Investigating bacterial virulence using an alternative host-pathogen model

使用替代宿主病原体模型研究细菌毒力

• 批准号: RGPIN-2014-04595
• 负责人: Charette, Steve
• 金额: $ 3.86万
• 依托单位: Université Laval
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2014
• 资助国家: 加拿大
• 起止时间: 2014-01-01 至 2015-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=566621
• 关键词: Investigating; bacterial; virulence; using; alternative

Virulence is defined as the ability of a bacterial pathogen to injure a host organism by producing factors that are essential for the pathogen to invade and multiply in the host. These factors must be identified and functionally characterized in order to understand the behaviour of bacterial pathogens. The long-term objective of my research program is to identify the molecular basis of the virulence factors produced by pathogenic bacteria affecting fish as well as ways to inhibit or modulate them to prevent or treat bacterial infections in fish farms. I use an approach that combines bacterial genomics with the use of the alternative host model Dictyostelium discoideum, a soil amoeba that is a natural predator of bacteria, to achieve this goal. D. discoideum is a particularly useful tool for studying the virulence of psychrophilic pathogenic bacteria, including those that affect fish such as salmonids. Over the next five years, my research program will continue to focus on the virulence of Aeromonas salmonicida. This bacterium is a fish pathogen that has a major negative impact on aquaculture in Canada. Using the amoeba host model, we have shown that this bacterium is sensitive to stressful conditions, which cause genomic modifications that affect major virulence factors, including the type three secretion system, leading to a loss of virulence. The genomic plasticity of A. salmonicida results from the presence of abundant mobile genetic elements in its genome. Using next generation sequencing and genotyping of multiple A. salmonicida strains, we have preliminary evidence of the presence of new uncharacterized virulence factors. Based on the hypothesis that important virulence factors of A. salmonicida are affected by the activity of mobile genetic elements, we propose that other virulence factors in this bacterium are also affected by the activity of mobile genetic elements and may be targets for the prevention or treatment of A. salmonicida infections. The identification and characterization of A. salmonicida virulence factors and the evaluation of their sensitivity to the direct effect of compounds isolated from natural products or through the action of mobile genetic elements stimulated by these compounds is a promising approach. My research program will thus focus on the following short-term objectives: (1) Identify new virulence factors of A. salmonicida by genetic screening using D. discoideum as a host and by comparative genomics. (2) Functionally characterize these new virulence factors using various approaches, including challenges in fish. This part of the study will also involve the development of new genetic tools to efficiently manipulate A. salmonicida genes. (3) Identify aquaculture-compatible compounds that directly inhibit virulence factors or disrupt their genes by stimulating mobile genetic elements. The effect of these compounds on A. salmonicida virulence will be screened using D. discoideum as a host. The most promising compounds will then be tested on fish. My research program will provide a better understanding of the virulence factors of A. salmonicida. Based on our on-going effort to understand the pathogenic basis of A. salmonicida and the objectives set out in this proposal, we are confident that we will be able to modulate and inhibit the virulence of this bacterium by taking advantage of its genomic plasticity. This research is crucial for the development of novel anti-bacterial drugs that target virulence factors and that can be used as alternatives to antibiotics not only in the veterinary field but also for human health.

毒力被定义为细菌病原体通过产生病原体侵入宿主并在宿主中繁殖所必需的因子来伤害宿主生物体的能力。必须识别这些因素并对其进行功能表征，以便了解细菌病原体的行为。我的研究计划的长期目标是确定影响鱼类的病原菌产生的毒力因子的分子基础，以及抑制或调节它们以预防或治疗养鱼场细菌感染的方法。我使用一种将细菌基因组学与替代宿主模型盘基网柄菌（一种土壤变形虫，细菌的天然捕食者）相结合的方法来实现这一目标。 D. discoideum 是研究嗜冷病原菌毒力的特别有用的工具，包括那些影响鱼类（如鲑鱼）的病原菌。在接下来的五年中，我的研究项目将继续关注杀鲑气单胞菌的毒力。这种细菌是一种鱼类病原体，对加拿大的水产养殖产生重大负面影响。使用阿米巴宿主模型，我们发现这种细菌对压力条件敏感，压力条件会导致基因组修饰，影响主要毒力因子，包括三型分泌系统，导致毒力丧失。杀鲑鱼的基因组可塑性源于其基因组中存在丰富的可移动遗传元件。通过对多种杀鲑鱼菌株进行下一代测序和基因分型，我们获得了存在新的未表征毒力因子的初步证据。基于杀鲑鱼重要毒力因子受移动遗传元件活性影响的假设，我们提出该细菌的其他毒力因子也受移动遗传元件活性影响，并可能成为预防或治疗的靶点杀鲑鱼感染。杀鲑鱼毒力因子的鉴定和表征以及评估它们对从天然产物中分离的化合物的直接作用或通过这些化合物刺激的移动遗传元件的作用的敏感性是一种有前途的方法。因此，我的研究计划将集中于以下短期目标：（1）通过使用盘状 D. discoideum 作为宿主的遗传筛选和比较基因组学来鉴定杀鲑鱼的新毒力因子。 (2) 使用各种方法（包括鱼类的挑战）对这些新毒力因子进行功能表征。这部分研究还将涉及开发新的遗传工具来有效操纵杀鲑鱼基因。 (3) 鉴定可直接抑制毒力因子或通过刺激可移动遗传元件破坏其基因的与水产养殖相容的化合物。这些化合物对杀鲑鱼毒力的影响将使用盘状鲑鱼作为宿主进行筛选。然后，最有希望的化合物将在鱼身上进行测试。我的研究计划将有助于更好地了解杀鲑鱼的毒力因子。基于我们对杀鲑鱼致​​病基础的持续努力以及本提案中提出的目标，我们相信我们将能够利用其基因组可塑性来调节和抑制这种细菌的毒力。这项研究对于开发针对毒力因子的新型抗菌药物至关重要，这些药物不仅可以在兽医领域而且可以用于人类健康作为抗生素的替代品。