Pathogen Synergy Through Cross-Species Induction of Outer Membrane Vesicle Biogenesis

通过跨物种诱导外膜囊泡生物发生的病原体协同作用

• 批准号: 10043365
• 负责人: Jeffrey Schertzer
• 金额: $ 21.44万
• 依托单位: STATE UNIVERSITY OF NY,BINGHAMTON
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-29 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10043365
• 关键词: ANXA5 gene; Antibiotics; Apoptosis; Area; Bacteria; Behavior; Biogenesis; Cells; Coculture Techniques; Communicable Diseases; Communication; Communities; Complex; Data; Disease; Disease model; Enzymes; Epidemiology; Epithelial Cells; Escherichia coli; Foundations; Future; Gammaproteobacteria; Human; Immune response; Immune system; In Vitro; Individual; Induction of Apoptosis; Infection; Label; Lead; Life; Lipids; Lung; Mass Fragmentography; Mediator of activation protein; Membrane; Methods; Modeling; Organism; Pathogenesis; Pathogenicity; Patient-Focused Outcomes; Patients; Phenotype; Play; Population; Problem Solving; Production; Pseudomonas aeruginosa; Reporting; Role; Signal Transduction; Site; Testing; Time; Toxin; Trypan Blue; Vesicle; Virulence; Virulence Factors; Virulent; Work; antibiotic tolerance; bacterial community; cell type; co-infection; cytotoxic; cytotoxicity; experimental study; follow-up; genetic element; human pathogen; in vivo; insight; macrophage; microbiota; novel; pathogen; pathogenic bacteria; small molecule; synergism

Abstract Bacteria have been cooperating and competing for all of evolutionary time. Only recently, however, have we seriously begun to take this into account in the context of infectious disease mechanisms. The expanding ability to study bacteria in complex consortia (i.e. how they normally live) has sparked a renewed appreciation for the complexity of real bacterial populations and an examination of how they interact. Co-infection studies show that pathogen communities are more virulent, and this aligns with epidemiological reports that connect multispecies infections to worse patient outcomes. A common hypothesis is that this pathogen synergy comes about because of competition and communication between multiple species at an infection site. We hypothesize that Outer Membrane Vesicles (OMVs) serve as a major mediator of these interactions because they are known to facilitate both competition and communication between bacteria (in addition to direct virulence against host cells). We proposed a model in P. aeruginosa where OMV biogenesis is driven by the secretion and insertion of a self- produced small molecule into the outer leaflet of the outer membrane. We recently showed that this molecule could induce related species to overproduce OMVs when given at low concentration, and that those recipient species produced their own OMV cross-inducing factors. This raised the possibility that cross-species induction of OMV biogenesis could serve as a mechanism for multispecies communities to synergize for increased virulence. Guided by strong preliminary data, we will pursue two specific aims to characterize how multiple pathogens at an infection site might interact through cross-species induction of OMV biogenesis and what effects this would have on host cells: (1) We will begin by testing for small molecule-induced OMV formation using the known P. aeruginosa OMV-inducing compound PQS against a broad panel of important human pathogens likely to encounter each other at an infection site. Our established methods have already identified several such interactions among the γ-proteobacteria. We will then test whether actually growing species together (in contrast to using monocultures with exogenous compounds as above) will result in increased OMV production for the community. Preliminary results show that this is true for our pilot pairing of P. aeruginosa + E. coli. (2) OMVs from mono- vs. co-culture will be tested for their cytotoxic potential against macrophage and lung epithelial cells as well as their ability to degrade or sequester antibiotics (another disease-related function of OMVs). Preliminary results with P. aeruginosa + E. coli co-culture OMVs show that cytotoxicity and induction of apoptosis are both increased over monoculture OMVs, demonstrating feasibility of the approach and providing support for the hypothesis that cross-species OMV induction may contribute to pathogen synergy. Understanding pathogen interactions at infection sites is critical to understanding why multispecies infections lead to more severe disease. Given the ubiquity of OMV production among Gram-negative organisms, the insights gained from this study promise to broadly impact our understanding of multi-species pathogenesis.

抽象的 细菌一直在整个进化时期合作和竞争。但是，直到最近我们才有 在传染病机制的背景下，认真地开始考虑这一点。扩展的能力 在复杂的联盟中研究细菌（即它们通常的生活方式）引发了人们对 实际细菌种群的复杂性以及对它们相互作用的研究。共感染研究表明 病原体群落更具毒性，这与连接多人的流行病学报告保持一致 感染对患者预后更糟的感染。一个常见的假设是这种病原体协同作用是因为 在感染部位的多个物种之间的竞争和沟通。我们假设外部 膜囊泡（OMV）是这些相互作用的主要中介者，因为已知它们有助于 细菌之间的竞争和通信（除了针对宿主细胞的直接病毒外）。 我们在铜绿假单胞菌中提出了一个模型，其中OMV生物发生是由分泌和插入自我驱动的 产生的小分子进入外膜的外部小叶。我们最近表明该分子 在低浓度下给出时，可能会诱导相关物种过度生产OMV，而那些受体 物种产生了自己的OMV交叉诱导因素。这增加了跨物种诱导的可能性 OMV生物发生可以作为多物种社区协同作用以增加的机制 毒力。在强大的初步数据的指导下，我们将追求两个具体的目标，以表征多个 感染部位的病原体可能通过跨物种诱导OMV生物发生以及什么影响相互作用 这将在宿主细胞上具有：（1）我们将使用使用该分子诱导的小分子诱导的OMV形成开始 已知的铜绿假单胞菌OMV诱导化合物PQs针对一组重要的人类病原体可能 在感染部位相遇。我们既定的方法已经确定了几种 γ-蛋白菌之间的相互作用。然后，我们将测试是否真正种植物种（相反） 使用上述外源化合物的单一培养物将导致OMV产生增加 社区。初步结果表明，对于我们的铜绿假单胞菌 +大肠杆菌的试验配对是正确的。 （2）OMV 从单与培养物中，将测试其针对巨噬细胞和肺上皮细胞的细胞毒性潜力 以及它们降解或隔离抗生素（OMV的另一个功能）的能力。初步的 铜绿假单胞菌 +大肠杆菌共培养OMV的结果表明，细胞毒性和凋亡的诱导都是 对单一文化OMV的增加，证明了该方法的可行性并为 跨物种OMV诱导可能有助于病原体协同作用的假设。 了解感染部位的病原体相互作用对于理解为什么多物种感染至关重要 导致更严重的疾病。考虑到革兰氏阴性生物中OMV生产的普遍性， 从这项研究中获得的见解有望广泛影响我们对多物种发病机理的理解。