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Optimising the detection of marine taxonomic richness using environmental DNA metabarcoding: the effects of filter material, pore size and extraction method

Optimising the detection of marine taxonomic richness using environmental DNA metabarcoding: the effects of filter material, pore size and extraction method
Optimising the detection of marine taxonomic richness using environmental DNA metabarcoding: the effects of filter material, pore size and extraction method
The analysis of environmental DNA (eDNA) using metabarcoding has increased in use as a method for tracking biodiversity of ecosystems. Little is known about eDNA in marine human-modified environments, such as commercial ports, which are key sites to monitor for anthropogenic impacts on coastal ecosystems. To optimise an eDNA metabarcoding protocol in these environments, seawater samples were collected in a commercial port and methodologies for concentrating and purifying eDNA were tested for their effect on eukaryotic DNA yield and subsequent richness of Operational Taxonomic Units (OTUs). Different filter materials [Cellulose Nitrate (CN) and Glass Fibre (GF)], with different pore sizes (0.5 µm, 0.7 µm and 1.2 µm) and three previously published liquid phase extraction methods were tested. The number of eukaryotic OTUs detected differed by a factor of three amongst the method combinations. The combination of CN filters with phenol-chloroform-isoamyl alcohol extractions recovered a higher amount of eukaryotic DNA and OTUs compared to GF filters and the chloroform-isoamyl alcohol extraction method. Pore size was not independent of filter material but did affect the yield of eukaryotic DNA. For the OTUs assigned to a highly successful non-indigenous species, Styela clava, the two extraction methods with phenol significantly outperformed the extraction method without phenol; other experimental treatments did not contribute significantly to detection. These results highlight that careful consideration of methods is warranted because choice of filter material and extraction method create false negative detections of marine eukaryotic OTUs and underestimate taxonomic richness from environmental samples.
Deiner, Kristy
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Lopez, Jacqueline
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Bourne, Steve
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Holman, Luke, Earl
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Seymour, Mathew
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Grey, Erin K.
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Lacoursière, Anaïs
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Li, Yiyuan
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Renshaw, Mark A.
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Pfrender, Michael E.
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Rius, Marc
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Bernatchez, Louis
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Lodge, David M.
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Deiner, Kristy
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Lopez, Jacqueline
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Bourne, Steve
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Holman, Luke, Earl
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Seymour, Mathew
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Grey, Erin K.
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Lacoursière, Anaïs
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Li, Yiyuan
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Renshaw, Mark A.
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Pfrender, Michael E.
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Rius, Marc
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Bernatchez, Louis
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Lodge, David M.
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Deiner, Kristy, Lopez, Jacqueline, Bourne, Steve, Holman, Luke, Earl, Seymour, Mathew, Grey, Erin K., Lacoursière, Anaïs, Li, Yiyuan, Renshaw, Mark A., Pfrender, Michael E., Rius, Marc, Bernatchez, Louis and Lodge, David M. (2018) Optimising the detection of marine taxonomic richness using environmental DNA metabarcoding: the effects of filter material, pore size and extraction method. Metabarcoding and Metagenomics, 2 (e28963). (doi:10.3897/mbmg.2.28963).

Record type: Article

Abstract

The analysis of environmental DNA (eDNA) using metabarcoding has increased in use as a method for tracking biodiversity of ecosystems. Little is known about eDNA in marine human-modified environments, such as commercial ports, which are key sites to monitor for anthropogenic impacts on coastal ecosystems. To optimise an eDNA metabarcoding protocol in these environments, seawater samples were collected in a commercial port and methodologies for concentrating and purifying eDNA were tested for their effect on eukaryotic DNA yield and subsequent richness of Operational Taxonomic Units (OTUs). Different filter materials [Cellulose Nitrate (CN) and Glass Fibre (GF)], with different pore sizes (0.5 µm, 0.7 µm and 1.2 µm) and three previously published liquid phase extraction methods were tested. The number of eukaryotic OTUs detected differed by a factor of three amongst the method combinations. The combination of CN filters with phenol-chloroform-isoamyl alcohol extractions recovered a higher amount of eukaryotic DNA and OTUs compared to GF filters and the chloroform-isoamyl alcohol extraction method. Pore size was not independent of filter material but did affect the yield of eukaryotic DNA. For the OTUs assigned to a highly successful non-indigenous species, Styela clava, the two extraction methods with phenol significantly outperformed the extraction method without phenol; other experimental treatments did not contribute significantly to detection. These results highlight that careful consideration of methods is warranted because choice of filter material and extraction method create false negative detections of marine eukaryotic OTUs and underestimate taxonomic richness from environmental samples.

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e-pub ahead of print date: 2 November 2018
Published date: 2 November 2018

Identifiers

Local EPrints ID: 425947
URI: http://eprints.soton.ac.uk/id/eprint/425947
PURE UUID: ea373c60-ff26-443f-ad78-e25061b8ac42
ORCID for Luke, Earl Holman: ORCID iD orcid.org/0000-0002-8139-3760

Catalogue record

Date deposited: 07 Nov 2018 17:30
Last modified: 26 Nov 2021 02:32

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Contributors

Author: Kristy Deiner
Author: Jacqueline Lopez
Author: Steve Bourne
Author: Luke, Earl Holman ORCID iD
Author: Mathew Seymour
Author: Erin K. Grey
Author: Anaïs Lacoursière
Author: Yiyuan Li
Author: Mark A. Renshaw
Author: Michael E. Pfrender
Author: Marc Rius
Author: Louis Bernatchez
Author: David M. Lodge

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