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Environmental DNA metabarcoding for freshwater bivalves biodiversity assessment: methods and results for the Western Palearctic (European sub-region)

Abstract

Freshwater ecosystems are the most vulnerable worldwide and freshwater bivalves rank amongst the most threatened animals in the world. Surveying and monitoring freshwater bivalves are difficult tasks: they are difficult to find, hard to identify (taxonomic expertise is needed), and working underwater is technically challenging. It is therefore crucial to find more efficient methods to survey and monitor these species. Here, we present the first metabarcoding approach for freshwater bivalves and compare environmental DNA (eDNA) and traditional surveys. We describe two sets of primers (for Unionida and Venerida) developed for freshwater bivalves eDNA metabarcoding. These primers have been tested in the field, with about 300 studied sites. Results were compared to freshwater bivalves’ surveys using traditional methods, with eDNA always detecting more species than traditional surveys, especially when Sphaerids were taken into account. While our study initially focused on Western Palearctic freshwater bivalve species, our primers were confronted in silico with available sequences and have proven to be effective at a global scale. The results show that eDNA metabarcoding, with our developed primers, is a remarkable tool allowing for non-invasive surveys, detection of rare and inconspicuous species, absence data and overall freshwater bivalves routine monitoring.

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Acknowledgements

The development of the eDNA analysis method for bivalves was carried out jointly by Caracol NGO, Spygen company and Biotope consultancy. The Agence Française pour la Biodiversité and the Direction Régionale de l’Environnement, de l’Aménagement et du Logement d’Occitanie contributed to its financing. The data presented here include work carried out by Biotope and Aquascop consultancies, Caracol NGO, SpyGen, the Compagnie Nationale du Rhône, the Regional Natural Park of Limousin, the Conservatoires d'Espaces Naturels Midi-Pyrénées and Nouvelle Aquitaine, the LIFE + Giant Freshwater Mussel program, GRT Gaz, Voies Navigables de France, EPTB Vilaine, CPIE Loire-Anjou for the main studies. The DREAL Grand Est, Nouvelle Aquitaine, Occitanie and the DDT du Tarn also financed part of the studies. We would also like to thank all those who participated in the sampling and laboratory analysis of the samples. This research was also developed under ConBiomics: the missing approach for the Conservation of Freshwater Bivalves Project Nº NORTE-01-0145-FEDER-030286, co-financed by COMPETE 2020, Portugal 2020 and the European Union through the ERDF, and by Portuguese Foundation for Science and Technology (FCT) through national funds.

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Online Resource1: Western Palearctic 16S fragment reference database. Supplementary file1 (XLSX 27 kb)

10750_2020_4260_MOESM2_ESM.xlsx

Online Resource 2: In silico PCR (Ficetola et al. 2010) ran on a collection of all mitochondrial DNA sequences, and all available DNA sequences in release 138 (standard sequences) of the EMBL database. Supplementary file2 (XLSX 126 kb)

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Prié, V., Valentini, A., Lopes-Lima, M. et al. Environmental DNA metabarcoding for freshwater bivalves biodiversity assessment: methods and results for the Western Palearctic (European sub-region). Hydrobiologia 848, 2931–2950 (2021). https://doi.org/10.1007/s10750-020-04260-8

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  • DOI: https://doi.org/10.1007/s10750-020-04260-8

Keywords

  • eDNA
  • Non-invasive survey
  • Metabarcoding primers
  • Freshwater bivalve survey
  • Invasive species monitoring