Investigating bacterial virulence using an alternative host-pathogen model

毒力被定义为细菌病原体通过产生病原体在宿主中入侵和繁殖所必需的因子来伤害宿主生物体的能力。这些因素必须被识别和功能特性，以了解细菌病原体的行为。我的研究计划的长期目标是确定致病菌影响鱼类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。我使用一种方法，结合细菌基因组学与使用的替代主机模型Dictyosteelium discoideum，土壤变形虫，是一种天然的捕食者的细菌，以实现这一目标。D. discoideum是研究嗜冷病原菌毒力的一个特别有用的工具，包括那些影响鱼类的细菌，如鲑鱼。在接下来的五年里，我的研究计划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业产生重大负面影响。使用变形虫宿主模型，我们已经表明，这种细菌是敏感的压力条件下，这会导致基因组的修改，影响主要的毒力因子，包括3型分泌系统，导致毒力的损失。A.杀鲑纲是由于其基因组中存在丰富的移动的遗传元件。使用下一代测序和基因分型的多个A。杀鲑毒素菌株，我们有新的未知毒力因子存在的初步证据。基于A.杀鲑毒素受移动的遗传因子的影响，我们认为杀鲑毒素中的其他毒力因子也受移动的遗传因子的影响，可能是防治杀鲑毒素的靶点。沙门氏菌感染对A.杀鲑鱼毒力因子和评价它们对从天然产物分离的化合物的直接作用或通过由这些化合物刺激的移动的遗传元件的作用的敏感性是一种有前途的方法。因此，本课题的近期研究工作主要集中在以下几个方面：（1）鉴定新的毒力因子;利用D. discoideum作为宿主，并通过比较基因组学。(2)使用各种方法对这些新的毒力因子进行功能表征，包括在鱼类中的挑战。这部分研究还将涉及开发新的遗传工具来有效地操纵A。杀鲑基因(3)鉴定与水产养殖相容的化合物，通过刺激移动的遗传元件直接抑制毒力因子或破坏其基因。研究了这些化合物对A.用D. discoideum作为宿主。最有希望的化合物将在鱼类身上进行测试。我的研究计划将有助于更好地了解A.杀鲑鱼的。基于我们对A.鉴于杀鲑素和本提案中提出的目标，我们相信，我们将能够通过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，这些药物不仅可用于兽医领域，还可用于人类健康。

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.