Multicenter benchmarking of short and long read wet lab protocols for clinical viral metagenomics
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Multicenter benchmarking of short and long read wet lab protocols for clinical viral metagenomics. / Lopez-Labrador, F Xavier; Huber, Michael; Sidorov, Igor A; Brown, Julianne R; Cuypers, Lize; Laenen, Lies; Vanmechelen, Bert; Maes, Piet; Fischer, Nicole; Pichler, Ian; Storey, Nathaniel; Atkinson, Laura; Schmutz, Stefan; Kufner, Verena; van Boheemen, Sander; Mulders, Claudia E; Grundhoff, Adam; Blümke, Patrick; Robitaille, Alexis; Cinek, Ondrej; Hubáčková, Klára; Mourik, Kees; Boers, Stefan A; Stauber, Lea; Salmona, Maud; Cappy, Pierre; Ramette, Alban; Franze', Alessandra; LeGoff, Jerome; Claas, Eric C J; Rodriguez, Christophe; de Vries, Jutte J C; European Society of Clinical Virology (ESCV) Network on Next-Generation Sequencing (ENNGS).
in: J CLIN VIROL, Jahrgang 173, 08.2024, S. 105695.Publikationen: SCORING: Beitrag in Fachzeitschrift/Zeitung › SCORING: Zeitschriftenaufsatz › Forschung › Begutachtung
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TY - JOUR
T1 - Multicenter benchmarking of short and long read wet lab protocols for clinical viral metagenomics
AU - Lopez-Labrador, F Xavier
AU - Huber, Michael
AU - Sidorov, Igor A
AU - Brown, Julianne R
AU - Cuypers, Lize
AU - Laenen, Lies
AU - Vanmechelen, Bert
AU - Maes, Piet
AU - Fischer, Nicole
AU - Pichler, Ian
AU - Storey, Nathaniel
AU - Atkinson, Laura
AU - Schmutz, Stefan
AU - Kufner, Verena
AU - van Boheemen, Sander
AU - Mulders, Claudia E
AU - Grundhoff, Adam
AU - Blümke, Patrick
AU - Robitaille, Alexis
AU - Cinek, Ondrej
AU - Hubáčková, Klára
AU - Mourik, Kees
AU - Boers, Stefan A
AU - Stauber, Lea
AU - Salmona, Maud
AU - Cappy, Pierre
AU - Ramette, Alban
AU - Franze', Alessandra
AU - LeGoff, Jerome
AU - Claas, Eric C J
AU - Rodriguez, Christophe
AU - de Vries, Jutte J C
AU - European Society of Clinical Virology (ESCV) Network on Next-Generation Sequencing (ENNGS)
N1 - Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.
PY - 2024/8
Y1 - 2024/8
N2 - Metagenomics is gradually being implemented for diagnosing infectious diseases. However, in-depth protocol comparisons for viral detection have been limited to individual sets of experimental workflows and laboratories. In this study, we present a benchmark of metagenomics protocols used in clinical diagnostic laboratories initiated by the European Society for Clinical Virology (ESCV) Network on NGS (ENNGS). A mock viral reference panel was designed to mimic low biomass clinical specimens. The panel was used to assess the performance of twelve metagenomic wet lab protocols currently in use in the diagnostic laboratories of participating ENNGS member institutions. Both Illumina and Nanopore, shotgun and targeted capture probe protocols were included. Performance metrics sensitivity, specificity, and quantitative potential were assessed using a central bioinformatics pipeline. Overall, viral pathogens with loads down to 104 copies/ml (corresponding to CT values of 31 in our PCR assays) were detected by all the evaluated metagenomic wet lab protocols. In contrast, lower abundant mixed viruses of CT values of 35 and higher were detected only by a minority of the protocols. Considering the reference panel as the gold standard, optimal thresholds to define a positive result were determined per protocol, based on the horizontal genome coverage. Implementing these thresholds, sensitivity and specificity of the protocols ranged from 67 to 100 % and 87 to 100 %, respectively. A variety of metagenomic protocols are currently in use in clinical diagnostic laboratories. Detection of low abundant viral pathogens and mixed infections remains a challenge, implying the need for standardization of metagenomic analysis for use in clinical settings.
AB - Metagenomics is gradually being implemented for diagnosing infectious diseases. However, in-depth protocol comparisons for viral detection have been limited to individual sets of experimental workflows and laboratories. In this study, we present a benchmark of metagenomics protocols used in clinical diagnostic laboratories initiated by the European Society for Clinical Virology (ESCV) Network on NGS (ENNGS). A mock viral reference panel was designed to mimic low biomass clinical specimens. The panel was used to assess the performance of twelve metagenomic wet lab protocols currently in use in the diagnostic laboratories of participating ENNGS member institutions. Both Illumina and Nanopore, shotgun and targeted capture probe protocols were included. Performance metrics sensitivity, specificity, and quantitative potential were assessed using a central bioinformatics pipeline. Overall, viral pathogens with loads down to 104 copies/ml (corresponding to CT values of 31 in our PCR assays) were detected by all the evaluated metagenomic wet lab protocols. In contrast, lower abundant mixed viruses of CT values of 35 and higher were detected only by a minority of the protocols. Considering the reference panel as the gold standard, optimal thresholds to define a positive result were determined per protocol, based on the horizontal genome coverage. Implementing these thresholds, sensitivity and specificity of the protocols ranged from 67 to 100 % and 87 to 100 %, respectively. A variety of metagenomic protocols are currently in use in clinical diagnostic laboratories. Detection of low abundant viral pathogens and mixed infections remains a challenge, implying the need for standardization of metagenomic analysis for use in clinical settings.
KW - Metagenomics/methods
KW - Humans
KW - Benchmarking
KW - Viruses/genetics
KW - Sensitivity and Specificity
KW - High-Throughput Nucleotide Sequencing/methods
KW - Virus Diseases/diagnosis
KW - Computational Biology/methods
U2 - 10.1016/j.jcv.2024.105695
DO - 10.1016/j.jcv.2024.105695
M3 - SCORING: Journal article
C2 - 38823290
VL - 173
SP - 105695
JO - J CLIN VIROL
JF - J CLIN VIROL
SN - 1386-6532
ER -
