Characterizing the molecular composition and diagnostic potential of Mycobacterium tuberculosis urinary cell-free DNA using next-generation sequencing.
Cell-free DNA
DNA sequencing
Diagnostics
Molecular diagnosis
Mycobacterium tuberculosis
Transrenal DNA
Urine
Journal
International journal of infectious diseases : IJID : official publication of the International Society for Infectious Diseases
ISSN: 1878-3511
Titre abrégé: Int J Infect Dis
Pays: Canada
ID NLM: 9610933
Informations de publication
Date de publication:
Nov 2021
Nov 2021
Historique:
received:
21
07
2021
revised:
16
09
2021
accepted:
17
09
2021
pubmed:
26
9
2021
medline:
15
12
2021
entrez:
25
9
2021
Statut:
ppublish
Résumé
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. 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. 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. TB urine cfDNA is a potentially powerful biomarker but is highly fragmented, necessitating special procedures to maximize its recovery and detection.
Sections du résumé
BACKGROUND
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
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
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
CONCLUSIONS
TB urine cfDNA is a potentially powerful biomarker but is highly fragmented, necessitating special procedures to maximize its recovery and detection.
Identifiants
pubmed: 34562627
pii: S1201-9712(21)00750-5
doi: 10.1016/j.ijid.2021.09.042
pmc: PMC8627387
pii:
doi:
Substances chimiques
Biomarkers
0
Cell-Free Nucleic Acids
0
DNA, Bacterial
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
330-337Subventions
Organisme : NIAID NIH HHS
ID : K23 AI140918
Pays : United States
Informations de copyright
Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.
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