Structure and Diversity of Urinary Cell-Free DNA Informative of Host-Pathogen Interactions in Human Urinary Tract Infection

The risk of acquiring urinary tract infection (UTI) is very high for patients who have received kidney transplants; nearly half will develop bacteriuria and an eighth viruria via BK polyomavirus in the first three years following transplantation. Current methodology for diagnosing viral and bacterial infections, while well practiced, is often limited in scope. The advent of next generation sequencing of cell-free DNA has shown promise in its ability to capture multiple perspectives of human health from discovering the origin of cell-free DNA in plasma; such a diagnostic has not been readily applied to urinary cell-free DNA. In this study, we selected 141 urine samples from a cohort of kidney transplant patients, which were collected and stored at Weill Cornell Medical College - New York Presbyterian Hospital. Of these samples, 43 samples were determined to be culture positive (> 10,000 CFU) for one or more bacterial species, including E. coli, Enterococcus, Klebsiella, Haemophilus, and Raoultella. A separate 23 samples were confirmed for BK virus nephropathy, with presence of BK polyomavirus confirmed via qPCR. We also included 29 samples from 14 patients who did not develop UTIs in the first three months following transplantation, and 11 samples that were negative for BK virus nephropathy. For a full description of samples, please see the Supplementary Data 1 in https://pubmed.ncbi.nlm.nih.gov/29925834/. From these urine samples, we aimed to capture and sequence urinary cell-free DNA using a single-stranded library preparation. In addition, we performed whole-genome bisulfite sequencing to 170 plasma samples collected from 27 hematopoietic cell transplant patients who developed graft-versus host disease and BK polyomavirus infection. We also performed Sample-Intrinsic microbial DNA Found by Tagging and sequencing (SIFT-seq) on 196 cell-free DNA (cfDNA) samples (154 plasma and 42 urine) collected from five groups of subjects. For a full description of samples, please see publications in PMID: 35864089 and PMID: 35058359.]]>
For urinary cell-free DNA:Inclusion criteria:age at 18 - 90 years oldmale or femalekidney transplant recipient ability to provide informed consentExclusion criteria: inability to provide informed consent For amniotic fluid cell-free DNA:Inclusion criteria: pregnant mothers with or without chorioamnionitis, determined at the time of amniocentesis Exclusion criteria: inability to provide informed consentChildren were excluded.For plasma cell-free DNA:Inclusion criteria:meet the study criteria ability to provide informed consentExclusion criteria: inability to provide informed consentChildren were excluded. ]]>
Frozen urine samples were acquired in the past several years. Samples were processed (DNA extracted and corresponding libraries prepared and sequenced) from June 2016 to December 2021.Amniotic fluid samples were collected during amniocentesis and stored at -800C. Samples were processed (DNA extracted and corresponding libraries prepared and sequenced) from January 2017 to January 2019.Plasma sample were collected from from August 2018 to August 2021. ]]>