SeaDNA - Assessing marine biodiversity and structure using environmental DNA: from groundtruthing to food web structure and stability

Lead Research Organisation: University of Salford
Department Name: Sch of Environment & Life Sciences

Abstract

DNA evidence has revolutionised our understanding of the natural world. It has helped us to appreciate how species are related to one other, how environmental change can lead to species divergence and how individual populations become adapted through evolutionary processes to their local environments. It has also been particularly useful in quantifying the diversity of species in communities of microorganisms that cannot readily be seen and assessed using standard microscopy.

Importantly, DNA in the natural environment can also be used in a "forensic" manner. Traces of DNA from skin, blood, faeces or mucous can be used to identify which species have recently been present in the local environment. Given recent developments in DNA sequencing technology, this "environmental DNA" (eDNA) promises to revolutionise the way we probe biodiversity in our environment, particularly in marine environments that can be very difficult to sample reliably. Traditionally we have used specialist grabs and nets to survey species larger than microbes in marine communities. However, sampling free eDNA in surrounding water is potentially faster, less expensive and less destructive than such gears. Use of trace eDNA also holds potential to identify species that are not reliably sampled in the environment, either because they are rare, small, or adept at avoiding nets and grabs.

The utility of eDNA as a tool for sampling aquatic environments has been mostly tested in freshwater systems, and there are only a handful of studies that have tested the approach in the marine environment. Thus, there is a need to further evaluate the potential using a combination of laboratory experiments and field surveys. As an important first stage, we need to establish how long eDNA from fish and invertebrates persists in the marine environment before it is broken down beyond the point of detectability. This will tell us how well an eDNA-derived species list reflects the species community at the sampling site. We will conduct a set of laboratory experiments that will enable us to quantify the rate of eDNA break-down, and identify main environmental variables that influence this rate of decay. We will then aim to develop the laboratory and field methods needed to reliably detect DNA from these species groups, before testing these methods in experimental communities that we will assemble in laboratory aquaria.

An important stage in testing the ability of eDNA to be used as a tool in surveying and monitoring marine species is to survey the natural environment using both traditional methods (e.g. nets), and eDNA methods. We will do this in two UK marine habitats that are important for fisheries, conservation and environmental monitoring, namely estuaries and inshore shelf seas. We will also do this in an open ocean habitat, the Southern Ocean, which is an important habitat for fisheries and oceanic megafauna such as whales. We will directly compare data from eDNA methods to those from traditional methods to ask if eDNA accurately captures the fish and invertebrate communities, and if the method has the added ability to inform us on the presence of species that are typically rare or difficult to sample, some of which may be new to science.

Finally, we will use the eDNA derived species lists to reconstruct the food webs present in our sampling locations. We will use these data to test how stable marine communities are over space and time, and how environmental variables such as temperature affect their composition and stability. The results of these analyses will provide insight into the role of eDNA in helping us to understand how future climate change may affect fished species.

Planned Impact

The main beneficiaries will be:
1) Governmental Biodiversity/Conservation/Environmental Agencies & NGOs. The development of robust marine eDNA profiles could prove to be a powerful tool in biodiversity discovery, long-term monitoring and evaluation of the success of conservation and management initiatives. In addition to drawing on the wealth of contacts that the project team have with environmental agencies to seek knowledge exchange opportunities, we aim to use the existing Environment Agency-led "Environmental DNA network" as a forum for discussion and sharing of results.
2) Fisheries management organisations. Organisations such as DEFRA, CEFAS, Marine Scotland and the Marine Management Organisation have shown interest in scientific advances that can support enhancements to fisheries management and marine environmental monitoring. The project has strong potential to clarify the role that environmental DNA could have in the spatial and temporal mapping of target fished species, and the resilience of the food webs upon which they rely. We aim to share knowledge primarily through regular meeting exchanges, including the SEAFISH "Common Language Group" which includes representatives from NGOs, Fishermen Associations, Retailers, Consumer Groups and Scientists. This will ensure that a very broad base of stakeholders will be aware of the project tasks and achievements.
3) The General Public. We will play to the considerable interest that the general public have in marine life, and how new technologies can be used in species discovery, monitoring, conservation and exploitation. We will communicate specific research findings using institutional press offices while coordinating with the NERC communications team. We will use events such as National Science Week, the Manchester Science Festival and Bristol Festival of Nature to engage at the local level within our respective cities. We will generate and maintain a blog throughout the project, with an attached twitter account, which will give updates on project activities. All project researchers will contribute to the blog through short posts, photos, videos, news items and travel/meeting reports. We also aim to create a documentary, using existing links between project researchers and the science film making community, which will be accessible via websites of the project partners.

Publications

10 25 50
 
Description We have produced five publications, and several more are in preparation.
All initial results demonstrate that our goals will be achieved and in some cases surpassed.
- Data from UK estuaries show that eDNA can detect between two to three times more number of fish species than netting techniques and also detect spatial and seasonal patterns that would not be apparent using traditional techniques. It also shows that a great deal of additional important faunal information can be obtained from the same data, especially pertaining to seabirds and mammals of conservation importance.
- Data from the English Channel illustrate seasonal changes in fish assemblages and identify different habitat and life history related eDNA signatures. While the eDNA signatures correlate with trawl data for several species, we find that in others (such as sardines and plaice), eDNA peaks are associated with the spawning period, and hence gamete release. In other instances, differences in eDNA signatures between surface and bottom collections reflect the preferred habitat of the species (benthic/pelagic).
- Data from the Southern Ocean reveal dominance of different pelagic species at different depths, and for some of them an association with certain key planktonic invertebrates. In this case, also, eDNA data provide additional information on what can be termed 'molecular by-catch', by detecting several important species of marine mammals and sea birds. DNA metabarcoding of lanternfish stomach contents, paired with visual identification and stable isotope analysis, allow for a redefinition of the trophic niche of these important species, by showing a much more diverse food spectrum, and in particular a far greater importance of gelatinous carnivore zooplankton in the diet of lanternfishes.
- Further scientific papers have been produced since the last submission, and several more are in preparation, after the completion of the project.
Exploitation Route We are currently preparing several more publications to showcase all the project achievements.
We have also consistently engaged with the UK DNA Working Group (now represented under the UK EOF: http://www.ukeof.org.uk/our-work/ukdna) in order to translate evidence from 'SeaDNA' into practical solutions for marine environmental monitoring.
We are also currently engaging with stakeholders to initiate projects aimed at devising standard operating protocols for eDNA-based marine surveys.
Sectors Aerospace

Defence and Marine

Agriculture

Food and Drink

Digital/Communication/Information Technologies (including Software)

Environment

Leisure Activities

including Sports

Recreation and Tourism

Government

Democracy and Justice

Culture

Heritage

Museums and Collections

Pharmaceuticals and Medical Biotechnology

URL https://twitter.com/SeaDNAproject
 
Description 2022: Both the WWF Global Shark & Ray Initiative and the Save Our Seas Foundation have requested PI Stefano Mariani to give advisory seminars on the application of eDNA best practice for monitoring elasmobranch assemblages in different habitats and circumstances. The now launched project "Understanding the Ecological Diversity of the Chumash Heritage National Marine Sanctuary" (https://www.lenfestocean.org/en/research-projects/understanding-the-ecological-diversity-of-the-chumash-heritage-national-marine-sanctuary), under the Lenfest Ocean Program, has a strong socio-cultural dimension, aimed at embedding novel eDNA monitoring technologies with traditional indigenous knowledge, with knowledge transfer initiatives and the co-creation of blended conservation strategies, which stem from primary research findings and outcomes from the SeaDNA project. 2020: We have recently established a collaboration with Italian scientists, which is devoted to partnering with commercial trawlers as "ocean sentinels" and eDNA samplers, for assessing catch composition and the structure of the benthic community affected by trawling. We have consistently engaged with the UK DNA Working Group (now represented under the UK EOF: http://www.ukeof.org.uk/our-work/ukdna) in order to translate evidence from 'SeaDNA' into practical solutions for marine environmental monitoring. In particular, S. Mariani participated in a DNA-focused "marine benthic monitoring workshop" organised by the JNCC in Manchester, Feb 25-26. And also attended the UK DNA WG Steering Committee group at the NHM, London, March 11th. 2019: New developments: 1) Discussions with colleagues at Southampton, Bristol, NOCS, CEFAS and JNCC have led to the preparation of a Large Grant Outline bid (not successful). 2) Preliminary data generated through the collaboration with the NHM are leading to a new grant proposal (now awarded). 2016: The project is at n early phase, but meetings with CEFAS, Natural England, the Environment Agency, as well as a number of NGOs outside the UK, indicate that the project approaches and findings will have a significant impact on environmental monitoring activities and management planning potentially at a global scale. As a result of liaising with CEFAS, we developed a new project aimed at strengthening our understanding of efficacy of the eDNA approach to monitor and assess pelagic stocks, which was funded under the NERC-CASE initiative and will start in October 2018. We have also organised the 5th UK DNA Working Group Meeting in Salford (https://www.salford.ac.uk/events/event/2017/uk-dna-working-group), which catalysed further discussions and burgeoning collaborations between academics and stakeholders interested in putting eDNA approaches into practice.
First Year Of Impact 2022
Sector Agriculture, Food and Drink,Education,Environment,Leisure Activities, including Sports, Recreation and Tourism,Culture, Heritage, Museums and Collections
Impact Types Cultural

Societal

Policy & public services

 
Description Chief Scientific Advisers network dinner discussion at the Royal Society: Environmental DNA monitoring
Geographic Reach National 
Policy Influence Type Contribution to a national consultation/review
 
Description Consolidation and advancement of the knowledgevbase and transfer through the UK DNA Working Group
Geographic Reach National 
Policy Influence Type Membership of a guideline committee
URL https://www.ukeof.org.uk/our-work/ukdna
 
Description NERC CASE
Amount £89,114 (GBP)
Funding ID NE/R007861/1 
Organisation Natural Environment Research Council 
Sector Public
Country United Kingdom
Start 09/2018 
End 09/2022
 
Description SpongeDNA - Bolstering marine biodiversity exploration and monitoring through natural environmental DNA samplers
Amount £599,140 (GBP)
Funding ID NE/T007028/1 
Organisation Natural Environment Research Council 
Sector Public
Country United Kingdom
Start 06/2020 
End 06/2023
 
Title A generalised, dynamic DNA reference library for metabarcoding of fishes 
Description This reference sequence library and associated bioinformatic scripts offer a standardised, open-to-all, workflow to robustly identify North-East Atlantic and UK marine and freshwater fishes from DNA metabarcoding data. 
Type Of Material Improvements to research infrastructure 
Year Produced 2021 
Provided To Others? Yes  
Impact Reduction of uncertainty in taxonomic assignment of DNA sequences from environmental projects. Facilitation of computational analysis of environmental metabarcoding data sets. 
URL https://github.com/genner-lab/meta-fish-lib
 
Title Data from: All is fish that comes to the net: metabarcoding for rapid fisheries catch assessment 
Description AbstractMonitoring marine resource exploitation is a key activity in fisheries science and biodiversity conservation. Since research surveys are time-consuming and costly, fishery-dependent data (i.e. derived directly from fishing vessels) are increasingly credited with a key role in expanding the reach of ocean monitoring. Fishing vessels may be seen as widely ranging data-collecting platforms, which could act as a fleet of sentinels for monitoring marine life, in particular exploited stocks. Here, we investigate the possibility of assessing catch composition of single hauls carried out by trawlers by applying DNA metabarcoding to the "slush" collected from fishing nets just after the end of hauling operations. We assess the performance of this approach in portraying ß-diversity and examining the quantitative relationship between species abundances in the catch and DNA amount in the slush (reads counts generated by amplicon sequencing). We demonstrate that the assemblages identified using DNA in the slush mirror those returned by visual inspection of net content and detect a strong relationship between read counts and species abundances in the catch. We therefore argue that this approach could be upscaled to serve as a powerful source of information on the structure of demersal assemblages and the impact of fisheries. 
Type Of Material Database/Collection of data 
Year Produced 2020 
Provided To Others? Yes  
URL http://datadryad.org/stash/dataset/doi:10.5061/dryad.7m0cfxpsh
 
Title Data from: DNA metabarcoding unveils multi-scale trophic variation in a widespread coastal opportunist 
Description A thorough understanding of ecological networks relies on comprehensive information on trophic relationships among species. Since unpicking the diet of many organisms is unattainable using traditional morphology-based approaches, the application of high-throughput sequencing methods represents a rapid and powerful way forward. Here, we assessed the application of DNA-metabarcoding with nearly universal primers for the mitochondrial marker cytochrome c oxidase I (COI) in defining the trophic ecology of adult brown shrimp, Crangon crangon, in six European estuaries. The exact trophic role of this abundant and widespread coastal benthic species is somewhat controversial, while information on geographical variation remains scant. Results revealed a highly opportunistic behaviour. Shrimp stomach contents contained hundreds of taxa (>1000 molecular operational taxonomic units), of which 291 were identified as distinct species, belonging to 35 phyla. Only twenty ascertained species had a mean relative abundance of more than 0.5%. Predominant species included other abundant coastal and estuarine taxa, including the shore crab Carcinus maenas and the amphipod Corophium volutator. Jacobs' selectivity index estimates based on DNA extracted from both shrimp stomachs and sediment samples were used to assess the shrimp's trophic niche indicating a generalist diet, dominated by crustaceans, polychaetes and fish. Spatial variation in diet composition, at regional and local scales, confirmed the highly flexible nature of this trophic opportunist. Furthermore, the detection of a prevalent, possibly endoparasitic fungus (Purpureocillium lilacinum) in the shrimp's stomach demonstrates the wide range of questions that can be addressed using metabarcoding, towards a more robust reconstruction of ecological networks. 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? Yes  
URL https://datadryad.org/stash/dataset/doi:10.5061/dryad.sk2155m
 
Title Data from: Metabarcoding of shrimp stomach content: harnessing a natural sampler for fish biodiversity monitoring 
Description Given their positioning and biological productivity, estuaries have long represented key providers of ecosystem services, and consequently remain under remarkable pressure from numerous forms of anthropogenic impact. The monitoring of fish communities in space and time are one of the most widespread and established approaches to assess the ecological status of estuaries and other coastal habitats, but traditional fish surveys are invasive, costly, labour intensive and highly selective. Recently, the application of metabarcoding techniques, on either sediment or aqueous environmental DNA, has rapidly gained popularity. Here, we evaluate the application of a novel, high through-put DNA-based monitoring tool to assess fish diversity, based on the analysis of the gut contents of a generalist predator/scavenger, the European brown shrimp, Crangon crangon. Sediment and shrimp samples were collected from eight European estuaries and DNA metabarcoding (using both 12S and COI markers) was carried out to infer fish assemblage composition. We detected 32 teleost species (16 and 20, for 12S and COI respectively). Twice as many species were recovered using metabarcoding than by traditional net surveys. By comparing and interweaving trophic, environmental DNA and traditional survey-based techniques, we show that the DNA-assisted gut content analysis of a ubiquitous, easily accessible, generalist species may serve as a powerful, rapid and cost-effective tool for large scale, routine estuarine biodiversity monitoring. 
Type Of Material Database/Collection of data 
Year Produced 2018 
Provided To Others? Yes  
URL https://datadryad.org/stash/dataset/doi:10.5061/dryad.7498b88
 
Title Data from: Non-specific amplification compromises environmental DNA metabarcoding with COI 
Description 1. Metabarcoding extra-organismal DNA from environmental samples is now a key technique in aquatic biomonitoring and ecosystem health assessment. However, choice of genetic marker and primer set is a critical consideration when designing experiments, especially so when developing community standards and legislative frameworks. Mitochondrial cytochrome c oxidase subunit I (COI), the standard DNA barcode marker for animals, with its extensive reference library, taxonomic discriminatory power, and predictable sequence variation, is the natural choice for many metabarcoding applications such as the bulk sequencing of invertebrates. However, the overall utility of COI for environmental sequencing of targeted taxonomic groups has yet to be fully scrutinised. 2. Here, by using a case study of marine and freshwater fishes from the British Isles, we quantify the in silico performance of twelve mitochondrial primer pairs from COI, cytochrome b, 12S and 16S, in terms of reference library coverage, taxonomic discriminatory power, and primer universality. We subsequently test in vitro three COI primer pairs and one 12S pair for their specificity, reproducibility, and congruence with independent datasets derived from traditional survey methods at five estuarine and coastal sites in the English Channel and North Sea coast. 3. Our results show that for aqueous extra-organismal DNA at low template concentrations, both metazoan and fish-targeted COI primers perform poorly in comparison to 12S, exhibiting low levels of reproducibility due to non-specific amplification of prokaryotic and non-target eukaryotic DNAs. 4. An ideal metabarcode would have an extensive reference library for which custom primer sets can be designed for either broad assessments of biodiversity or taxon specific surveys, but unfortunately, low primer specificity hinders the use of COI, while the paucity of reference sequences is problematic for 12S. The latter, however, can be mitigated by expanding the concept of DNA barcodes to include whole mitochondrial genomes generated by genome-skimming existing tissue collections. 
Type Of Material Database/Collection of data 
Year Produced 2019 
Provided To Others? Yes  
URL https://datadryad.org/stash/dataset/doi:10.5061/dryad.b8f6s44
 
Title Reproduction influences seasonal eDNA variation in a temperate marine fish community 
Description All code and scripts required to reproduce the analysis of data in the paper. 
Type Of Material Data analysis technique 
Year Produced 2022 
Provided To Others? Yes  
Impact Opportunity to better understand the factors that explain the amount of fish eDNA detectable in temperate shelf seas. 
URL https://zenodo.org/record/6858158#.ZAnC8-zP2lM
 
Title Reproduction influences seasonal eDNA variation in a temperate marine fish community 
Description Full data sets for the above-mentioned paper 
Type Of Material Database/Collection of data 
Year Produced 2022 
Provided To Others? Yes  
Impact Understanding temporal variation of eDNA signatures in relation to fish life histories and reproduction 
URL https://www.ncbi.nlm.nih.gov/sra/PRJNA725897
 
Description Co-supervision of doctoral students 
Organisation University of Bologna
Country Italy 
Sector Academic/University 
PI Contribution I am an official co-supervisor of two PhD students registered at the University of Bologna. They will both visit my lab to process some of their samples.
Collaborator Contribution Sampling, provision of reagents, full-time labour of early-career researchers.
Impact data are yet to be generated. Outputs have not been produced yet.
Start Year 2023
 
Description Natural History Museum, London 
Organisation Natural History Museum
Country United Kingdom 
Sector Public 
PI Contribution We metabarcoded sponge samples, analysed data and drafted a manuscript for submission. This initial activity was at the foundation of the whole project proposal.
Collaborator Contribution Procurement of invaluable sponge tissue samples from the Mediterranean and Antarctica. After this initial study, the NHM team engaged with the co-creation of the project proposal for this grant.
Impact A paper was published in 2019 (see relevant section). A grant proposal based on these ideas was submitted to the July 2019 NERC Standard Grant round (NE/T007028/1), then becoming the present project.
Start Year 2018
 
Description University of La Reunion 
Organisation University of La Réunion
Country Réunion 
Sector Academic/University 
PI Contribution We have completed a study on eDNA metabarcoding around the island of La Reunion.
Collaborator Contribution Invaluable sampling activities from a remote tropical location.
Impact https://conbio.onlinelibrary.wiley.com/doi/full/10.1111/csp2.407
Start Year 2018
 
Description University of Palermo 
Organisation University of Palermo
Country Italy 
Sector Academic/University 
PI Contribution A network of researchers investigating fish biodiversity in 11 Mediterranean Marine Protected Areas, led by Prof. M. Milazzo from the University of Palermo, requested our team contribution to produce eDNA metabarcoding screening of the localities investigated by their project, using a range of 'traditional' methods, such as visual census and baited cameras.
Collaborator Contribution The partners carried out eDNA water sampling in 11 locations in Spain, France, Italy, Slovenia and Greece. They also sent a postdoctoral fellow to my lab to carry out most of the required lab work.
Impact We have completed the first draft manuscript from the project, which will be submitted in the coming weeks. An oral presentation was contributed to the FSBI SYmposium in HUll, July 2019, based on the above manuscript: "Integrated Biodiversity Assessment in Mediterranean Marine Protected Areas".
Start Year 2019
 
Description University of Rome 
Organisation University of Rome Tor Vergata
Department Department of Biology
Country Italy 
Sector Academic/University 
PI Contribution We have metabarcoded samples of "trawl slush" from survey vessels, in order to generate total biodiversity data.
Collaborator Contribution carried out sampling from Southern Italian continental shelf and slope.
Impact A manuscript has been submitted to the journal Frontiers in Ecology & the Environment, titled "All is fish that comes to the net: metabarcoding for rapid fisheries catch assessment".
Start Year 2018
 
Description BBC radio 4 Inside Science 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Extensive interview about the use of DNA reads from a shrimp's stomach to monitor fish diversity in estuaries.
Year(s) Of Engagement Activity 2018
 
Description European Researchers Night: Planet 2.0 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact - stand with demonstration materials for keen public.
- round-table discussion about the topic "can you study animals without seeing them"?
Year(s) Of Engagement Activity 2018
 
Description Radio Interview with the popular CBC science programme "Quirks and Quarks" 
Form Of Engagement Activity A press release, press conference or response to a media enquiry/interview
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact Live interview with a popular science radio broadcast, explaining and discussing the "natural sampler DNA" approach.
Link here:
https://www.cbc.ca/radio/quirks/june-22-is-your-wi-fi-watching-you-dog-s-manipulative-eyebrows-darwin-s-finches-in-danger-and-more-1.5182752/a-research-assistant-named-spongebob-sea-sponges-collect-data-for-science-1.5182754
Year(s) Of Engagement Activity 2019
URL https://www.cbc.ca/radio/quirks/june-22-is-your-wi-fi-watching-you-dog-s-manipulative-eyebrows-darwi...