Identification of genetic factors affecting the survival of Legionella pneumophila in water systems

嗜肺军团菌（Legionella pneumophila，Lp）是引起肺炎的军团菌病（Legionnaires' disease，LD）。LD的发病率相当低，但它正在增加，并且在世界各地的主要城市定期发生大规模爆发，期间可能有数百人被感染。死亡率通常在10%左右。脂蛋白存在于热水分配系统和冷却塔中，它生长在吞噬原生动物体内。通过气溶胶传播给人类。* 我的长期目标是了解影响细菌病原体在水系统中生存的遗传和环境因素，并制定战略，以加强对水传播细菌病原体的控制。目前，氯和热处理用于控制水系统中的Lp。这些治疗有助于暂时减少Lp的数量，从而减少对人类的传播，但不能消除Lp。据推测，脂蛋白具有在这种治疗中生存的反应系统，但对脂蛋白对热休克和氯的反应知之甚少。此外，随着时间的推移，在水系统中的LP种群结构的菌株和变化之间的遗传多样性水平高，建议适应LP的特定条件下存在于水系统和所使用的治疗。这可能是由点突变和从常驻微生物获得基因驱动的。因此，我在未来五年的短期目标是：1）确定Lp对热休克的转录反应，2）确定参与Lp对氯和热休克反应的基因，3）确定Lp在周期性处理的复杂水系统中生长期间的遗传适应。对于第一个目标，我们将使用转录组学的方法来研究我们以前确定的调节器在热休克反应中发挥作用的作用。对于第二个目标，将使用基于转座子的方法来识别在热处理和氯消毒中存活所需的基因。对于第三个目标，将在纯水和冷却塔微生物群中进行进化实验，以产生与祖先菌株相比对热处理和氯具有高度抗性的克隆。将通过全基因组测序鉴定与耐药性增加相关的基因突变和基因获得。* 我的研究计划描述了一种强有力的方法，应该阐明使Lp抵抗控制措施的遗传因素。这将有助于改进用于控制人造水系统内Lp和其他细菌病原体复制的策略，以防止人类感染。此外，这项工作将有助于我们了解Lp和其他水媒病原体的进化，特别是水平基因转移的作用。在这项研究计划中，一名硕士，3名博士和至少5名本科生将接受微生物学，分子遗传学和生物信息学的培训。

Legionella pneumophila (Lp) causes the pneumonia Legionnaires' disease (LD). Incidence of LD is rather low, but it is increasing, and large outbreaks occur periodically in major cities worldwide during which hundreds of people can be infected. The mortality rate is typically around 10%. Lp is found in hot water distribution systems and cooling towers where it grows inside phagocytic protozoans. Transmission to humans occurs via aerosols. ***My long-term goals are to understand the genetic and environmental factors affecting the survival of bacterial pathogens in water systems and to develop strategies to enhance control of water-borne bacterial pathogens. Currently, chlorine and thermal treatments are used to control Lp in water systems. These treatments help reduce momentarily the population of Lp, and hence the transmission to humans, but fail to eliminate Lp. Presumably, Lp possesses response systems to survive such treatments, but not much is know about Lp's response to heat shock and chlorine. In addition, high level of genetic diversity between isolates and changes in Lp population structure in water systems over time suggest adaptation of Lp to specific conditions present in water systems and the treatment used. This is likely driven by point mutations and by acquisition of genes from resident microorganisms. Therefore, my short-term objectives for the next five years are to: 1) determine the transcriptional response of Lp to heat shock, 2) identify genes involved in the response of Lp to chlorine and heat shock, and 3) identify genetic adaptation of Lp during growth in complex water system undergoing periodic treatment.***For the first objective, we will use a transcriptomic approach to study the role of regulators that we have previously identified to play a role in heat shock response. For the second objectives, a transposon-based approach will be used to identify genes required for surviving thermal treatment and chlorine disinfection. For the third objective, evolution experiments will be performed in pure water and in cooling tower microbiota to produce clones highly resistant to thermal treatment and chlorine compare to the ancestor strain. Genetic mutations and gene acquisitions associated with increased resistance will be identified by whole genome sequencing. ***My research plan describes a powerful approach that should shed lights on the genetic factors enabling Lp to resist control measures. This should help refine the strategies used to control the replication of Lp and other bacterial pathogens inside man-made water systems to prevent human infections. In addition, this work will contribute to our understanding of the evolution of Lp and other water-borne pathogens, particularly the role of horizontal gene transfer. During this research program, one MSc, 3 PhD and at least 5 undergraduate students will be trained in microbiology, molecular genetics, and bioinformatics.