Host-Pathogen Interactions From Measurements of Urinary Cell Free DNA in Kidney Transplantation

肾移植中尿细胞游离 DNA 测量的宿主-病原体相互作用

• 批准号: 9375180
• 负责人: Iwijn De Vlaminck
• 金额: $ 24.87万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-06-26 至 2019-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9375180
• 关键词: Allografting; Antibiotic Therapy; Bacteria; Bacterial Genome; Biological Assay; Cells; Clinical; Complication; DNA Library; Data; Data Set; Development; End stage renal failure; Evaluation; Gene Expression; Genes; Genetic Fingerprintings; Genomics; Goals; Gold; Growth; Immune response; Immunocompromised Host; Immunologics; Incidence; Infection; Injury; Kidney Transplantation; Longevity; Measurement; Messenger RNA; Metagenomics; Monitor; Morbidity - disease rate; Multi-Drug Resistance; Natural Immunity; Nucleosomes; Pathogenesis; Patients; Pilot Projects; Population; Preparation; Recruitment Activity; Recurrence; Regulator Genes; Replication Origin; Resolution; Sampling; Sequence Analysis; Shotgun Sequencing; Shotguns; Single-Stranded DNA; Specimen; Structure; Study Section; Symbiosis; Techniques; Testing; Time; Tissues; Transcription Initiation Site; Transplant Recipients; Transplantation; Urinalysis; Urinary tract infection; Urine; Visit; Work; adaptive immunity; base; biobank; cell free DNA; cell injury; co-infection; cohort; diagnosis standard; differential expression; microbiome; microbiota; mortality; novel; pathogen; promoter; resistance mutation; response; tool; urinary

Kidney transplantation is the preferred treatment option for end stage kidney disease, but infectious and immunological complications limit the lifespan of kidney transplants. Urinary tract infections (UTI) occur at an alarmingly high rate after kidney transplantation and are associated with urosepsis, allograft loss, and mortality. The current gold standard for diagnosis of UTI is bacterial culture. Culture-based tests are, however, limited by their ability to detect only culturable bacteria, do not inform about co-infections and commensal microbiota, do not inform about bacterial growth dynamics, and do not inform about the host's response to infection. The goal of this proposal is to invent and apply genomic assays of urinary cell-free DNA that can simultaneously inform about the composition of the urinary microbiome, growth dynamics of pathogens associated with UTI, tissue injury in the graft, and the host's immune response to infection. A unique opportunity for this work is our large cohort of over 250 kidney transplant recipients in which we have serial biobanked urine specimens as well as routine conventional urine cultures in the first 3 months of transplantation. Using a newly developed single-stranded DNA library preparation technique, we will profile 290 urinary samples for cfDNA: i) serial samples from transplant recipients with UTI (prior to, during, and after UTI) ii) serial samples from recipients with recurrent UTIs; iii) samples from recipients with asymptomatic UTI and negative urinanalysis; and iv) samples from recipients who do not develop UTI in the first 3 months of transplantation. We will profile the dynamic urinary microbiome prior to, during, and after UTI in kidney transplantation. We will investigate whether cfDNA profiles are associated with UTI as well as whether they can anticipate UTI, UTI recurrence, and multi-drug resistant UTI. Based on the concept that bacterial genome replication rates and growth dynamics can be estimated from analyses of metagenomic sequence coverage, we will estimate growth dynamics over time and in response to antibiotic therapy in patients with UTI and recurrent UTI. We will monitor the host response to UTI via analyses of nucleosome footprints comprised within cfDNA and via donor specific cfDNA. Our pilot studies indicate that i) the structures of nucleosomal arrays in transcriptional regulatory elements are preserved and comprised within urinary cfDNA, and ii) that an analysis of nucleosomal occupancy in gene promoters can be used to quantify gene expression. We will investigate the differential host's response in subjects who have symptomatic UTI with positive urinalysis and asymptomatic UTI with negative urinalysis and the differential host's response in UTI patients after antibiotic treatment. We will assess tissue injury to the allograft in the scope of UTI, using donor specific cfDNA as a marker of allograft injury. Successful implementation of this proposal will provide a novel understanding of the pathogenesis of UTI as well as open new avenues for the utilization of cfDNA profiling as a sensitive and noninvasive transplant monitoring tool.

肾移植是终末期肾病的首选治疗选择，但具有传染性和传染性 免疫并发症限制了肾移植的寿命。尿路感染（UTI）发生在 肾移植后发生率高得惊人，并与尿脓毒症、同种异体移植物丢失和死亡率相关。 目前诊断尿路感染的金标准是细菌培养。然而，基于文化的测试受到以下因素的限制： 它们仅检测可培养细菌的能力，不会告知合并感染和共生微生物群， 不告知细菌生长动态，也不告知宿主对感染的反应。 该提案的目标是发明并应用尿液游离 DNA 的基因组分析方法， 同时了解尿液微生物组的组成、病原体的生长动态 与尿路感染、移植物组织损伤以及宿主对感染的免疫反应有关。一个独特的 这项工作的机会是我们有超过 250 名肾移植受者的大队列，其中我们有连续的 移植前 3 个月的生物库尿液标本以及常规常规尿液培养。 使用新开发的单链 DNA 文库制备技术，我们将分析 290 个尿液样本 对于 cfDNA： i) 来自患有 UTI 的移植受者的连续样本（UTI 之前、期间和之后） ii) 连续样本 来自患有复发性尿路感染的接受者； iii) 来自无症状尿路感染且尿液分析呈阴性的接受者的样本； iv) 移植前 3 个月内未发生尿路感染的受者的样本。 我们将分析肾移植尿路感染之前、期间和之后的动态泌尿微生物组。 我们将研究 cfDNA 谱是否与 UTI 相关以及它们是否可以预测 UTI， 尿路感染复发和多重耐药尿路感染。基于细菌基因组复制率和 生长动态可以通过宏基因组序列覆盖度的分析来估计，我们将估计生长 尿路感染和复发性尿路感染患者随时间的动态变化以及对抗生素治疗的反应。 我们将通过分析 cfDNA 中包含的核小体足迹来监测宿主对 UTI 的反应 并通过供体特异性 cfDNA。我们的初步研究表明 i) 核小体阵列的结构 转录调控元件被保留并包含在尿液 cfDNA 中，并且 ii) 分析 基因启动子中核小体占据的变化可用于量化基因表达。我们将调查 尿液分析阳性的有症状尿路感染受试者和无症状尿路感染受试者的宿主反应不同 尿路感染患者在抗生素治疗后尿液分析结果呈阴性，宿主反应也不同。我们将 使用供体特异性 cfDNA 作为同种异体移植物损伤的标记，评估 UTI 范围内同种异体移植物的组织损伤。 该提案的成功实施也将为尿路感染的发病机制提供新的认识 为利用 cfDNA 分析作为敏感和非侵入性移植监测开辟了新途径 工具。