Elucidation of colonization and virulence through a novel comparative approach

描述(申请人提供)：副溶血性弧菌是世界范围内引起胃肠炎的主要病原体。虽然已经确定了一些毒力机制，但这些机制并不能完全解释这种病原体致病的能力。在这项研究中，我们将通过一种新的比较方法来阐明导致定殖和毒力的机制。包括副溶血性弧菌在内的人类致病弧菌的一个共同特点是，它们的许多毒力因子是水平获得的，但获得性毒力特性必须与其他定植特性一起协调调节。彻底瞄准所有独特的序列将是低效的，而且对连接毒力和定殖力的调控网络提供的洞察力很小。我们认为，将该病原体与一种不能致病的相关细菌的定植进行比较，将表明哪些定殖机制在该属中是古老和保守的。更重要的是，这一新颖的比较将揭示与副溶血性弧菌致病生活方式有关的独特调控转录本。我们推测，副溶血弧菌中不受GacA调控或不存在GacA调控的基因与副溶血弧菌的毒力和定植有关。我们的假设和预测是基于这样一个事实：1)包括霍乱毒素在内的致病弧菌中水平获得的毒力因子受整个属中保守的调节因子控制，包括共生菌，费氏弧菌；2)GacA调节共生因子和寄主在裂叶弧菌中的定植，以及在许多病原体包括霍乱弧菌中的毒力和定植；3)在所有弧菌中未保守的关键共生因子(Lux)受裂谷弧菌中的GacA调节，表明生活方式特有的特征映射到这一调节因子。这项研究的具体目的是： *通过GacA的突变产生全球定植缺陷的副溶血性弧菌菌株。 *利用费氏弧菌和副溶血性弧菌中GacA调节子的比较DNA阵列分析来确定可能的定植和毒力特征。 *确定GacA调控的靶向基因在细胞毒性中的作用。 与公共卫生相关：过去十年出现了一种大流行的副溶血性弧菌菌株，它在以前被认为没有危险的地区引起了疫情，预示着气候变化对人类健康的威胁越来越大。评估感染风险和制定预防方案的一个主要障碍是缺乏对副溶血性弧菌致病性的了解。我们的发现将为了解这种重要的新兴病原体的毒力机制提供宝贵的见解，揭示宿主的脆弱性，并为预防和治疗感染提供新的机会。

DESCRIPTION (provided by applicant): The emergent human pathogen Vibrio parahaemolyticus is a major cause of gastroenteritis worldwide. Although some virulence mechanisms have been identified, these do not fully explain the ability of this pathogen to cause disease. In this study we will elucidate mechanisms that contribute to colonization and virulence by a novel comparative approach. A common feature of human pathogenic Vibrios, including V. parahaemolyticus, is that many of their virulence factors are horizontally acquired, and yet acquired virulence traits must be coordinately regulated along with other colonization traits. Exhaustively targeting all unique sequences would be inefficient, and would provide little insight into regulatory networks that connect virulence and colonization. We propose that a comparison of colonization between this pathogen and a related bacterium that is incapable of causing disease, such as the well characterized light organ symbiont Vibrio fischeri, will indicate which mechanisms of colonization are ancient and conserved in the genera. More importantly, the novel comparison will reveal uniquely regulated transcripts that contribute specifically to the pathogenic lifestyle of V. parahaemolyticus. We hypothesize that GacA-regulated genes in V. parahaemolyticus that are not GacA-regulated or absent in V. fischeri contribute to the virulence and colonization by V. parahaemolyticus. We base our hypothesis and predictions on the fact that 1) horizontally acquired virulence factors in pathogenic Vibrios including cholera toxin are controlled by regulators conserved throughout the genus including the symbiotic bacterium, Vibrio fischeri, 2) GacA regulates symbiosis factors and host-colonization in V. fischeri and virulence and colonization in many pathogens including Vibrio cholerae, 3) key symbiotic factors that are not conserved in all Vibrios (lux) are regulated by GacA in V. fischeri indicating lifestyle-specific traits map to this regulon. The specific aims of the study are to: * Generate a global colonization-defective strain of V. parahaemolyticus through a mutation in GacA. * Use comparative DNA array analysis of GacA regulons in V. fischeri and V. parahaemolyticus to identify putative colonization and virulence traits. * Determine the role of targeted GacA-regulated genes in cytotoxicity. PUBLIC HEALTH RELEVANCE: The last decade marked the emergence of a pandemic strain of V. parahaemolyticus, which caused outbreaks in regions that were previously not considered to be at risk, foreshadowing the increasing threat to human health due to climate changes. A major obstacle to assessing risk of infection and developing preventative protocols is a lack of understanding of the pathogenicity of V. parahaemolyticus. Our findings will provide invaluable insight into the virulence mechanisms of this important emergent pathogen revealing host vulnerabilities and providing new opportunities to prevent and treat infections.