鸡源性大肠埃希菌耐β内酰胺类药物机制的研究

发布时间：2018-01-22 16:14

本文关键词： 大肠杆菌 β-内酰胺类抗生素耐药性　出处：《山东大学》2017年硕士论文　论文类型：学位论文

【摘要】：近年来,抗生素的普遍使用使人们在治疗一些感染性疾病时面对的抗生素失效问题日趋严峻,抗生素耐药性对人类健康构成的威胁已经引起了普遍关注。在全部的抗生素中,包含头孢菌素类和碳青霉烯类抗生素的β-内酰胺类抗生素,因为其最早被发现并且运用最为广泛而成为研究的核心。而且大肠埃希菌是能够引起胃肠道疾病和尿道感染的重要的食物病原菌之一,通常我们会用大量的抗生素来治疗大肠杆菌引起的各种感染,这就使得大肠杆菌的耐药性问题给人类健康提出的挑战愈加严峻。我们对鸡源性大肠埃希菌对β-内酰胺类抗生素的耐药性及其耐药性传播机制展开了研究。通过研究待检大肠杆菌对14种抗生素的耐药性统计分析发现,158株非重复鸡源性大肠埃希菌基本上都为多重耐药菌,大部分的大肠杆菌菌株都对10～13种抗生素呈现耐药状态。同时80%的菌株对3种以上的β-内酰胺类抗生素表现出耐药,只有4株菌对β-内酰胺类抗生素表现敏感状态,说明从肉鸡养殖厂中分离出的耐药大肠杆菌对β-内酰胺类抗生素的耐药性尤其显著。整合子是在耐药基因的转移中是尤为重要的一类可移动元件,我们通过检测发现,在158株大肠杆菌中,1型整合子和2型整合子的存在率分别为85.44%、18.35%,3型整合子未被检测到。同时,我们发觉检测到了 7种不同的基因盒,其中1型整合子可变区的基因盒为dfrA17-aadA5,dfrA12-orfF-aadA2,dfrA1-aadA1,catB3-arr3,aacA4-cmLA 1-variant dfrA17-aadA5/aacA4-cmLA1-variant,2型整合子对应基因盒仅发现一种,为dfrA1-sat2-aadA1。耐药决定子的检测结果显示,blaCTX-M和blaTEM耐药基因存在最为普遍,存在率分别为86.7%、81%。但是blaCTX-M与大肠杆菌对β-内酰胺类抗生素的耐药性呈现显著相关性。我们又对存在率最高的blaCTX-M进行了分型,其中有77株菌属于blaaCTX-M-1家族,60株菌属于blaCTX-M-9家族。在blaaCTX-M-1家族中70株为blaCTX-M-55 亚型,7 株为 blaCTX-M-123 亚型;在 blaCTX-M-9 家族中,35 株为 blaCTX-M-65亚型,18株为blaCTX-M-14亚型,7株为blaCTX-M-27亚型。AmpC类耐药基因blaCMY在5株菌中被发现并且都为blaCMY-2亚型。虽然不同的blaCTX-M基因的氨基酸序列具有很高的相似度,但是它们对不同的β-内酰胺类抗生素耐药程度显著不同。blaCTX-M-55主要对AMC、AMP、CAZ、CTX和CPE五种β-内酰胺类抗生素耐药,blaCTX-M-14、blaCTX-M-65、blalCTX-M-123 主要对 AMC、AMP、CTX、CPE 四种β-内酰胺类抗生素耐药,blaCTX-M-27则主要对AMC抗生素耐药,对AMP、CTX的耐药程度介于耐药与敏感之间。进而我们通过接合转移实验发现,可移动元件整合子虽然在耐药大肠杆菌中普遍存在,但是其并没有伴随β-内酰胺类耐药基因的转移而转移到受体菌株。158株大肠杆菌中β-内酰胺类耐药基因接合转移成功率达到53.16%,β内酰胺类耐药基因接合转移成功率显著高于喹诺酮类耐药基因和1型、2型整合子。最后,我们发现检测到的5株大肠杆菌中的blaCMY-2基因与blaCTX-M基因在对碳青霉烯类抗生素亚胺培南的耐药性方面存在协同效应。这些结果表明,肉制品中耐β-内酰胺类抗生素的大肠杆菌会对人类健康构成严重的威胁,整合子在大肠杆菌耐β-内酰胺类抗生素的机制中有重要作用,为解决对碳青霉烯类抗生素耐药性问题提供了新的思路。本研究为耐药性机制研究提供了一定的理论基础,说明解决食品安全和细菌耐药性问题的紧迫性。
[Abstract]:In recent years, the widespread use of antibiotics that people face in the treatment of some infectious diseases of the antibiotic failure problem of antibiotic resistance is becoming more and more serious threat to human health has aroused widespread concern. In all antibiotics, including cephalosporins and carbapenems of beta lactam antibiotics, because the most discovered early and the most widely used and become the core of the study. And Escherichia coli is one of the most important food can cause gastrointestinal diseases and pathogens of urinary tract infection, usually we will all use a lot of antibiotics to treat infections caused by Escherichia coli, which makes the problem of drug resistance of Escherichia coli to human health challenges more and more serious. We launched the study of Chicken Escherichia coli to beta lactam antibiotic resistance and resistance mechanism of transmission. The study tested Escherichia coli resistance to 14 antibiotics of statistical analysis found that 158 strains of non repetitive Chicken Escherichia coli basically are multi drug resistant strains of Escherichia coli, most of the 10 to 13 kinds of antibiotics were resistant state. While 80% strains of 3 species of the beta lactam antibiotics showed drug resistance, only 4 strains were sensitive to beta lactam antibiotics, that drug resistance Escherichia coli isolated from broiler breeding plants of beta lactam antibiotics especially significant. Integron is the transfer of resistance genes is very important for a class of transposable elements. We detected in 158 strains of Escherichia coli, 1 integrons and 2 integrons exist rates were 85.44%, 18.35%, 3 integrons were not detected. At the same time, we found that 7 genes were detected in 1 different boxes. Integron gene cassettes of variable region of dfrA17-aadA5, dfrA12-orfF-aadA2, dfrA1-aadA1, catB3-arr3, aacA4-cmLA 1-variant dfrA17-aadA5/aacA4-cmLA1-variant, 2 integron gene cassettes found only a corresponding decision, the test results show dfrA1-sat2-aadA1. resistance, blaCTX-M and blaTEM resistance genes are the most common, there were 86.7%, but the drug resistance of blaCTX-M 81%. with Escherichia coli to beta lactam antibiotics showed significant correlation. We have to have the highest rates of blaCTX-M were classified, including 77 strains belonging to blaaCTX-M-1 family, 60 strains belonging to the blaCTX-M-9 family. The blaaCTX-M-1 family in 70 were blaCTX-M-55 subtype, 7 strains of blaCTX-M-123 subtype in blaCTX-M-9; in the family, 35 were blaCTX-M-65 subtype, 18 were blaCTX-M-14 subtype, 7 strains of blaCTX-M-27 subtype.AmpC class resistance gene blaCMY in 5 Strains were found and they are all blaCMY-2 subtypes. Although the amino acid sequence of different blaCTX-M genes with high similarity, but they have different beta lactam antibiotic resistance was significantly different.BlaCTX-M-55 of AMC, AMP, CAZ, CTX and CPE five kinds of - lactam antibiotics, blaCTX-M-14 blaCTX-M-65, blalCTX-M-123, AMP, CTX of AMC, CPE, four kinds of beta lactam antibiotics, blaCTX-M-27 is mainly about AMC of antibiotic resistance, AMP, CTX resistance between resistant and sensitive. Then we through the joint transfer experiment found that the movable element integron although in antibiotic resistant Escherichia coli widespread, but the transfer was not accompanied by a beta lactam resistance gene and transferred to the receptor strain.158 in Escherichia coli strain of beta lactam resistance gene conjugative transfer success rate reached 53.16%, with beta Amine resistance gene conjugative transfer success rate was significantly higher than the quinolone resistance gene and type 1, type 2 integron. Finally, we find that there is synergistic effect of blaCMY-2 gene and blaCTX-M gene of 5 strains of Escherichia coli in the detected in resistance to carbapenems imipenem. These results suggest that beta - resistant meat in the products of lactam antibiotics of Escherichia coli will pose a serious threat to human health, the integron in Escherichia coli resistance to beta lactam antibiotics mechanism plays an important role, to solve the carbapenem resistance problem provides a new way of thinking. This study provides a theoretical basis for the mechanism of drug resistance research shows that food safety and urgency to solve the problem of bacterial resistance.

【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R378

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