Contaminating viral sequences in high-throughput sequencing viromics: a linkage study of 700 sequencing libraries

Maria Asplund, Kristín Rós Kjartansdóttir, Sarah Mollerup, Lasse Vinner, Helena Fridholm, José A. R. Herrera, Jens Friis-Nielsen, Thomas Arn Hansen, Randi Holm Jensen, Ida Broman Nielsen, Stine Raith Richter, Alba Rey-Iglesia, Maria Luisa Matey-Hernandez, David E. Alquezar-Planas, Pernille V. S. Olsen, Thomas Sicheritz-Pontén, Eske Willerslev, Ole Lund, Søren Brunak, Tobias Mourier & 3 others Lars Peter Nielsen, Jose M. G. Izarzugaza, Anders Johannes Hansen*

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

Sample preparation for High-throughput sequencing (HTS) includes treatment with various laboratory components, potentially carrying viral nucleic acids, the extent of which has not been thoroughly investigated. Our aim was to systematically examine a diverse repertoire of laboratory components used to prepare samples for HTS in order to identify contaminating viral sequences. A total of 322 samples of mainly human origin were analysed using eight protocols, applying a wide variety of laboratory components. Several samples (60% of human specimens) were processed by different protocols. In total 712 sequencing libraries were investigated for viral sequence contamination. Among sequences showing similarity to viruses, 493 were significantly associated to the use of laboratory components. Each of these viral sequences showed sporadic appearance, only being identified in a subset of the samples treated with the linked laboratory component, and some were not identified in the non-template control (NTC) samples. Remarkably, more than 65% of all viral sequences identified were within viral clusters linked to the use of laboratory components. We show that high prevalence of contaminating viral sequences can be expected in HTS-based virome data and provide an extensive list of novel contaminating viral sequences that can be used for evaluation of viral findings in future virome and metagenome studies. Moreover we show that detection can be problematic due to stochastic appearance and limited NTCs. Although the exact origin of these viral sequences requires further research, our results support laboratory component linked viral sequence contamination of both biological and synthetic origin.
Original languageEnglish
JournalClinical Microbiology and Infection
Volume25
Issue number10
Pages (from-to)1277-1285
ISSN1198-743X
DOIs
Publication statusPublished - 2019

Keywords

  • Contamination
  • Cluster
  • High-throughput sequencing
  • Laboratory component
  • Metagenomic
  • Next generation sequencing
  • Nucleic acid
  • Virome
  • Virus

Cite this

Asplund, M., Kjartansdóttir, K. R., Mollerup, S., Vinner, L., Fridholm, H., Herrera, J. A. R., ... Hansen, A. J. (2019). Contaminating viral sequences in high-throughput sequencing viromics: a linkage study of 700 sequencing libraries. Clinical Microbiology and Infection, 25(10), 1277-1285. https://doi.org/10.1016/j.cmi.2019.04.028
Asplund, Maria ; Kjartansdóttir, Kristín Rós ; Mollerup, Sarah ; Vinner, Lasse ; Fridholm, Helena ; Herrera, José A. R. ; Friis-Nielsen, Jens ; Hansen, Thomas Arn ; Jensen, Randi Holm ; Nielsen, Ida Broman ; Richter, Stine Raith ; Rey-Iglesia, Alba ; Matey-Hernandez, Maria Luisa ; Alquezar-Planas, David E. ; Olsen, Pernille V. S. ; Sicheritz-Pontén, Thomas ; Willerslev, Eske ; Lund, Ole ; Brunak, Søren ; Mourier, Tobias ; Nielsen, Lars Peter ; Izarzugaza, Jose M. G. ; Hansen, Anders Johannes. / Contaminating viral sequences in high-throughput sequencing viromics: a linkage study of 700 sequencing libraries. In: Clinical Microbiology and Infection. 2019 ; Vol. 25, No. 10. pp. 1277-1285.
@article{40cc8bdfd3754d97aa6a13890b84e5d9,
title = "Contaminating viral sequences in high-throughput sequencing viromics: a linkage study of 700 sequencing libraries",
abstract = "Sample preparation for High-throughput sequencing (HTS) includes treatment with various laboratory components, potentially carrying viral nucleic acids, the extent of which has not been thoroughly investigated. Our aim was to systematically examine a diverse repertoire of laboratory components used to prepare samples for HTS in order to identify contaminating viral sequences. A total of 322 samples of mainly human origin were analysed using eight protocols, applying a wide variety of laboratory components. Several samples (60{\%} of human specimens) were processed by different protocols. In total 712 sequencing libraries were investigated for viral sequence contamination. Among sequences showing similarity to viruses, 493 were significantly associated to the use of laboratory components. Each of these viral sequences showed sporadic appearance, only being identified in a subset of the samples treated with the linked laboratory component, and some were not identified in the non-template control (NTC) samples. Remarkably, more than 65{\%} of all viral sequences identified were within viral clusters linked to the use of laboratory components. We show that high prevalence of contaminating viral sequences can be expected in HTS-based virome data and provide an extensive list of novel contaminating viral sequences that can be used for evaluation of viral findings in future virome and metagenome studies. Moreover we show that detection can be problematic due to stochastic appearance and limited NTCs. Although the exact origin of these viral sequences requires further research, our results support laboratory component linked viral sequence contamination of both biological and synthetic origin.",
keywords = "Contamination, Cluster, High-throughput sequencing, Laboratory component, Metagenomic, Next generation sequencing, Nucleic acid, Virome, Virus",
author = "Maria Asplund and Kjartansd{\'o}ttir, {Krist{\'i}n R{\'o}s} and Sarah Mollerup and Lasse Vinner and Helena Fridholm and Herrera, {Jos{\'e} A. R.} and Jens Friis-Nielsen and Hansen, {Thomas Arn} and Jensen, {Randi Holm} and Nielsen, {Ida Broman} and Richter, {Stine Raith} and Alba Rey-Iglesia and Matey-Hernandez, {Maria Luisa} and Alquezar-Planas, {David E.} and Olsen, {Pernille V. S.} and Thomas Sicheritz-Pont{\'e}n and Eske Willerslev and Ole Lund and S{\o}ren Brunak and Tobias Mourier and Nielsen, {Lars Peter} and Izarzugaza, {Jose M. G.} and Hansen, {Anders Johannes}",
year = "2019",
doi = "10.1016/j.cmi.2019.04.028",
language = "English",
volume = "25",
pages = "1277--1285",
journal = "Clinical Microbiology and Infection",
issn = "1198-743X",
publisher = "Elsevier",
number = "10",

}

Asplund, M, Kjartansdóttir, KR, Mollerup, S, Vinner, L, Fridholm, H, Herrera, JAR, Friis-Nielsen, J, Hansen, TA, Jensen, RH, Nielsen, IB, Richter, SR, Rey-Iglesia, A, Matey-Hernandez, ML, Alquezar-Planas, DE, Olsen, PVS, Sicheritz-Pontén, T, Willerslev, E, Lund, O, Brunak, S, Mourier, T, Nielsen, LP, Izarzugaza, JMG & Hansen, AJ 2019, 'Contaminating viral sequences in high-throughput sequencing viromics: a linkage study of 700 sequencing libraries', Clinical Microbiology and Infection, vol. 25, no. 10, pp. 1277-1285. https://doi.org/10.1016/j.cmi.2019.04.028

Contaminating viral sequences in high-throughput sequencing viromics: a linkage study of 700 sequencing libraries. / Asplund, Maria; Kjartansdóttir, Kristín Rós; Mollerup, Sarah; Vinner, Lasse; Fridholm, Helena; Herrera, José A. R.; Friis-Nielsen, Jens; Hansen, Thomas Arn; Jensen, Randi Holm; Nielsen, Ida Broman; Richter, Stine Raith; Rey-Iglesia, Alba; Matey-Hernandez, Maria Luisa; Alquezar-Planas, David E.; Olsen, Pernille V. S.; Sicheritz-Pontén, Thomas; Willerslev, Eske; Lund, Ole; Brunak, Søren; Mourier, Tobias; Nielsen, Lars Peter; Izarzugaza, Jose M. G.; Hansen, Anders Johannes.

In: Clinical Microbiology and Infection, Vol. 25, No. 10, 2019, p. 1277-1285.

Research output: Contribution to journalJournal articleResearchpeer-review

TY - JOUR

T1 - Contaminating viral sequences in high-throughput sequencing viromics: a linkage study of 700 sequencing libraries

AU - Asplund, Maria

AU - Kjartansdóttir, Kristín Rós

AU - Mollerup, Sarah

AU - Vinner, Lasse

AU - Fridholm, Helena

AU - Herrera, José A. R.

AU - Friis-Nielsen, Jens

AU - Hansen, Thomas Arn

AU - Jensen, Randi Holm

AU - Nielsen, Ida Broman

AU - Richter, Stine Raith

AU - Rey-Iglesia, Alba

AU - Matey-Hernandez, Maria Luisa

AU - Alquezar-Planas, David E.

AU - Olsen, Pernille V. S.

AU - Sicheritz-Pontén, Thomas

AU - Willerslev, Eske

AU - Lund, Ole

AU - Brunak, Søren

AU - Mourier, Tobias

AU - Nielsen, Lars Peter

AU - Izarzugaza, Jose M. G.

AU - Hansen, Anders Johannes

PY - 2019

Y1 - 2019

N2 - Sample preparation for High-throughput sequencing (HTS) includes treatment with various laboratory components, potentially carrying viral nucleic acids, the extent of which has not been thoroughly investigated. Our aim was to systematically examine a diverse repertoire of laboratory components used to prepare samples for HTS in order to identify contaminating viral sequences. A total of 322 samples of mainly human origin were analysed using eight protocols, applying a wide variety of laboratory components. Several samples (60% of human specimens) were processed by different protocols. In total 712 sequencing libraries were investigated for viral sequence contamination. Among sequences showing similarity to viruses, 493 were significantly associated to the use of laboratory components. Each of these viral sequences showed sporadic appearance, only being identified in a subset of the samples treated with the linked laboratory component, and some were not identified in the non-template control (NTC) samples. Remarkably, more than 65% of all viral sequences identified were within viral clusters linked to the use of laboratory components. We show that high prevalence of contaminating viral sequences can be expected in HTS-based virome data and provide an extensive list of novel contaminating viral sequences that can be used for evaluation of viral findings in future virome and metagenome studies. Moreover we show that detection can be problematic due to stochastic appearance and limited NTCs. Although the exact origin of these viral sequences requires further research, our results support laboratory component linked viral sequence contamination of both biological and synthetic origin.

AB - Sample preparation for High-throughput sequencing (HTS) includes treatment with various laboratory components, potentially carrying viral nucleic acids, the extent of which has not been thoroughly investigated. Our aim was to systematically examine a diverse repertoire of laboratory components used to prepare samples for HTS in order to identify contaminating viral sequences. A total of 322 samples of mainly human origin were analysed using eight protocols, applying a wide variety of laboratory components. Several samples (60% of human specimens) were processed by different protocols. In total 712 sequencing libraries were investigated for viral sequence contamination. Among sequences showing similarity to viruses, 493 were significantly associated to the use of laboratory components. Each of these viral sequences showed sporadic appearance, only being identified in a subset of the samples treated with the linked laboratory component, and some were not identified in the non-template control (NTC) samples. Remarkably, more than 65% of all viral sequences identified were within viral clusters linked to the use of laboratory components. We show that high prevalence of contaminating viral sequences can be expected in HTS-based virome data and provide an extensive list of novel contaminating viral sequences that can be used for evaluation of viral findings in future virome and metagenome studies. Moreover we show that detection can be problematic due to stochastic appearance and limited NTCs. Although the exact origin of these viral sequences requires further research, our results support laboratory component linked viral sequence contamination of both biological and synthetic origin.

KW - Contamination

KW - Cluster

KW - High-throughput sequencing

KW - Laboratory component

KW - Metagenomic

KW - Next generation sequencing

KW - Nucleic acid

KW - Virome

KW - Virus

U2 - 10.1016/j.cmi.2019.04.028

DO - 10.1016/j.cmi.2019.04.028

M3 - Journal article

VL - 25

SP - 1277

EP - 1285

JO - Clinical Microbiology and Infection

JF - Clinical Microbiology and Infection

SN - 1198-743X

IS - 10

ER -