (100%) and even in the sensitive group (66.7%), and the accumulation of QRDR mutation was significantly higher than that of Klebsiella pneumoniae (73.9% in the drug-resistant group). The genetic characteristics of two strains of ciprofloxacin-resistant E. coli and 46 ciprofloxacin-resistant Klebsiella pneumoniae and the transmission of drug-resistant plasmids of multiple PQR strains in 302 ciprofloxacin-resistant E. coli and 46 ciprofloxacin-resistant strains of ciprofloxacin in the second part of the study The characteristics of the seeding were studied. The UCC database (http://mlst.ucc.ie/mlst/dbs/Ecoli) was used for MLST typing of E. coli. The model of MLST of Klebsiella pneumoniae was selected as the pateur database (

http://bigsdb.web.pastor. fr/klebsiella/ klebsilla.html) ). The sequencing results are online specific in the database to obtain the allelic type and Sequence-type (ST). The analysis of the genetic evolution of the strain-resistant strain of the fluoronone-resistant strain uses eBURST V3 and commercial Bionum. The multi-PQR-resistant plasmid replicon type is based on the PCR-based typing method (PBRT + ColE + IncerR), which can be used for 20 species of Enterobacteriaceae. The replicon was genotyped and these multiple PQR-resistant plasmids were fed The results of the test show that 302 ciprofloxacin-resistant E. coli A total of 60 ST-types were found in the bacteria. The common prevalence was ST131 (14.6%), ST1193 (7.9%), ST405 (6.6%), ST69 (6.3%), ST 648 (6.3%). There are 15,26,28,11,17,22 and 13 species in East China, South China, North China, Northeast, Northwest, South China and Southwest.46 strains of ciprofloxacin-resistant Klebsiella pneumoniae There were 23 types of ST-type. The clone complex CC11 (ST11, ST340, ST258) was 15.2%, followed by ST1 (13.0%), ST15 (10.9%), ST147 (10.9%), and ST. 875 (8.7%). East China, Central South, North China, Northeast, Northwest, South China and Southwest China are 3,2,5, respectively. 4,8,5 and 7. The MPMQR-resistant plasmid replicon of E. coli 83.3% was IncF, most of which co-existed with other replicon, 45.8%. The transfer-resistant gene can be engaged. The multi-PQR-resistant plasmid replicon of Klebsiella pneumoniae is ColE-like,75% In this study, the genetic characteristics of the second part of the drug-resistant fluobanone and the type and the characteristics of the multi-PQR-resistant plasmid replicon were studied. The results of the study found that the sequence type of the drug-resistant E. coli was diverse, and it was scattered in each area. in that case of Klebsiella pneumoniae, only 46 of the strains were resistant to ciprofloxacin, but the order Nematic is also very rich, up to 23 species. No clonal propagation trend in the distribution and no clonal propagation trend. The prevalence and genetic characteristics of the drug-resistant E. coli and Klebsiella pneumoniae in the community of 30 county-level hospitals in seven regions of China have been found in this study. The difference between the high-frequency resistance of E. coli to the high-frequency, E. coli and K. pneumoniae in the QRDR mutation may be both. One of the most important factors of the disparity in drug rate. The drug-resistant level of high-flurolenin is a long-term drug-resistant gene mutation accumulation.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2015
【分類(lèi)號(hào)】：R446.5

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2 姚芬;肺炎克雷伯菌和假單胞菌耐藥機(jī)制研究[D];汕頭大學(xué);2007年

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6 王京;引起肝膿腫的肺炎克雷伯菌毒力基因分析與分子流行特征研究[D];中國(guó)人民解放軍醫(yī)學(xué)院;2013年

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2 唐小紅;2010-2013年我院肺炎克雷伯菌感染的臨床及耐藥分析[D];重慶醫(yī)科大學(xué);2014年

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