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

By

Deiner, Kristy
Lopez, Jacqueline
Bourne, S. J. (Steve J.)
Holman, Luke
Seymour, Mathew
Grey, Erin K.
Lacoursière, Anaïs
Li, Yiyuan
Renshaw, Mark A.
Pfrender, Michael E.
Rius, Marc
Bernatchez, Louis
Lodge, David M.

Type

Article

Date of Publication

2018-11-2

Original Publication

Metabarcoding and metagenomics

Volume

2

Pages

1--15

Subjects

18S ribosomal , Edna , high-throughput-sequencing , metazoan eukaryotes , non-indigenous species , Seawater

Contributed by

Pensoft Publishers

Language

English

Published by

Bulgaria Pensoft Publishers 2017-

Identifiers

DOI: https://doi.org/10.3897/mbmg.2.28963

 

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, Styelaclava, 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.