PAPM EAGER: Microwell array platform for high-throughput screening and discovery of microbial interactions

PAPM EAGER：用于高通量筛选和发现微生物相互作用的微孔阵列平台

• 批准号: 1650187
• 负责人: Ryan Hansen
• 金额: $ 30万
• 依托单位: Kansas State University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-15 至 2019-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1650187&HistoricalAwards=false
• 关键词: PAPM; EAGER; Microwell; array; platform

This project aims to develop a new tool for the discovery of interactions occurring within root-associated bacterial communities. Thousands of different bacteria persistently live on plant roots where bacterial interactions help shape these communities and are a critical factor in determining plant health. Traditional approaches test only a few interactions at a time, leaving many bacterial communities poorly characterized. By simultaneously testing thousands of different bacterial interactions, our approach will greatly accelerate the pace of discovery. Uncovering these interactions will aid efforts to manipulate bacterial communities to improve food production and environmental decontamination efforts. Our method will be highly adaptable, allowing for examination of any microbial community and for use in any microbiology laboratory. The project involves an interdisciplinary team of engineers and microbiologists that will apply recent advances in microfabrication with concepts from microbial ecology and genetics, thereby providing interdisciplinary training for a post-doctoral researcher and a graduate student. In addition, the project will support an interactive public outreach program at the Flint Hills Discovery Center illustrating the impact of land use management practices on microbiome diversity in the Flint Hills ecoregion, as well as how micro- and nanotechnologies can be used to improve our understanding of microbes.Microbe-microbe interactions influence microbial community dynamics, composition, and impact on the host. This project aims to develop a high-throughput screening approach for identifying bacterial species that impact a focal bacterial species. The screening platform will use a microwell array to create thousands of unique pairings between the focal species and different bacterial species within a microbiome. Cell pairs will be trapped within their respective wells using a polymer membrane and then monitored with a fluorescent microscope for effects on the focal species. Wells showing enhancement or suppression of focal species function will be extracted and the antagonizing or promoting species will be sequenced for identification. We will validate this technology platform with two screens of Helianthus annuus microbiomes sampled from sites under different land use management regimes at the Konza Long-Term Ecological Research (LTER) site. The first screen will identify bacteria that antagonize or promote the growth of the generalist plant pathogen Agrobacterium tumefaciens. To target specific mechanisms of microbial interaction, the second screen will identify microbiome members that either positively or negatively influence induction of A. tumefaciens' quorum sensing system. The established platform will be low-cost, simple to operate, and applicable for discovery in any microbiome.

该项目旨在开发一种新的工具，用于发现根相关细菌群落内发生的相互作用。成千上万种不同的细菌持续生活在植物根部，其中细菌相互作用有助于形成这些群落，并且是决定植物健康的关键因素。传统的方法一次只能测试几种相互作用，使得许多细菌群落的特征很差。通过同时测试数千种不同的细菌相互作用，我们的方法将大大加快发现的步伐。揭示这些相互作用将有助于操纵细菌群落，以改善食品生产和环境净化工作。我们的方法将具有高度的适应性，允许检查任何微生物群落并在任何微生物学实验室中使用。该项目涉及一个由工程师和微生物学家组成的跨学科团队，他们将利用微生物生态学和遗传学的概念应用微制造的最新进展，从而为博士后研究人员和研究生提供跨学科培训。此外，该项目还将支持弗林特山探索中心的互动式公共宣传计划，说明土地利用管理实践对弗林特山生态区微生物多样性的影响，以及如何使用微生物技术和纳米技术来提高我们对微生物的理解。微生物-微生物相互作用影响微生物群落动态，组成和对宿主的影响。该项目旨在开发一种高通量筛选方法，用于识别影响焦点细菌物种的细菌物种。该筛选平台将使用微孔阵列在微生物组内的焦点物种和不同细菌物种之间创建数千个独特的配对。使用聚合物膜将细胞对捕获在其各自的威尔斯孔内，然后用荧光显微镜监测对焦点物质的影响。将提取显示焦点物质功能增强或抑制的威尔斯孔，并对拮抗或促进物质进行测序以进行鉴定。我们将验证这一技术平台与向日葵微生物组的两个屏幕从不同的土地利用管理制度下的网站在Konza长期生态研究（LTER）网站采样。第一次筛选将鉴定拮抗或促进通用植物病原体根癌农杆菌生长的细菌。为了靶向微生物相互作用的特定机制，第二次筛选将鉴定对A的诱导产生积极或消极影响的微生物组成员。根瘤菌的群体感应系统。建立的平台将是低成本的，操作简单，并适用于发现任何微生物组。

项目成果

On Demand Release and Retrieval of Bacteria from Microwell Arrays Using Photodegradable Hydrogel Membranes

使用可光降解水凝胶膜从微孔阵列中按需释放和回收细菌

• DOI: 10.1021/acsabm.8b00592
• 发表时间: 2018
• 期刊: ACS Applied Bio Materials
• 影响因子: 4.7
• 作者: van der Vlies, André J.;Barua, Niloy;Nieves-Otero, Priscila A.;Platt, Thomas G.;Hansen, Ryan R.
• 通讯作者: Hansen, Ryan R.

Photo-addressable microwell devices for rapid functional screening and isolation of pathogen inhibitors from bacterial strain libraries

• DOI: 10.1063/5.0188270
• 发表时间: 2024-01-01
• 期刊: BIOMICROFLUIDICS
• 影响因子: 3.2
• 作者: Barua，Niloy;Herken，Ashlee M.;Hansen，Ryan R.
• 通讯作者: Hansen，Ryan R.