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Unravelling mechanisms by which Zinc influences diversity and multi-resistance of porcine intestinal Escherichia coli populations

揭示锌影响猪肠道大肠杆菌群多样性和多重耐药性的机制

基本信息

批准号：
292118477
负责人：
Professor Dr. Lothar H. Wieler
金额：
--
依托单位：
HPI Digital Health Center
依托单位国家：
德国
项目类别：
Research Grants
财政年份：
2016
资助国家：
德国
起止时间：
2015-12-31 至 2019-12-31
项目状态：
已结题

来源：
https://gepris.dfg.de/gepris/projekt/292118477?language=en
关键词：
Unravelling mechanisms Zinc influences diversity

项目摘要

Despite the ban of antimicrobial growth promoters the prevalence of MDR bacteria, especially MRSA and ESBL-producing Escherichia (E.) coli, raised tremendously in livestock. Instead alternative feed additives - like the cationic trace element zinc - are increasingly used. Genes conferring resistance against cationic trace elements are often co-localized with genes conferring antimicrobial resistance. Therefore we raised the hypothesis that zinc feeding is one cause of the increase of MDR bacteria. As E. coli is both a major commensal and pathogenic bacterial species of the intestinal microbiota, in two feeding trials of piglets with zinc-oxid (ZnO) of particle sizes between 100-200 µm within the SFB852, we molecularly analyzed nearly 1,500 intestinal E. coli to the clonal level. Indeed, zinc feeding had a strong impact on E. coli, as strains from zinc fed piglets showed (i) a greater genetic diversity, and even more importantly (ii) an increased rate of MDR E. coli. Unexpectedly, antimicrobial resistance of E. coli was not significantly associated with zinc tolerance. The analyzed intestinal habitats did also not contain an increased E. coli density, so that increased physical contact does not explain a possible higher conjugation rate. In conclusion, co-selection of zinc tolerance and drug resistance is not the sole explanation for our findings.To understand the mechanisms underlying our observations of a higher diversity and an increase in MDR associated with zinc feeding, the presented project aims at:(i) Identifying genetic differences between E. coli strains isolated from the feeding and the control group of the mentioned feeding trials by whole genome sequence analysis. (ii) Characterizing and functionally proving the mechanisms responsible for the higher rate of MDR E. coli by initial whole genome sequence analysis and in vitro assays.As basis for these experiments we utilize E. coli strains that were isolated from the mentioned feeding trials and which have been extensively characterized by macrorestriction analysis (defining clones), by Multiplex-PCR (defining pathotypes) and by MLST) defining phylotypes). We thereby circumvent biases of previous studies which either performed functional investigations on E. coli K-12 lab strains only or judged on the intestinal E. coli population by random colony picking. Taking into consideration the growing knowledge on the genetic diversity of E. coli genomes, ranging in size from 4.5 to 5.5 Mbp and in gene contents from ~4,100 to ~5,800 genes, our aand in gene contents from ~4,100 to ~5,800 genes, our approach will tremendously increase the knowledge on the influence of zinc on natural E. coli populations in piglets and beyond.This proposal is a straightforward continuation of previous work performed in the institute. Most importantly, unravelling the mechanisms by which zinc increases the occurrence of MDR E. coli will have fundamental impact on feeding procedures in livestock.

尽管禁止使用抗菌生长促进剂，但耐多药细菌的流行，特别是MRSA和产ESBL的大肠杆菌（E.）大肠杆菌，在家畜中大量繁殖。替代饲料添加剂-如阳离子微量元素锌-越来越多地使用。赋予阳离子微量元素抗性的基因通常与赋予抗菌药物抗性的基因共定位。因此，我们提出了一个假设，即锌喂养是MDR细菌增加的原因之一。例如E.大肠杆菌是肠道菌群的主要致病菌，在两次仔猪饲喂SFB 852中粒径为100-200 μm的氧化锌（ZnO）试验中，我们对近1,500株肠道大肠杆菌进行了分子分析。coli的克隆水平。事实上，锌饲料对E。大肠杆菌，因为来自锌饲喂仔猪的菌株显示（i）更大的遗传多样性，并且更重要的是（ii）MDR E.杆菌出乎意料的是，E. coli与锌耐受性无显著相关性。分析的肠道栖息地也不包含增加的E。大肠杆菌密度，因此增加的物理接触并不能解释可能的较高接合率。总之，锌耐受和耐药的共同选择不是我们发现的唯一解释。为了理解我们观察到的与锌喂养相关的更高多样性和MDR增加的机制，本项目旨在：（i）鉴定E.通过全基因组序列分析，对上述饲养试验的饲养组和对照组分离的大肠杆菌菌株进行了比较。（二）表征并功能性证明MDR E较高发生率的机制。大肠杆菌的全基因组序列分析和体外实验。大肠杆菌菌株，其分离自所提及的饲养试验，并且已经通过宏观限制性分析（确定克隆）、通过多重PCR（确定致病型）和通过MLST（确定致病型）进行了广泛表征。因此，我们避免了以往研究的偏见，无论是进行功能调查E。coliK-12实验室菌株或对肠E.大肠杆菌种群随机菌落挑取。考虑到对E.大肠杆菌基因组大小为4.5 ~ 5.5Mbp，基因含量为4，100 ~ 5，800个基因。这个建议是对研究所以前工作的直接延续。最重要的是，阐明锌增加MDR E发生的机制。大肠杆菌对牲畜的饲养程序有根本性的影响。