Characterizing the molecular composition and diagnostic potential of Mycobacterium tuberculosis urinary cell-free DNA using next-generation sequencing

Background: Urine cell-free DNA (cfDNA) is an attractive target for diagnosing pulmonary Mycobacterium tuberculosis (MTB) infection, but has not been thoroughly characterized as a biomarker. Methods: This study was performed to investigate the size and composition of urine cfDNA from tuberculosis (T...

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Autores principales:	Amy Oreskovic, Adam Waalkes, Elizabeth A. Holmes, Christopher A. Rosenthal, Douglas P.K. Wilson, Adrienne E. Shapiro, Paul K. Drain, Barry R. Lutz, Stephen J. Salipante
Formato:	article
Lenguaje:	EN
Publicado:	Elsevier 2021
Materias:	Mycobacterium tuberculosis DNA sequencing Cell-free DNA Transrenal DNA Urine Diagnostics Infectious and parasitic diseases RC109-216
Acceso en línea:	https://doaj.org/article/673829f55a02480389481fb7b5aca49a
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spelling	oai:doaj.org-article:673829f55a02480389481fb7b5aca49a2021-11-30T04:14:20ZCharacterizing the molecular composition and diagnostic potential of Mycobacterium tuberculosis urinary cell-free DNA using next-generation sequencing1201-971210.1016/j.ijid.2021.09.042https://doaj.org/article/673829f55a02480389481fb7b5aca49a2021-11-01T00:00:00Zhttp://www.sciencedirect.com/science/article/pii/S1201971221007505https://doaj.org/toc/1201-9712Background: Urine cell-free DNA (cfDNA) is an attractive target for diagnosing pulmonary Mycobacterium tuberculosis (MTB) infection, but has not been thoroughly characterized as a biomarker. Methods: This study was performed to investigate the size and composition of urine cfDNA from tuberculosis (TB) patients with minimal bias using next-generation sequencing (NGS). A combination of DNA extraction and single-stranded sequence library preparation methods demonstrated to recover short, highly degraded cfDNA fragments was employed. Urine cfDNA from 10 HIV-positive patients with pulmonary TB and two MTB-negative controls was examined. Results: MTB-derived cfDNA was identifiable by NGS from all MTB-positive patients and was absent from negative controls. MTB cfDNA was significantly shorter than human cfDNA, with median fragment lengths of ≤19–52 bp and 42–92 bp, respectively. MTB cfDNA abundance increased exponentially with decreased fragment length, having a peak fragment length of ≤19 bp in most samples. In addition, we identified a larger fraction of short human genomic cfDNA, ranging from 29 to 53 bp, than previously reported. Urine cfDNA fragments spanned the MTB genome with relative uniformity, but nucleic acids derived from multicopy elements were proportionately over-represented. Conclusions: TB urine cfDNA is a potentially powerful biomarker but is highly fragmented, necessitating special procedures to maximize its recovery and detection.Amy OreskovicAdam WaalkesElizabeth A. HolmesChristopher A. RosenthalDouglas P.K. WilsonAdrienne E. ShapiroPaul K. DrainBarry R. LutzStephen J. SalipanteElsevierarticleMycobacterium tuberculosisDNA sequencingCell-free DNATransrenal DNAUrineDiagnosticsInfectious and parasitic diseasesRC109-216ENInternational Journal of Infectious Diseases, Vol 112, Iss , Pp 330-337 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Mycobacterium tuberculosis DNA sequencing Cell-free DNA Transrenal DNA Urine Diagnostics Infectious and parasitic diseases RC109-216
spellingShingle	Mycobacterium tuberculosis DNA sequencing Cell-free DNA Transrenal DNA Urine Diagnostics Infectious and parasitic diseases RC109-216 Amy Oreskovic Adam Waalkes Elizabeth A. Holmes Christopher A. Rosenthal Douglas P.K. Wilson Adrienne E. Shapiro Paul K. Drain Barry R. Lutz Stephen J. Salipante Characterizing the molecular composition and diagnostic potential of Mycobacterium tuberculosis urinary cell-free DNA using next-generation sequencing
description	Background: Urine cell-free DNA (cfDNA) is an attractive target for diagnosing pulmonary Mycobacterium tuberculosis (MTB) infection, but has not been thoroughly characterized as a biomarker. Methods: This study was performed to investigate the size and composition of urine cfDNA from tuberculosis (TB) patients with minimal bias using next-generation sequencing (NGS). A combination of DNA extraction and single-stranded sequence library preparation methods demonstrated to recover short, highly degraded cfDNA fragments was employed. Urine cfDNA from 10 HIV-positive patients with pulmonary TB and two MTB-negative controls was examined. Results: MTB-derived cfDNA was identifiable by NGS from all MTB-positive patients and was absent from negative controls. MTB cfDNA was significantly shorter than human cfDNA, with median fragment lengths of ≤19–52 bp and 42–92 bp, respectively. MTB cfDNA abundance increased exponentially with decreased fragment length, having a peak fragment length of ≤19 bp in most samples. In addition, we identified a larger fraction of short human genomic cfDNA, ranging from 29 to 53 bp, than previously reported. Urine cfDNA fragments spanned the MTB genome with relative uniformity, but nucleic acids derived from multicopy elements were proportionately over-represented. Conclusions: TB urine cfDNA is a potentially powerful biomarker but is highly fragmented, necessitating special procedures to maximize its recovery and detection.
format	article
author	Amy Oreskovic Adam Waalkes Elizabeth A. Holmes Christopher A. Rosenthal Douglas P.K. Wilson Adrienne E. Shapiro Paul K. Drain Barry R. Lutz Stephen J. Salipante
author_facet	Amy Oreskovic Adam Waalkes Elizabeth A. Holmes Christopher A. Rosenthal Douglas P.K. Wilson Adrienne E. Shapiro Paul K. Drain Barry R. Lutz Stephen J. Salipante
author_sort	Amy Oreskovic
title	Characterizing the molecular composition and diagnostic potential of Mycobacterium tuberculosis urinary cell-free DNA using next-generation sequencing
title_short	Characterizing the molecular composition and diagnostic potential of Mycobacterium tuberculosis urinary cell-free DNA using next-generation sequencing
title_full	Characterizing the molecular composition and diagnostic potential of Mycobacterium tuberculosis urinary cell-free DNA using next-generation sequencing
title_fullStr	Characterizing the molecular composition and diagnostic potential of Mycobacterium tuberculosis urinary cell-free DNA using next-generation sequencing
title_full_unstemmed	Characterizing the molecular composition and diagnostic potential of Mycobacterium tuberculosis urinary cell-free DNA using next-generation sequencing
title_sort	characterizing the molecular composition and diagnostic potential of mycobacterium tuberculosis urinary cell-free dna using next-generation sequencing
publisher	Elsevier
publishDate	2021
url	https://doaj.org/article/673829f55a02480389481fb7b5aca49a
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Characterizing the molecular composition and diagnostic potential of Mycobacterium tuberculosis urinary cell-free DNA using next-generation sequencing

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