Circulating Bacteriophages for the Diagnosis of Sepsis

用于诊断脓毒症的循环噬菌体

• 批准号: 10673035
• 负责人: Paul L Bollky
• 金额: $ 23.2万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10673035
• 关键词: Academic Medical Centers; Accident and Emergency department; Address; Algorithm Design; Antibiotic Therapy; Antibiotics; Area; Bacteria; Bacterial DNA; Bacterial Infections; Bacteriophages; Cells; Chronically Ill; Circulation; Cohort Studies; Computing Methodologies; Confusion; Consumption; DNA; DNA Sequence; DNA sequencing; Data; Data Set; Decision Making; Development; Diagnosis; Diagnostic; Disease; Early treatment; Elderly; Grant; High-Throughput Nucleotide Sequencing; Human; Individual; Infection; Lead; Lung; Medicine; Methods; Opportunistic Infections; Pathogenesis; Patient-Focused Outcomes; Patients; Performance; Phase; Plasma; Population Dynamics; Protocols documentation; Pseudomonas aeruginosa infection; Resolution; Risk; Role; Sampling; Sepsis; Serum; Signal Transduction; Site; Skin; Speed; Technology; Testing; Time; Transcript; Uncertainty; Virus; Work; biobank; cell free DNA; cohort; communicable disease diagnosis; computational pipelines; diagnostic accuracy; empowerment; human DNA; immunosuppressed; improved; insight; interest; microbial; molecular diagnostics; nanopore; next generation sequencing; novel; novel strategies; pathogen; pathogenic bacteria; pathogenic microbe; prospective; rapid diagnosis; scale up; septic patients; tool

Project Summary The rapid diagnosis of bacterial pathogens in septic patients is critical for early treatment decision making. Delayed diagnoses lead to a high rate of unnecessary antibiotic prescriptions and worse patient outcomes. One approach that has been used to improve sepsis diagnoses is circulating free DNA (cfFNA). Here, bacterial DNA present in serum is used to identify microbial pathogens and inform antibiotic treatment decisions. Unfortunately, while cfDNA approaches are good at identifying some sepsis pathogens, cfDNA does poorly at distinguishing bacterial infection from bacterial colonization. In settings where substantial background signal exists from related bacteria present in the gut, skin or lungs, existing approaches often confuse colonization as infection. Similarly, sensitivity can also be compromised by misinterpreting infection as colonization. We propose that this lack of resolution exists because cfDNA diagnostics only identify bacteria at the species level. They are unable to provide insight into the bacterial strain dynamics that underly infection. To address this, we have identified a novel approach for improving the performance of cfDNA in sepsis using bacteriophage –viruses produced by bacteria. Because bacteriophages are exquisitely specific to their particular host strain, the quantification of unique phages can provide insights into bacterial population dynamics at the strain level. Our preliminary data reveal that bacteriophage sequences are present in cfDNA collected from individuals with sepsis. Using previously collected and sequenced cfDNA data, we find that we can diagnose Pseudomonas aeruginosa infections using unique phage sequences that were not possible to diagnose with bacterial sequences alone. It may be possible to extend this approach to work with other bacterial pathogens. However, first we must develop robust computational pipelines for studying phages in cfDNA as the existing algorithms are designed to work with human and bacterial DNA. A further issue with cfDNA sequencing is speed. For cfDNA to be helpful in identifying sepsis pathogens, delivering timely results is critical. To this end, newer nanopore technologies have advantages over more time consuming illumina sequencing methods. However, the utility of nanopore sequencing for phage cfDNA is untested. We must demonstrate that nanopore sequencing of phages is both accurate and timely. Our hypothesis is that bacteriophage cfDNA can provide insight into the bacterial pathogenesis of sepsis. To test this, in the R21 portion of this grant we will develop computational protocols for studying phages in existing sepsis cfDNA datasets. Then, in the R33 portion of these studies we will develop rapid sequencing protocols for characterizing phage cfDNA in existing sepsis biorepository samples Together, these studies will generate the tools and conceptual frameworks needed to investigate the role of bacterial strains in sepsis. Moreover, these studies will set the stage for large, prospective human cohort studies to test the value of phage cfDNA in sepsis diagnosis.

项目摘要 化粪池患者中细菌病原体的快速诊断对于早期治疗决策至关重要。 延迟诊断导致不必要的抗生素处方和患者预后较差。 一种用于改善败血症诊断的方法是循环自由DNA（CFFNA）。这里， 血清中存在的细菌DNA用于鉴定微生物病原体并为抗生素治疗决定提供了信息。 不幸的是，尽管CFDNA方法擅长识别某些败血症病原体，但CFDNA的确很差 在区分细菌感染与细菌定植时。在背景信号的设置中 来自肠道，皮肤或肺中存在的相关细菌，现有方法通常将定植混淆为 感染。同样，通过将感染作为定殖，敏感性也可能会损害。我们建议 由于CFDNA诊断仅在物种水平上识别细菌，因此存在这种分辨率的缺乏。他们是 无法深入了解潜在感染的细菌菌株动力学。 为了解决这个问题，我们已经确定了一种新的方法，用于改善败血症中CfDNA的性能 细菌 - 细菌产生的病毒。因为细菌噬菌体完全特定于其特定 宿主应变，独特的噬菌体数量可以提供对细菌种群动态的见解 应变水平。我们的初步数据表明，从从 患有败血症的人。使用先前收集的和测序的CFDNA数据，我们发现我们可以诊断 使用独特的噬菌体序列铜绿假单胞菌感染，无法诊断 仅细菌序列。可以扩展这种方法与其他细菌病原体一起使用。 但是，首先，我们必须开发可靠的计算管道，以研究CFDNA中的噬菌体 算法旨在与人类和细菌DNA一起使用。 CFDNA测序的另一个问题是速度。 CFDNA有助于识别败血症病原体， 提供及时的结果至关重要。为此，较新的纳米技术在更多时间内具有优势 消费光明测序方法。但是，纳米孔测序对噬菌体CFDNA的实用性为 未经测试。我们必须证明噬菌体的纳米孔测序既准确又及时。 我们的假设是噬菌体CFDNA可以洞悉败血症的细菌发病机理。到 测试这一点，在这笔赠款的R21部分中，我们将开发用于研究现有噬菌体的计算协议 败血症CFDNA数据集。然后，在这些研究的R33部分中，我们将开发快速测序方案 表征现有的败血症生物剂样品中的噬菌体cfDNA 这些研究将共同​​生成研究研究作用所需的工具和概念框架 败血症中的细菌菌株。此外，这些研究将为大型，前瞻性人类队列研究奠定阶段 测试噬菌体CFDNA在败血症诊断中的值。