Single metabarcoding multiplex captures community‐level freshwater biodiversity and beyond
Abstract Cost‐effective and accurate quantification of biodiversity is important for biodiversity conservation, resource management, and forecasting. Traditional monitoring approaches have relied on direct observations, remote sensing, and mark‐recapture techniques, providing insights into species e...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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Wiley
2024-01-01
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| Series: | Environmental DNA |
| Subjects: | |
| Online Access: | https://doi.org/10.1002/edn3.515 |
| _version_ | 1797277052989079552 |
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| author | Niamh Eastwood Stephen Kissane Lewis Campbell Andrew G. Briscoe Bastian Egeter Luisa Orsini |
| author_facet | Niamh Eastwood Stephen Kissane Lewis Campbell Andrew G. Briscoe Bastian Egeter Luisa Orsini |
| author_sort | Niamh Eastwood |
| collection | DOAJ |
| description | Abstract Cost‐effective and accurate quantification of biodiversity is important for biodiversity conservation, resource management, and forecasting. Traditional monitoring approaches have relied on direct observations, remote sensing, and mark‐recapture techniques, providing insights into species ecology and the impact of pollution and climate change on indicator species. However, these techniques are typically low throughput, expensive, and can be invasive. In addition, they cannot detect cryptic diversity and are biased toward species that leave identifiable remains. DNA‐based methods, such as metabarcoding or single marker gene assays, have enabled high throughput screening of a wide range of taxonomic groups, including ones without well‐preserved remains. When compared with traditional techniques, these approaches have high throughput, can resolve cryptic diversity, do not require taxonomic specialist skills, and are non‐invasive. However, although they are comparatively cheaper than traditional approaches, they are expensive when applied at the community‐level as single marker assays are amplified and sequenced independently. Multilocus approaches in which multiple gene markers are amplified in a single reaction are desirable to deliver community‐level assessments in a cost‐effective manner. Yet, they are uncommon because of technical challenges that may lead to biases in downstream analyses, such as index hopping and unbalanced representation of taxonomic groups. Here, we developed a highly multiplexed protocol that combines the early pooling of marker genes that target broad taxonomic groups and taxon‐specific markers in a single tube reaction. This step is followed by the pooling of up to 384 samples per locus (N = 15,636 samples) with unique dual‐indexed sequencing adapters in a single sequencing run. This approach dramatically reduces the costs of community‐level biodiversity quantification and lowers the need for input DNA without compromising output quality. We optimized the multiplex assay on lake freshwater sediment samples and benchmarked the assay on samples from other environmental matrices, demonstrating its direct application to the river and marine communities. |
| first_indexed | 2024-03-07T15:38:07Z |
| format | Article |
| id | doaj.art-e9f76cebda8145b095a05b3ba5634bde |
| institution | Directory Open Access Journal |
| issn | 2637-4943 |
| language | English |
| last_indexed | 2024-03-07T15:38:07Z |
| publishDate | 2024-01-01 |
| publisher | Wiley |
| record_format | Article |
| series | Environmental DNA |
| spelling | doaj.art-e9f76cebda8145b095a05b3ba5634bde2024-03-05T09:32:37ZengWileyEnvironmental DNA2637-49432024-01-0161n/an/a10.1002/edn3.515Single metabarcoding multiplex captures community‐level freshwater biodiversity and beyondNiamh Eastwood0Stephen Kissane1Lewis Campbell2Andrew G. Briscoe3Bastian Egeter4Luisa Orsini5Environmental Genomics Group, School of Biosciences University of Birmingham Birmingham UKEnvironmental Genomics Group, School of Biosciences University of Birmingham Birmingham UKNatureMetrics Ltd. Surrey Research Park Guildford UKNatureMetrics Ltd. Surrey Research Park Guildford UKNatureMetrics Ltd. Surrey Research Park Guildford UKEnvironmental Genomics Group, School of Biosciences University of Birmingham Birmingham UKAbstract Cost‐effective and accurate quantification of biodiversity is important for biodiversity conservation, resource management, and forecasting. Traditional monitoring approaches have relied on direct observations, remote sensing, and mark‐recapture techniques, providing insights into species ecology and the impact of pollution and climate change on indicator species. However, these techniques are typically low throughput, expensive, and can be invasive. In addition, they cannot detect cryptic diversity and are biased toward species that leave identifiable remains. DNA‐based methods, such as metabarcoding or single marker gene assays, have enabled high throughput screening of a wide range of taxonomic groups, including ones without well‐preserved remains. When compared with traditional techniques, these approaches have high throughput, can resolve cryptic diversity, do not require taxonomic specialist skills, and are non‐invasive. However, although they are comparatively cheaper than traditional approaches, they are expensive when applied at the community‐level as single marker assays are amplified and sequenced independently. Multilocus approaches in which multiple gene markers are amplified in a single reaction are desirable to deliver community‐level assessments in a cost‐effective manner. Yet, they are uncommon because of technical challenges that may lead to biases in downstream analyses, such as index hopping and unbalanced representation of taxonomic groups. Here, we developed a highly multiplexed protocol that combines the early pooling of marker genes that target broad taxonomic groups and taxon‐specific markers in a single tube reaction. This step is followed by the pooling of up to 384 samples per locus (N = 15,636 samples) with unique dual‐indexed sequencing adapters in a single sequencing run. This approach dramatically reduces the costs of community‐level biodiversity quantification and lowers the need for input DNA without compromising output quality. We optimized the multiplex assay on lake freshwater sediment samples and benchmarked the assay on samples from other environmental matrices, demonstrating its direct application to the river and marine communities.https://doi.org/10.1002/edn3.515eDNAfreshwatermarinemetabarcodingmultiplexsediment |
| spellingShingle | Niamh Eastwood Stephen Kissane Lewis Campbell Andrew G. Briscoe Bastian Egeter Luisa Orsini Single metabarcoding multiplex captures community‐level freshwater biodiversity and beyond Environmental DNA eDNA freshwater marine metabarcoding multiplex sediment |
| title | Single metabarcoding multiplex captures community‐level freshwater biodiversity and beyond |
| title_full | Single metabarcoding multiplex captures community‐level freshwater biodiversity and beyond |
| title_fullStr | Single metabarcoding multiplex captures community‐level freshwater biodiversity and beyond |
| title_full_unstemmed | Single metabarcoding multiplex captures community‐level freshwater biodiversity and beyond |
| title_short | Single metabarcoding multiplex captures community‐level freshwater biodiversity and beyond |
| title_sort | single metabarcoding multiplex captures community level freshwater biodiversity and beyond |
| topic | eDNA freshwater marine metabarcoding multiplex sediment |
| url | https://doi.org/10.1002/edn3.515 |
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