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

项目摘要 微生物耐药性(AR)的传播是由广泛的 抗生素的临床和农业应用。自然环境是抗生素的巨大港湾 阻性油层，被称为“阻力器”。编码AR的基因存在于移动遗传元件上 并通过水平基因转移(HGT)在微生物之间转移；这种转移可以 因接触抗生素而触发。沿海环境增加了对抗生素的接触 来自人为输入的废物，如废物和雨水径流，这些废物影响了 AR基因从非致病微生物到人类病原体。暴露在沿海水域的人类在 风暴过后或排污口附近以及进食时，耐药感染的风险增加 具有抗药性微生物群的海鲜。阐明AR基因转移的来源和方式 因此，在环境水库中应对这些径流是非常重要的。在增加的同时 在AR中对径流的反应已经建立，将AR基因的转移映射到特定的 社区成员尚未在沿海微生物群中建立。在这项建议中，我们寻求 探索沿海微生物种群中耐药组分布的变化，特别是对 抗生素和风暴径流，通过元基因组方法的组合。1)我们将聘请一名 微基因组染色质构象捕捉技术(MetaHiC)追踪移动基因的运动 微生物群落成员之间的元素和AR基因对处理的响应 四环素，一种在径流中常见的抗生素，已知可诱发HGT。MetaHiC可以绘制移动地图 元素到特定的微生物宿主，提供了抗生素污染如何影响的详细查看 复杂微生物群落中耐药小体的分布。2)我们将结合MetaHiC数据和Long- 阅读元基因组测序以定义整合结合元件(ICES，以下之一)的比率 携带AR基因的主要可移动遗传元件)被切除或整合到微生物基因组中 对四环素治疗的反应。3)在暴雨期间对沿海微生物群使用MetaHiC，我们 将建立在初步数据的基础上(这些数据揭示了分类和抗性组分的变化 对风暴径流的响应)，以开发包含AR基因的沿海微生物图谱 暴雨。我们预计，a)四环素将导致沿海地区冰层的切除和转移。 微生物组以非AR基因特有的方式，b)增加已知病原体的丰度， 抗药性将在暴雨径流的24-72小时内出现，c)增加 暴雨过后，会观察到具有抗药性的共生细菌，形成一个抵抗剂储存库。 用于临床相关的人类病原体作为AR的遗传源。这项研究将进行 并将阐明抗生素耐药性在沿海微生物群中的传播。

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.