Species-level biodiversity assessment using marine environmental DNA metabarcoding requires protocol optimization and standardization
DNA extraction from environmental samples (environmental DNA; eDNA) for metabarcoding-based biodiversity studies is gaining popularity as a noninvasive, time-efficient, and cost-effective monitoring tool. The potential benefits are promising for marine conservation, as the marine biome is frequently...
| Main Authors: | , , , , , , , |
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| Format: | Journal Article |
| Language: | English |
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WILEY
2019
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| Online Access: | http://purl.org/au-research/grants/arc/LP160101508 http://hdl.handle.net/20.500.11937/90909 |
| _version_ | 1848765457880842240 |
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| author | Jeunen, G.J. Knapp, M. Spencer, H.G. Taylor, H.R. Lamare, M.D. Stat, Michael Bunce, Michael Gemmell, N.J. |
| author_facet | Jeunen, G.J. Knapp, M. Spencer, H.G. Taylor, H.R. Lamare, M.D. Stat, Michael Bunce, Michael Gemmell, N.J. |
| author_sort | Jeunen, G.J. |
| building | Curtin Institutional Repository |
| collection | Online Access |
| description | DNA extraction from environmental samples (environmental DNA; eDNA) for metabarcoding-based biodiversity studies is gaining popularity as a noninvasive, time-efficient, and cost-effective monitoring tool. The potential benefits are promising for marine conservation, as the marine biome is frequently under-surveyed due to its inaccessibility and the consequent high costs involved. With increasing numbers of eDNA-related publications have come a wide array of capture and extraction methods. Without visual species confirmation, inconsistent use of laboratory protocols hinders comparability between studies because the efficiency of target DNA isolation may vary. We determined an optimal protocol (capture and extraction) for marine eDNA research based on total DNA yield measurements by comparing commonly employed methods of seawater filtering and DNA isolation. We compared metabarcoding results of both targeted (small taxonomic group with species-level assignment) and universal (broad taxonomic group with genus/family-level assignment) approaches obtained from replicates treated with the optimal and a low-performance capture and extraction protocol to determine the impact of protocol choice and DNA yield on biodiversity detection. Filtration through cellulose-nitrate membranes and extraction with Qiagen's DNeasy Blood & Tissue Kit outperformed other combinations of capture and extraction methods, showing a ninefold improvement in DNA yield over the poorest performing methods. Use of optimized protocols resulted in a significant increase in OTU and species richness for targeted metabarcoding assays. However, changing protocols made little difference to the OTU and taxon richness obtained using universal metabarcoding assays. Our results demonstrate an increased risk of false-negative species detection for targeted eDNA approaches when protocols with poor DNA isolation efficacy are employed. Appropriate optimization is therefore essential for eDNA monitoring to remain a powerful, efficient, and relatively cheap method for biodiversity assessments. For seawater, we advocate filtration through cellulose-nitrate membranes and extraction with Qiagen's DNeasy Blood & Tissue Kit or phenol-chloroform-isoamyl for successful implementation of eDNA multi-marker metabarcoding surveys. |
| first_indexed | 2025-11-14T11:35:34Z |
| format | Journal Article |
| id | curtin-20.500.11937-90909 |
| institution | Curtin University Malaysia |
| institution_category | Local University |
| language | English |
| last_indexed | 2025-11-14T11:35:34Z |
| publishDate | 2019 |
| publisher | WILEY |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | curtin-20.500.11937-909092023-05-01T06:15:19Z Species-level biodiversity assessment using marine environmental DNA metabarcoding requires protocol optimization and standardization Jeunen, G.J. Knapp, M. Spencer, H.G. Taylor, H.R. Lamare, M.D. Stat, Michael Bunce, Michael Gemmell, N.J. Science & Technology Life Sciences & Biomedicine Ecology Evolutionary Biology Environmental Sciences & Ecology biodiversity assessment eDNA extraction filtration metabarcoding EXTRACTION PCR QUANTIFICATION CAPTURE CONSERVATION PRIMERS TOOL biodiversity assessment eDNA extraction filtration metabarcoding DNA extraction from environmental samples (environmental DNA; eDNA) for metabarcoding-based biodiversity studies is gaining popularity as a noninvasive, time-efficient, and cost-effective monitoring tool. The potential benefits are promising for marine conservation, as the marine biome is frequently under-surveyed due to its inaccessibility and the consequent high costs involved. With increasing numbers of eDNA-related publications have come a wide array of capture and extraction methods. Without visual species confirmation, inconsistent use of laboratory protocols hinders comparability between studies because the efficiency of target DNA isolation may vary. We determined an optimal protocol (capture and extraction) for marine eDNA research based on total DNA yield measurements by comparing commonly employed methods of seawater filtering and DNA isolation. We compared metabarcoding results of both targeted (small taxonomic group with species-level assignment) and universal (broad taxonomic group with genus/family-level assignment) approaches obtained from replicates treated with the optimal and a low-performance capture and extraction protocol to determine the impact of protocol choice and DNA yield on biodiversity detection. Filtration through cellulose-nitrate membranes and extraction with Qiagen's DNeasy Blood & Tissue Kit outperformed other combinations of capture and extraction methods, showing a ninefold improvement in DNA yield over the poorest performing methods. Use of optimized protocols resulted in a significant increase in OTU and species richness for targeted metabarcoding assays. However, changing protocols made little difference to the OTU and taxon richness obtained using universal metabarcoding assays. Our results demonstrate an increased risk of false-negative species detection for targeted eDNA approaches when protocols with poor DNA isolation efficacy are employed. Appropriate optimization is therefore essential for eDNA monitoring to remain a powerful, efficient, and relatively cheap method for biodiversity assessments. For seawater, we advocate filtration through cellulose-nitrate membranes and extraction with Qiagen's DNeasy Blood & Tissue Kit or phenol-chloroform-isoamyl for successful implementation of eDNA multi-marker metabarcoding surveys. 2019 Journal Article http://hdl.handle.net/20.500.11937/90909 10.1002/ece3.4843 English http://purl.org/au-research/grants/arc/LP160101508 http://purl.org/au-research/grants/arc/LP160100839 http://creativecommons.org/licenses/by/4.0/ WILEY fulltext |
| spellingShingle | Science & Technology Life Sciences & Biomedicine Ecology Evolutionary Biology Environmental Sciences & Ecology biodiversity assessment eDNA extraction filtration metabarcoding EXTRACTION PCR QUANTIFICATION CAPTURE CONSERVATION PRIMERS TOOL biodiversity assessment eDNA extraction filtration metabarcoding Jeunen, G.J. Knapp, M. Spencer, H.G. Taylor, H.R. Lamare, M.D. Stat, Michael Bunce, Michael Gemmell, N.J. Species-level biodiversity assessment using marine environmental DNA metabarcoding requires protocol optimization and standardization |
| title | Species-level biodiversity assessment using marine environmental DNA metabarcoding requires protocol optimization and standardization |
| title_full | Species-level biodiversity assessment using marine environmental DNA metabarcoding requires protocol optimization and standardization |
| title_fullStr | Species-level biodiversity assessment using marine environmental DNA metabarcoding requires protocol optimization and standardization |
| title_full_unstemmed | Species-level biodiversity assessment using marine environmental DNA metabarcoding requires protocol optimization and standardization |
| title_short | Species-level biodiversity assessment using marine environmental DNA metabarcoding requires protocol optimization and standardization |
| title_sort | species-level biodiversity assessment using marine environmental dna metabarcoding requires protocol optimization and standardization |
| topic | Science & Technology Life Sciences & Biomedicine Ecology Evolutionary Biology Environmental Sciences & Ecology biodiversity assessment eDNA extraction filtration metabarcoding EXTRACTION PCR QUANTIFICATION CAPTURE CONSERVATION PRIMERS TOOL biodiversity assessment eDNA extraction filtration metabarcoding |
| url | http://purl.org/au-research/grants/arc/LP160101508 http://purl.org/au-research/grants/arc/LP160101508 http://hdl.handle.net/20.500.11937/90909 |