TY - JOUR

T1 - Detection of environmental DNA from amphibians in Northern Europe applied in citizen science

AU - Knudsen, Steen Wilhelm

AU - Hesselsøe, Martin

AU - Rytter, Maria

AU - Lillemark, Marie Rathcke

AU - Tøttrup, Anders P.

AU - Rahbek, Carsten

AU - Sheard, Julie Koch

AU - Thomsen, Philip Francis

AU - Agersnap, Sune

AU - Mortensen, Peter B.

AU - Møller, Peter Rask

N1 - Publisher Copyright: © 2023 The Authors. Environmental DNA published by John Wiley & Sons Ltd.

PY - 2023

Y1 - 2023

N2 - Many species of amphibians in Northern Europe are threatened and the local distributions are rarely described in detail. Application of modern molecular methods provides an important supplementary tool for monitoring the distribution and diversity of amphibians. For this purpose, we designed, tested, validated, and optimized 14 species-specific assays on genomic DNA extracted from tissue samples to use for quantitative polymerase chain reaction (qPCR) setups targeting mitochondrial DNA from amphibians in freshwater samples. The tests confirmed species specificity for all assays. Considering a systematic definition of the limit of detection for each of the assays, the presented qPCR assays are unlikely to return false positive detection from any co-occurring species in northern Europe. For field validation, the qPCR assays were applied in a large-scale nationwide citizen science project in which sampling and qPCR analysis was carried out by high school students. Data from the citizen science project returned the expected results when compared to the known regional distribution of the target species and confirmed the presence of nine out of 14 Danish species of amphibians in the collected freshwater samples. Four out of 2550 qPCR test sets carried out by the high school students required a professional reanalysis in multiple replicates due to initial unexpected results. This emphasizes that efforts from citizen science may generate large amounts of valuable data, as long as the results are carefully scrutinized by experts.

AB - Many species of amphibians in Northern Europe are threatened and the local distributions are rarely described in detail. Application of modern molecular methods provides an important supplementary tool for monitoring the distribution and diversity of amphibians. For this purpose, we designed, tested, validated, and optimized 14 species-specific assays on genomic DNA extracted from tissue samples to use for quantitative polymerase chain reaction (qPCR) setups targeting mitochondrial DNA from amphibians in freshwater samples. The tests confirmed species specificity for all assays. Considering a systematic definition of the limit of detection for each of the assays, the presented qPCR assays are unlikely to return false positive detection from any co-occurring species in northern Europe. For field validation, the qPCR assays were applied in a large-scale nationwide citizen science project in which sampling and qPCR analysis was carried out by high school students. Data from the citizen science project returned the expected results when compared to the known regional distribution of the target species and confirmed the presence of nine out of 14 Danish species of amphibians in the collected freshwater samples. Four out of 2550 qPCR test sets carried out by the high school students required a professional reanalysis in multiple replicates due to initial unexpected results. This emphasizes that efforts from citizen science may generate large amounts of valuable data, as long as the results are carefully scrutinized by experts.

KW - Amphibia, Anura

KW - eDNA

KW - qPCR

KW - quantitative PCR

KW - real-time polymerase chain reaction

U2 - 10.1002/edn3.462

DO - 10.1002/edn3.462

M3 - Journal article

AN - SCOPUS:85168884104

VL - 5

SP - 1429

EP - 1448

JO - Environmental DNA

JF - Environmental DNA

SN - 2637-4943

IS - 6

ER -