Employing Metagenomic and Chromatin Capture Techniques to Map the Evolution of Antibiotic Resistance in Coastal Microbiomes

项目摘要 微生物抗生素耐药性（AR）的传播是一个全球性的医疗保健问题， 抗生素的临床和农业应用。自然环境是抗生素的巨大港湾 阻力储层，称为“阻力区”。编码AR的基因存在于移动的遗传元件上 并且通过水平基因转移（HGT）在微生物之间转移;这种转移可以是 是由于接触抗生素而引发的沿海环境增加了抗生素的暴露 来自人为输入的废物，如废物和雨水径流，这影响了 AR基因从非病原微生物到人类病原体。暴露于沿海沃茨的人类 在暴风雨后或废水排放口附近以及进食时，耐药性感染的风险增加 具有抗微生物菌群的海鲜。阐明AR基因转移的来源和模式 因此，环境水库对这些径流的响应非常重要。而与 已经建立了AR对径流的反应，将AR基因转移到特定的 在沿海微生物群落中尚未建立社区成员。在本建议中，我们力求 探索沿海微生物种群中耐药基因组分布的变化，特别是在应对 抗生素和暴雨径流，通过宏基因组方法的组合。1)我们将雇用一名 宏基因组染色质构象捕获（MetaHiC）来追踪移动的遗传物质的运动， 元素和AR基因之间的微生物群落成员的处理响应 四环素，一种在径流中常见的抗生素，已知会引起HGT。MetaHiC可以映射移动的 元素对特定微生物宿主的影响，详细了解抗生素污染如何影响 耐药基因组分布在复杂的微生物群落中。2)我们将联合收割机MetaHiC数据与长- 读取宏基因组测序，以确定整合接合元件（ICE， 携带AR基因的主要移动的遗传元件）被切除或整合到微生物基因组中， 对四环素治疗的反应。3)使用MetaHiC在整个暴雨期间的沿海微生物群落中，我们 将建立在初步数据的基础上（这些数据揭示了 响应风暴径流），以开发含有AR基因的沿海微生物的概况， 暴雨我们预计，a）四环素将诱导切割和转移的ICE在沿海 B）增加已知病原体的丰度， 在暴雨径流后24-72小时内， 在暴雨之后，将观察到具有抗药性的细菌，从而形成一个抗药性水库 临床相关的人类病原体用作AR的遗传来源。这项研究将在 并将阐明抗生素耐药性在沿海微生物群落中的传播。

Project Summary The spread of microbial antibiotic resistance (AR) is a global healthcare problem driven by the extensive clinical and agricultural use of antibiotics. Natural environments serve as massive harbors of antibiotic resistance reservoirs, known as the “resistome”. Genes that encode AR exist on mobile genetic elements and are transferred between microbes through horizontal gene transfer (HGT); this transfer can be triggered in response to antibiotic exposure. Coastal environments have increased exposure to antibiotic waste from anthropogenic inputs such as waste- and stormwater runoff, which influence the spread of AR genes from non-pathogenic microbes to human pathogens. Humans exposed to coastal waters are at increased risk of drug-resistant infections after storms or near wastewater outlets, as well as when eating seafood with antibiotic-resistant microbiomes. Elucidating the sources and patterns of AR gene transfer in environmental reservoirs in response to these runoffs is therefore of great importance. While increases in AR in response to runoffs have been established, mapping the transfer of AR genes to particular community members has yet to be established in coastal microbiomes. In this proposal, we seek to explore the changes in resistome distribution in coastal microbial populations, specifically in response to antibiotics and storm runoff, through a combination of metagenomic approaches. 1) We will employ a metagenomic chromatin conformation capture (MetaHiC) to trace the movement of mobile genetic elements and AR genes between microbial community members in response to treatment with tetracycline, an antibiotic commonly in runoff and known to induce HGT. MetaHiC can map mobile elements to particular microbial hosts, providing a detailed look at how antibiotic contamination affects resistome distribution in complex microbial communities. 2) We will combine MetaHiC data with long- read metagenomic sequencing to define the ratio in which integrative conjugative elements (ICEs, one of the main mobile genetic elements that carry AR genes) are excised or integrated in microbial genomes in response to tetracycline treatment. 3) Using MetaHiC on coastal microbiomes throughout rainstorms, we will build on preliminary data (which revealed taxonomic and resistome composition changes in response to storm runoff) to develop a profile of coastal microbes containing AR genes following rainstorms. We anticipate that a) tetracycline will induce excision and transfer of ICEs in coastal microbiomes in a non-AR gene-specific manner, b) increases in the abundance of known pathogens that are antibiotic-resistant will appear within 24-72 hours of storm runoff, c) increases in the abundance of antibiotic-resistant commensal bacteria will be observed after rainstorms, creating a resistome reservoir for clinically relevant human pathogens to use as a genetic source of AR. This research will be performed by undergraduates and will elucidate the spread of antibiotic resistance in coastal microbiomes.