Exploiting Pf phage superinfection to lower Pseudomonas aeruginosa virulence via evolutionary tradeoffs

利用 Pf 噬菌体重复感染通过进化权衡降低铜绿假单胞菌毒力

• 批准号: 10748681
• 负责人: Nanami Kubota
• 金额: $ 4.77万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10748681
• 关键词: Acute; Affect; Antibiotics; Bacteria; Bacterial Chromosomes; Bacterial Infections; Bacteriophages; Binding; Biological Models; Capsid; Capsid Proteins; Cells; Characteristics; Charge; Chromosomes; Chronic; Clinical; Collection; Cytolysis; Data; Drug Tolerance; Environment; Evolution; Family; Filament; Funding; Genes; Genome; Goals; Growth; Human; Infection; Inoviridae; Inovirus; Learning; Length; Longitudinal Studies; Measurement; Measures; Metadata; Microbial Biofilms; Modeling; Mutation; Parasites; Pathogenesis; Patients; Pharmaceutical Preparations; Phenotype; Population; Prevalence; Production; Prophages; Pseudomonas aeruginosa; Pseudomonas aeruginosa infection; Pyocyanine; Recording of previous events; Reporting; Resources; Testing; Time; Viral Genome; Viral Proteins; Virulence; Virulence Factors; acute infection; antibiotic tolerance; bacterial fitness; clinically relevant; clinically significant; cost; cystic fibrosis patients; fitness; improved; insight; mutant; novel; novel strategies; pathogen; pathogenic bacteria; pressure; prevent; pyoverdin; reproductive; superinfection; whole genome

PROJECT SUMMARY Many clinically relevant pathogens have bacteriophage (phage) genomes integrated in their chromosome (prophages), which can have large effects on the bacteria’s phenotype and fitness. Since phage fitness is tied to bacterial host fitness, phages can become fitter by decreasing their burden on their host. However, when bacteria are infected by multiple phages (i.e., superinfection), competition for host resources may select phages that are more competitive against other phages, even at the increased burden to their host. The objective of this proposal is to understand and learn to exploit the evolutionary tradeoff between phage intracellular competitiveness and bacterial fitness during superinfection. Understanding this tradeoff will give better insight into how prophages influence their bacterial host’s phenotype and fitness, and potentially pave way for a novel approach in phage therapy that utilizes such tradeoffs to make bacterial infections easier to treat. We propose to use Pseudomonas aeruginosa (Pa) and its prophage, Pf phage, as a model system for better understanding the evolutionary tradeoff between phage competitiveness and bacterial fitness. More than half of Pa carry Pf prophages. Furthermore, cystic fibrosis (CF) patients who are chronically infected with Pa were more likely to have Pa that have Pf prophages than acutely infected patients. This hints that Pf phages are an integral part of Pa evolutionary history and pathogenesis. The first part of this project aims to understand whether Pa virulence factor production is correlated with the number of Pf prophages that Pa carries in its chromosome. From a previously funded study, we have a collection of >100 Pa clinical isolates from 33 CF patients that carry zero, one, or two Pf prophages. Using this collection and lab Pa strains, we will test for correlations between Pf copy number and the production of virulence factors like pyocyanin and pyoverdine. In the second aim, we will superinfect Pa lab strain PA14 with Pf phage that has a mutation in the prophage repressor gene. This Pf phage mutant replicates quickly and at high populations within the host cell, creating an environment that favors selection for defective interfering (DI) phages that lack capsid genes. These DI phages are cheaters that exploit full-length phages for public resources, like capsid, to selfishly propagate. Pf capsid is tied to clinically relevant phenotypes of Pa, such as biofilm robustness and antibiotic tolerance. Thus, the loss of capsid genes is not only a way Pf phages become more competitive against other Pf phages but can affect host fitness as well. We plan to evolve PA14 infected with this selfish Pf phage under biofilm and antibiotic selection to test whether capsid genes become lost over time, undermining biofilm stability and antibiotic tolerance. Completion of this project will provide valuable information on the influence prophages have on their bacterial host’s phenotype and fitness. Exploiting the evolutionary tradeoff between phage competitiveness and host fitness may potentially pave way for a novel approach in phage therapy, analogous to a gene drive, that exploits such tradeoffs to make bacterial infections easier to treat.

项目摘要 许多临床相关病原体的染色体中整合有噬菌体基因组 （原噬菌体），其可以对细菌的表型和适合度具有很大影响。既然噬菌体适应度 对于细菌宿主适合度，细菌可以通过减少它们对宿主的负担而变得更适合。然而当 细菌被多种病原体感染（即，重叠感染），对宿主资源的竞争可能选择重叠感染， 即使对他们的主人增加了负担，他们也比其他人更具竞争力。的目的 我们的建议是理解并学会利用噬菌体细胞内 竞争力和细菌适应性。理解这种权衡将使我们更好地了解 研究前噬菌体如何影响其细菌宿主的表型和适应性，并可能为一种新的 噬菌体疗法中的一种方法，利用这种权衡使细菌感染更容易治疗。 我们建议使用铜绿假单胞菌（Pa）及其前噬菌体Pf噬菌体作为模型系统， 理解噬菌体竞争力和细菌适应性之间的进化权衡。一半以上 Pa携带Pf原噬菌体。此外，囊性纤维化（CF）患者谁是慢性感染Pa更多 可能比急性感染患者更容易感染具有Pf原噬菌体的Pa。这暗示Pf是一个积分 Pa的部分进化史和发病机制。该项目的第一部分旨在了解Pa是否 毒力因子的产生与Pa在其染色体中携带的Pf原噬菌体的数量相关。 从先前资助的研究中，我们收集了来自33名携带PA的CF患者的>100个PA临床分离株， 零个、一个或两个Pf原噬菌体。使用该集合和实验室Pa菌株，我们将测试Pf之间的相关性。 拷贝数和毒力因子如绿脓菌素和绿脓菌荧光素的产生。第二个目标，我们将 用在原噬菌体阻遏物基因中具有突变的Pf噬菌体重复感染Pa实验室菌株PA14。这种Pf噬菌体 突变体在宿主细胞内快速复制并大量繁殖，创造了一个有利于 选择缺乏衣壳基因的缺陷型干扰（DI）引物。这些侦探是骗子， 为了公共资源，像衣壳一样，自私地繁殖。Pf衣壳与临床相关的 表型，如生物膜稳健性和抗生素耐受性。因此，衣壳基因的丢失不仅是 一种方式，Pf的球员变得更有竞争力，对其他Pf的球员，但可以影响主机的健身以及。我们计划 在生物膜和抗生素选择下进化感染这种自私的Pf噬菌体的PA14，以测试衣壳是否 基因随着时间的推移而丢失，破坏了生物膜的稳定性和抗生素耐受性。 该项目的完成将提供有关原噬菌体对细菌的影响的有价值的信息。 寄主的表型和适合度。利用噬菌体竞争性和宿主之间的进化权衡 适应性可能为噬菌体治疗的新方法铺平道路，类似于基因驱动， 这样的权衡使细菌感染更容易治疗。