Determining mechanisms underlying bacteriophage dynamics during abiotic perturbation.

确定非生物扰动期间噬菌体动力学的潜在机制。

• 批准号: RGPIN-2019-04591
• 负责人: Tropini, Carolina
• 金额: $ 2.7万
• 依托单位: University of British Columbia
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2020
• 资助国家: 加拿大
• 起止时间: 2020-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=715023
• 关键词: Determining; mechanisms; underlying; bacteriophage; dynamics

The long-term goal of my research program is to determine the mechanisms by which microbial communities remain resilient during disturbances (perturbations) in their physical (abiotic) environment. Abiotic environmental factors such as temperature, pH (acidity or alkalinity), and osmolality (the concentration of solute particles in a solution) strongly impact bacterial growth, yet we do not understand how they affect microbial communities. One of the ways that bacterial community dynamics are affected during abiotic perturbations, is through changes in the infection ability and reproduction of viruses that infect bacteria, known as bacteriophages or phages. Although phages are the most abundant organisms on Earth, we know very little about the complexities of the relationship between them and their bacterial hosts, particularly in the context of abiotic perturbations. Understanding the molecular mechanisms underlying this interaction is essential to understanding microbial communities associated with various hosts and environments. This research proposal focuses on unraveling the bacterial as well as phage-dependent mechanisms by which phage infection is resilient to environmental perturbations, specifically osmolality. We have previously shown that shifts in environmental parameters such as external osmolality lead to irreversible changes in both bacterial and phage populations. Our preliminary experimental and computational data indicate that phages that interact with bacterial osmolality sensing proteins may take advantage of a disrupted environment to increase infection. We will now determine the molecular links between bacterial and phage responses to abiotic perturbation: 1) We will investigate the bacterial genes involved in the response to osmotic stress and phage infection in E. coli by applying single-cell imaging and microfluidics approaches in combination with bulk assays. We will test the resilience of the system to osmolality perturbations in a mouse model to determine the in vivo relevance of these genes and their behavior in the context of a complex bacterial community. The combination of experimental and computational analyses will allow us to converge on general mechanisms that in the future we will test in other bacteria-phage pairs and perturbations. 2) We will identify phage genes involved in resilience to environmental stress by combining isolation approaches with in vitro assays and computational analyses. We will create a library of phages against hundreds of bacterial species to identify genetic features predictive of resilience to environmental perturbations. Finally, we will evolve phages to further identify genes involved in resilience to abiotic insults. The completion of this research will not only allow for assignment of novel features in currently largely under-characterized phage genomes, but it will also provide crucial missing knowledge to enable the prediction of microbial community dynamics.

我的研究计划的长期目标是确定微生物群落在其物理（非生物）环境中的干扰（扰动）期间保持弹性的机制。非生物环境因素，如温度，pH值（酸度或碱度）和渗透压（溶液中溶质颗粒的浓度）强烈影响细菌生长，但我们不知道它们如何影响微生物群落。在非生物扰动期间，细菌群落动态受到影响的方式之一是通过感染细菌的病毒（称为噬菌体或噬菌体）的感染能力和繁殖的变化。虽然细菌是地球上最丰富的生物，但我们对它们与细菌宿主之间关系的复杂性知之甚少，特别是在非生物扰动的背景下。了解这种相互作用的分子机制对于了解与各种宿主和环境相关的微生物群落至关重要。这项研究计划的重点是解开细菌以及噬菌体依赖的机制，噬菌体感染是弹性的环境扰动，特别是渗透压。我们以前已经表明，在环境参数的变化，如外部渗透压导致细菌和噬菌体种群的不可逆变化。 我们的初步实验和计算数据表明，与细菌渗透压传感蛋白相互作用的细菌可能会利用破坏的环境来增加感染。我们现在将确定细菌和噬菌体对非生物扰动的反应之间的分子联系：1）我们将研究在E.大肠杆菌通过应用单细胞成像和微流体方法与批量测定相结合。我们将在小鼠模型中测试系统对渗透压扰动的弹性，以确定这些基因及其在复杂细菌群落背景下的行为的体内相关性。实验和计算分析的结合将使我们能够收敛于一般的机制，在未来，我们将在其他细菌噬菌体对和扰动测试。2)我们将通过结合分离方法和体外测定及计算分析来鉴定参与环境压力恢复的噬菌体基因。我们将创建一个针对数百种细菌的细菌文库，以确定预测对环境扰动的适应性的遗传特征。最后，我们将进一步发展基因组学，以进一步确定参与对非生物伤害的恢复力的基因。 这项研究的完成不仅可以在目前基本上特征不足的噬菌体基因组中分配新特征，而且还将提供关键的缺失知识，以预测微生物群落动态。