Systematic approach to explore the microbial interactome

探索微生物相互作用组的系统方法

• 批准号: 9375816
• 负责人: Renate Lux
• 金额: $ 24.65万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-01 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9375816
• 关键词: Address; Adherence; Animal Model; Architecture; Bacteria; Behavior; Binding; Bioinformatics; Biological; Biological Assay; Communities; Complex; Computer software; Corynebacterium; Development; Disease; Disease Outcome; Ensure; Equipment and supply inventories; Fluorescent in Situ Hybridization; Fusobacterium nucleatum; Genome; Goals; Health; Image; Imagery; In Vitro; Individual; Investigation; Knowledge; Label; Laboratories; Membrane; Microbial Biofilms; Modification; Nature; Nucleotides; Oral; Oral cavity; Organism; Pathogenicity; Porphyromonas; Procedures; Property; Public Health; Publishing; Research; Resolution; Role; Sampling; Signal Transduction; Social Behavior; Streptococcus; Structure; System; Testing; Time; Tissues; Translating; Variant; base; combinatorial; comparative genomics; design; disorder prevention; driving force; experience; functional genomics; gene function; in vivo; interest; member; metagenomic sequencing; microbial; microbial community; microorganism; microorganism interaction; next generation sequencing; novel; oral biofilm; parallelization; social; spectrograph; subgingival biofilm; superparamagnetic beads; therapeutic target

Abstract Intermicrobial interactions are key driving forces for social behavior within polymicrobial biofilms that ultimately influences health or disease outcome for the host. The nature of these interactions are greatly influenced by the “Who”, “Where”, “What”, “When”, “How” and “With whom” factors. This application aims to develop an approach for comprehensive assessment of the “With Whom” to ultimately unravel the complex network of interacting microbial species (microbial interactome). Understanding which bacteria adhere to each other is particularly important for open flow systems such as most of the biofilms colonizing the oral cavity: physical interspecies binding ensures the proximity necessary for efficient intercellular signaling, which often occurs on a micron scale. Thus, microorganisms that coadhere are more likely to influence each other’s behavior and potentially take part in the community manipulation that recent research proposed for individual species. Building on our previous published research, we propose to develop a systematic approach and start building the species-level physical interaction network for oral microbial communities using Fusobacterium nucleatum (Fn), a well-established bridging organism for biofilm architecture as model organism. By establishing this approach for Fn we will start testing our working hypothesis that interspecies adherence within the microbial interactome is a major factor for the establishment of health- and disease-associated oral biofilms by dictating the co-localization of distinct subsets of species and thus their combined (beneficial or pathogenic) functional potential. This will be accomplished via the two aims proposed in this application. In Aim 1, we will develop a versatile pulldown assay that employs superparamagnetic beads as anchoring matrix for the “bait” species of interest (Fn) to allow the capture of interacting partner species from representative health- and disease-associated oral microbial “prey” communities. Interacting organisms will be identified using Illumina (MiSeq) next generation sequencing. In Aim 2, we will apply fluorescent in situ hybridization (FISH) or combinatorial labeling and spectral imaging fluorescent in situ hybridization (CLASI-FISH) followed by quantitative analysis of co-localization with the software daime to investigate the in vivo distribution of the Fn microbial interactomes in healthy and diseased subgingival biofilms. Further sequence bioinformatics analysis of the confirmed in vivo interacting species will evaluate their functional genomic potential. The physical microbial interactome is an important determinant in the social structure and behavior of biofilm communities which can potentially determine the health and disease outcome for the associated host. The proposed research will establish a versatile approach for comprehensive investigation of the microbial interactome that is not limited to the oral cavity but can easily be expanded to any biofilm community.

抽象的 微生物间相互作用是多微生物生物膜内社会行为的关键驱动力，最终 影响宿主的健康或疾病结果。这些相互作用的性质很大程度上受 “谁”、“何地”、“什么”、“何时”、“如何”和“与谁”因素。该应用程序旨在开发一种方法 全面评估“与谁”，最终揭开复杂的互动网络 微生物物种（微生物相互作用组）。了解哪些细菌相互粘附尤其重要 对于开放流动系统（例如口腔中的大多数生物膜）很重要：物理种间 结合确保了有效细胞间信号传导所需的邻近性，这通常发生在微米尺度上。 因此，粘附的微生物更有可能影响彼此的行为并可能参与 最近的研究建议对个体物种进行群落操纵。 基于我们之前发表的研究，我们建议开发一种系统方法并开始构建 使用具核梭杆菌的口腔微生物群落的物种级物理相互作用网络 (Fn)，一种成熟的桥接生物，用于生物膜结构作为模型生物。通过建立这个 对于 Fn 的方法，我们将开始测试我们的工作假设，即在 微生物相互作用组是建立与健康和疾病相关的口腔疾病的主要因素 生物膜通过规定不同物种子集的共定位以及它们的组合 （有益或致病）功能潜力。 这将通过本申请中提出的两个目标来实现。在目标 1 中，我们将开发一种多功能的 使用超顺磁珠作为感兴趣的“诱饵”物种 (Fn) 的锚定基质的 Pulldown 测定 允许从代表性的健康和疾病相关口腔中捕获相互作用的伙伴物种 微生物“猎物”群落。将使用下一代 Illumina (MiSeq) 来识别相互作用的生物体 测序。在目标 2 中，我们将应用荧光原位杂交 (FISH) 或组合标记和光谱 成像荧光原位杂交 (CLASI-FISH)，然后进行共定位定量分析 daime 软件用于研究健康和患病人群中 Fn 微生物相互作用组的体内分布 龈下生物膜。对已确认的体内相互作用物种的进一步序列生物信息学分析将 评估它们的功能基因组潜力。 物理微生物相互作用组是生物膜社会结构和行为的重要决定因素 可以潜在地决定相关宿主的健康和疾病结果的社区。这 拟议的研究将建立一种全面研究微生物的通用方法 相互作用组不仅限于口腔，而且可以轻松扩展到任何生物膜群落。