Development of an environmental DNA metabarcoding assay for aquatic vascular plant communities

Abstract

Environmental DNA (eDNA) metabarcodes allow for the simultaneous detection of multiple taxa if the barcode regions meet several key requirements including conserved primer‐binding sites, interspecific variability that exceeds intraspecific variability, and relatively short amplicons. Currently, there are no established metabarcoding assays for aquatic vascular plants, which could limit biodiversity assessments and the early detection of alien species. We used a combination of novel and pre‐existing primers to generate eDNA metabarcodes from three gene regions that are commonly used for plant barcoding: two regions of chloroplast DNA (rbcL and matK) plus a segment of an internal transcribed spacer (ITS2). We optimized the assay on a mock community of 25 known species and then applied it to wild samples collected from two waterbodies in southern Ontario, Canada (Black River and Seymour Lake). Our novel rbcL primers, which amplify a fragment of ~220 bp, provided the most comprehensive description of the mock community, identifying all but one of the taxa to species or genus. In addition, after pooling data from five sites within each sampled waterbody, metabarcodes generated by this same primer pair identified more taxa than all other primer pairs; more specifically, 20 and 26 taxa were identified from Black River and Seymour Lake, respectively, to species or genus. Across the two sites, nine of the identified taxa are alien invasive aquatic plants. Five of these invasive species have no previous reports from our sites, and in some cases have no known established Ontario populations; our data therefore suggest an urgent need to increase surveillance for these aliens. Overall, our study showed that eDNA metabarcoding with a novel rbcL primer pair provides a cost‐effective method for simultaneously detecting multiple aquatic vascular plant taxa and is a valuable tool for the early detection of invasive species.

Detection of Endangered Aquatic Plants in Rapid Streams Using Environmental DNA

“…In fact, Potamogeton crispus, a common submerged plant, was detected at three sites, although it could not be found visually during the field sampling. Distribution survey is essential for the conservation of rare aquatic plants, however, it is difficult to find them because of their submerged nature and aquatic habitats are often hard to approach (Kuzmina et al, 2018;Coghlan et al, 2020). eDNA metabarcoding using a short marker will quickly inform on the distribution of rare aquatic plants and provide useful information for their conservation.…”

“…Recently, the environmental DNA (eDNA) survey was developed as an efficient and sensitive approach to determine the distribution of rare aquatic species (Thomsen and Willerslev, 2015). Several studies applied the eDNA survey to aquatic plants and successfully detected the target species among collected samples in aquariums, rivers, and ponds (Scriver et al, 2015;Fujiwara et al, 2016;Matsuhashi et al, 2016;Gantz et al, 2018;Kuzmina et al, 2018;Anglès d'Auriac et al, 2019;Chase et al, 2020;Coghlan et al, 2020;Doi et al, 2020;Kuehne et al, 2020;Miyazono et al, 2020). Some of these works report the detection of some populations that had not been recorded using conventional observation (Matsuhashi et al, 2016;Kuzmina et al, 2018;Miyazono et al, 2020).…”

Airborne Environmental DNA Metabarcoding Detects More Diversity, More Efficiently, Than a Traditional Plant Community Survey

“…Finally, ASVs with 10 sequences or fewer were removed from our future BLAST analysis to limit sequencing and assignment errors (40). Taxonomic assignment for the remaining unique ASVs was completed using BLASTn and the NCBI nt database.…”

Looking for Hidden Enemies of Metabarcoding: Species Composition, Habitat and Management Can Strongly Influence DNA Extraction while Examining Grassland Communities

“…Total DNA extracted from the living belowground plant tissues can considerably improve our understanding of the community organization, dynamics and diversity [ 6 , 19 ]. A growing number of biologists are using metabarcoding DNA for plants detection in a given environmental sample [ 24 , 25 , 26 ]. The internal transcribed spacer (ITS2 ) subunit region has been often employed for plant metabarcoding as it has potentially high resolution at the genus and species level [ 26 , 27 ].…”