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Needle in a haystack? A comparison of eDNA metabarcoding and targeted qPCR for detection of the great crested newt (Triturus cristatus)

Harper, Lynsey R.; Lawson Handley, Lori; Hahn, Christoph; Boonham, Neil; Rees, Helen C.; Gough, Kevin C.; Lewis, Erin; Adams, Ian P.; Brotherton, Peter; Phillips, Susanna; Hänfling, Bernd


Lynsey R. Harper

Christoph Hahn

Neil Boonham

Helen C. Rees

Kevin C. Gough

Erin Lewis

Ian P. Adams

Peter Brotherton

Susanna Phillips

Bernd Hänfling


Bernd Haenfling


Environmental DNA (eDNA) analysis is a rapid, cost‐effective, non‐invasive biodiversity monitoring tool which utilises DNA left behind in the environment by organisms for species detection. The method is used as a species‐specific survey tool for rare or invasive species across a broad range of ecosystems. Recently, eDNA and “metabarcoding” have been combined to describe whole communities rather than focusing on single target species. However, whether metabarcoding is as sensitive as targeted approaches for rare species detection remains to be evaluated. The great crested newt Triturus cristatus is a flagship pond species of international conservation concern and the first UK species to be routinely monitored using eDNA. We evaluate whether eDNA metabarcoding has comparable sensitivity to targeted real‐time quantitative PCR (qPCR) for T. cristatus detection. Extracted eDNA samples (N = 532) were screened for T. cristatus by qPCR and analysed for all vertebrate species using high‐throughput sequencing technology. With qPCR and a detection threshold of 1 of 12 positive qPCR replicates, newts were detected in 50% of ponds. Detection decreased to 32% when the threshold was increased to 4 of 12 positive qPCR replicates. With metabarcoding, newts were detected in 34% of ponds without a detection threshold, and in 28% of ponds when a threshold (0.028%) was applied. Therefore, qPCR provided greater detection than metabarcoding but metabarcoding detection with no threshold was equivalent to qPCR with a stringent detection threshold. The proportion of T. cristatus sequences in each sample was positively associated with the number of positive qPCR replicates (qPCR score) suggesting eDNA metabarcoding may be indicative of eDNA concentration. eDNA metabarcoding holds enormous potential for holistic biodiversity assessment and routine freshwater monitoring. We advocate this community approach to freshwater monitoring to guide management and conservation, whereby entire communities can be initially surveyed to best inform use of funding and time for species‐specific surveys.


Harper, L. R., Lawson Handley, L., Hahn, C., Boonham, N., Rees, H. C., Gough, K. C., …Hänfling, B. (2018). Needle in a haystack? A comparison of eDNA metabarcoding and targeted qPCR for detection of the great crested newt (Triturus cristatus). Ecology and Evolution, 8(12), 6330-6341.

Acceptance Date Feb 9, 2018
Online Publication Date May 29, 2018
Publication Date 2018-06
Deposit Date Jul 5, 2018
Publicly Available Date Jul 6, 2018
Journal Ecology and Evolution
Electronic ISSN 2045-7758
Publisher Wiley
Peer Reviewed Peer Reviewed
Volume 8
Issue 12
Pages 6330-6341
Keywords Ecology; Ecology, Evolution, Behavior and Systematics; Nature and Landscape Conservation
Public URL
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Article (856 Kb)

Copyright Statement
© 2018 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd.

This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.

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