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A validation scale to determine the readiness of environmental DNA assays for routine species monitoring

View ORCID ProfileBettina Thalinger, View ORCID ProfileKristy Deiner, View ORCID ProfileLynsey R. Harper, Helen C. Rees, View ORCID ProfileRosetta C. Blackman, View ORCID ProfileDaniela Sint, View ORCID ProfileMichael Traugott, View ORCID ProfileCaren S. Goldberg, Kat Bruce
doi: https://doi.org/10.1101/2020.04.27.063990
Bettina Thalinger
1Centre for Biodiversity Genomics, University of Guelph, 50 Stone Road E, N1G 2W1, Guelph, Ontario, Canada
2Department of Integrative Biology, College of Biological Sciences, University of Guelph, 50 Stone Road E, N1G2W1, Guelph, Ontario, Canada
3Department of Zoology, University of Innsbruck, Technikerstr. 25, 6020, Innsbruck, Austria
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  • For correspondence: bettina.thalinger@gmail.com
Kristy Deiner
4Natural History Museum, Cromwell Road, London, SW7 5BD, UK
5Institute of Biogeochemistry and Pollutant Dynamics, ETH Zurich, Universitätstr. 16, 8092 Zurich, Switzerland
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Lynsey R. Harper
6Illinois Natural History Survey, Prairie Research Institute, University of Illinois at Urbana-Champaign, Champaign, IL, USA
7School of Biological and Environmental Sciences, Liverpool John Moores University, Liverpool, L3 3AF, UK
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Helen C. Rees
8RSK ADAS Ltd, School of Veterinary Medicine and Science, The University of Nottingham, Sutton Bonington Campus, Loughborough, LE12 5RD, UK
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Rosetta C. Blackman
9Eawag: Swiss Federal Institute of Aquatic Science and Technology, Department of Aquatic Ecology, Überlandstr. 133, 8600 Dübendorf, Switzerland
10Department of Evolutionary Biology and Environmental Studies, University of Zurich, Winterthurerstr. 190, CH-8057 Zurich, Switzerland
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Daniela Sint
3Department of Zoology, University of Innsbruck, Technikerstr. 25, 6020, Innsbruck, Austria
11Sinsoma GmbH, Lannes 6, 6176 Völs, Austria
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Michael Traugott
3Department of Zoology, University of Innsbruck, Technikerstr. 25, 6020, Innsbruck, Austria
11Sinsoma GmbH, Lannes 6, 6176 Völs, Austria
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Caren S. Goldberg
12School of the Environment, Washington State University, Pullman, WA 99164, USA
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Kat Bruce
13Nature Metrics Ltd, CABI Site, Bakeham Lane, Egham, TW20 9TY, UK
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Abstract

The use of environmental DNA (eDNA) analysis for species monitoring requires rigorous validation - from field sampling to the analysis of PCR-based results - for meaningful application and interpretation. Assays targeting eDNA released by individual species are typically validated with no predefined criteria to answer specific research questions in one ecosystem. Hence, the general applicability of assays as well as associated uncertainties and limitations, often remain undetermined. The absence of clear guidelines for assay validation prevents targeted eDNA assays from being incorporated into species monitoring and policy; thus, their establishment is essential for realizing the potential of eDNA-based surveys. We describe the measures and tests necessary for successful validation of targeted eDNA assays and the associated pitfalls to form the basis of guidelines. A list of 122 variables was compiled, consolidated into 14 thematic blocks, (e.g. “in silico analysis”), and arranged on a 5-level validation scale from “incomplete” to “operational” with defined minimum validation criteria for each level. These variables were evaluated for 546 published single-species assays. The resulting dataset was used to provide an overview of current validation practices and test the applicability of the validation scale for future assay rating. Of the 122 variables, 20% to 76% were reported; the majority (30%) of investigated assays were classified as Level 1 (incomplete), and 15% did not achieve this first level. These assays were characterised by minimal in silico and in vitro testing, but their share in annually published eDNA assays has declined since 2014. The meta-analysis demonstrates the suitability of the 5-level validation scale for assessing targeted eDNA assays. It is a user-friendly tool to evaluate previously published assays for future research and routine monitoring, while also enabling the appropriate interpretation of results. Finally, it provides guidance on validation and reporting standards for newly developed assays.

Competing Interest Statement

KB is the co-founder and CEO of Nature Metrics Ltd. a for profit company dedicated to the analysis of environmental DNA. HCR manages environmental DNA services for RSK ADAS Ltd. a for profit environmental consultancy. DS and MT are co-founders of Sinsoma GmbH. a for profit company dedicated to the analysis of environmental DNA.

Footnotes

  • https://doi.org/10.6084/m9.figshare.12184860.v1

  • https://edna-validation.com/

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted February 03, 2021.
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A validation scale to determine the readiness of environmental DNA assays for routine species monitoring
Bettina Thalinger, Kristy Deiner, Lynsey R. Harper, Helen C. Rees, Rosetta C. Blackman, Daniela Sint, Michael Traugott, Caren S. Goldberg, Kat Bruce
bioRxiv 2020.04.27.063990; doi: https://doi.org/10.1101/2020.04.27.063990
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A validation scale to determine the readiness of environmental DNA assays for routine species monitoring
Bettina Thalinger, Kristy Deiner, Lynsey R. Harper, Helen C. Rees, Rosetta C. Blackman, Daniela Sint, Michael Traugott, Caren S. Goldberg, Kat Bruce
bioRxiv 2020.04.27.063990; doi: https://doi.org/10.1101/2020.04.27.063990

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