Structure and diversity of urinary cell-free DNA informative of host-pathogen interactions in human urinary tract infection

Burnham, Philip; Dadhania, Darshana; Heyang, Michael; Chen, F; Lf, Westblade; Suthanthiran, Manikkam; Lee, Myeong Soo; Vlaminck, Iwijn De

doi:10.1101/177402

Cited by 2 publications

(11 citation statements)

References 63 publications

(413 reference statements)

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“…We further incorporated a known-template, negative control in the library preparation procedures to identify any remaining contaminant sequences. The use of a negative control is recommended for metagenomics studies [16], and was implemented in our previous work [2,17]. Here, we compared the microbial abundance detected in samples to those in controls to set a baseline for environmental contamination.…”

Section: Resultsmentioning

confidence: 99%

“…We evaluated and optimized LBBC using a dataset available from a recently published study that assessed the utility of urinary cfDNA for the monitoring of bacterial infection of the urinary tract [2].…”

Section: Resultsmentioning

confidence: 99%

“…The three filtering steps implemented in LLBC are appropriate for the analysis of any low-biomass sample, not limited to cfDNA isolates, and can be readily implemented, in a modular fashion, provided that: (1) the total DNA biomass going into the sample preparation is measured and recorded, (2) batch information is available, and (3) the microbial abundance is determined by a sequence-based alignment method [15]. This last point is of importance, because several popular algorithms for metagenomic sequence classification, including Metaphlan, which relies on reduced reference genomes [36].…”

Section: Discussionmentioning

confidence: 99%

“…Sample description -urinary cfDNA. Forty-four sample datasets were selected from a recent study [2]. (heart rate >100 beats/min), and (5) maternal leukocytosis (leukocyte count >15,000 cells/mm 3 ) [22,26].…”

Section: Methodsmentioning

confidence: 99%

“…Datasets were selected from the study from one of two groups: 1) UTI -those corresponding to a same-day positive urine culture (>10,000 CFU/mL) indicating monomicrobial E. coli, Enterococcus faecium, or Enterococcus faecalis UTI. A single sample from the original study[2] (GU14), was excluded due to the high likelihood that it was R. ornithinolytica infection incorrectly diagnosed as an E. coli UTI. 2) No UTI -samples from patients with same-day negative standard urine culture and no microorganisms detected at earlier or later dates.…”

mentioning

confidence: 99%

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Separating the signal from the noise in metagenomic cell-free DNA sequencing

Burnham

Gomez‐Lopez

Heyang

et al. 2020

Preprint

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Background: Cell-free DNA (cfDNA) in blood, urine and other biofluids provides a unique window into human health. A proportion of cfDNA is derived from bacteria and viruses, creating opportunities for the diagnosis of infection via metagenomic sequencing. The total biomass of microbial-derived cfDNA in clinical isolates is low, which makes metagenomic cfDNA sequencing susceptible to contamination and alignment noise. Results: Here, we report Low Biomass Background Correction (LBBC), a bioinformatics noise filtering tool informed by the uniformity of the coverage of microbial genomes and the batch variation in the absolute abundance of microbial cfDNA. We demonstrate that LBBC leads to a dramatic reduction in false positive rate while minimally affecting the true positive rate for a cfDNA test to screen for urinary tract infection. We next performed high throughput sequencing of cfDNA in amniotic fluid collected from term uncomplicated pregnancies or those complicated with clinical chorioamnionitis with and without intra-amniotic infection. Conclusions: The data provide unique insight into the properties of fetal and maternal cfDNA in amniotic fluid, demonstrate the utility of cfDNA to screen for intra-amniotic infection, support the view that the amniotic fluid is sterile during normal pregnancy, and reveal cases of intra-amniotic inflammation without infection at term.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%