Tag: eDNA

Dead Fish Shed More Intact Cells Into The Environment: A Promising Source For High-Resolution Genomic Analysis From Water

Study in Metabarcoding and Metagenomics investigated the availability of intact fish cells in water as a source of high-quality eDNA, using live and dead Rainbow Trout.

Guest blog post by Hiroki Yamanaka

Fish naturally release their DNA into surrounding waters, a phenomenon that has long powered environmental DNA (eDNA) analysis for detecting species without direct observation. Now, we are aiming for a leap beyond simple species identification toward fine-scale genomic analysis – a frontier that demands much higher-quality DNA samples.

Environmental DNA analysis as a biodiversity assessment tool

Environmental DNA (eDNA) analysis is a non-invasive technique that extracts and detects the genetic material of target species from environmental samples such as water and soil, and it has rapidly transformed biodiversity monitoring. Compared to traditional tools, like catch-and-release or visual surveys, this technique possesses remarkably higher sensitivity, demonstrating unparalleled utility in detecting rare, elusive, or invasive species. Because it is cost-effective and highly efficient, eDNA analysis has become a new biodiversity assessment tool for mapping species distribution on a broad scale.

However, the current primary objective of eDNA analysis largely remains at qualitative species identification – simply asking “what species are here?” – making it an urgent priority to advance the technique toward extracting detailed and much deeper ecological information.

The importance of the “individuality” of cells for further development

The next great frontier for eDNA analysis is its application to population genetics. To understand the health, evolutionary history, genetic diversity, inbreeding, or to identify hybrid individuals within specific populations, extracting high-quality genomic information across multiple genes (multilocus) is essential.

Chum salmon
Chum salmon. Photo credit to Jeff Duda, one of the co-authors of the study.

However, free-floating DNA in the environment (bulk eDNA) is rapidly degraded by microbial activity and environmental factors, leaving it highly fragmented. Furthermore, because it is a “genetic soup” mixed with shredded DNA from hundreds of different individuals, the loss of physical linkage – the “individuality” of the genetic material – remains the greatest roadblock.

To solve this, we focused on capturing “intact cells.” A physically undamaged cell membrane acts like a biological shield, protecting the fragile DNA inside from the harsh, degrading environment outside. By isolating these intact cells from environmental water and performing single-cell analysis, it would make it possible to read the complete, uncorrupted genetic fingerprint of a single individual. The study is now available in the open-access journal Metabarcoding & Metagenomics.

eDNA from dead fish is not just noise

To establish a method to specifically detect and quantify intact cells from environmental samples, we conducted a five-day laboratory experiment using live and dead Rainbow Trout (Oncorhynchus mykiss).

  • Experimental tanks used in the study. Photo by Hiroki Yamanaka, Ryukoku University.
    Experimental tanks used in the study. Photo by Hiroki Yamanaka, Ryukoku University.
  • Experimental tanks used in the study. Photo by Hiroki Yamanaka, Ryukoku University.
    Experimental tanks used in the study. Photo by Hiroki Yamanaka, Ryukoku University.

To distinguish the DNA inside intact cells from the free-floating, degraded DNA in the environment, we utilized propidium monoazide (PMA), a photoreactive DNA intercalating dye. PMA permeates into broken cell membrane and binds the DNA inside off the cell as well as the free-floating DNA outside of the cells; when exposed to strong visible light, it forms cross-links with DNA molecules that physically block subsequent PCR amplification. This effectively silences the “noise,” allowing us to selectively quantify only the pristine DNA protected inside intact cells with healthy membranes.

Rainbow Trout waiting for the introduction to the experimental tanks. Photo by Hiroki Yamanaka, Ryukoku University.

The results of this experiment were a revelation that flips conventional understanding in eDNA analysis. Historically, fish carcasses were viewed as a significant nuisance-biological noise that sheds massive amounts of DNA, causing false-positive detections or distorting biomass estimates.

But, this study demonstrated that carcasses release significantly higher concentrations of both total eDNA and PMA-resistant (intact-cell) eDNA compared to living fish. Particularly on Day 2 and Day 3 of the experiment, tanks containing carcasses yielded roughly 30 times more intact-cell DNA than tanks with live fish. In short, carcasses proved to be a promising source for the high-integrity cells that researchers desperately need.

Cell-by-cell analysis opens the door for the future

Counting and measuring Rainbow Trout used in the study. Photo by Hiroki Yamanaka, Ryukoku University.

This discovery presents a paradigm shift in our understanding of environmental sampling strategies for advanced genomic analysis. To acquire pristine genetic data, sampling waters during or immediately after known mass mortality events in nature-such as the natural die-offs after Pacific salmon migrate and spawn-could be a highly effective approach.

Access to perfectly preserved intact cells paves the way for a revolution in environmental monitoring. By integrating single-cell genomics with eDNA technology, we can overcome the loss of physical linkage that has plagued population genetics. Instead of looking at a mixed picture of DNA from multiple individuals, isolating individual cells for analysis makes it possible to compute exact inbreeding coefficients, map out local population structures, and identify hybrid species. This effectively bridges the gap between basic species detection and highly advanced ecological tracking, providing a technological foundation to deeply monitor the genetic health of entire ecosystems from just a few liters of water.

Limitations and further efforts required

Illustration of Double Helix DNA Structure
Illustration of Double Helix DNA Structure. Credit to Sumali Ibnu Chamid via Canva.

While this research provides pioneering insights, vital challenges requiring further validation remain. First, because this was a carefully controlled laboratory experiment using a single species (Rainbow Trout), further empirical studies involving a diverse array of taxa in complex, unpredictable natural environments are needed to ensure universal application.

Second, the technological tools – specifically the PMA dye method – require fine-tuning. Our observations confirmed that it sometimes struggles to fully suppress the PCR amplification of extracellular DNA, particularly when targeting the very short DNA fragments (short amplicons) often used in fish eDNA assays. Additionally, as a biological reality, DNA self-degradation by internal cellular enzymes continues even inside the cell. Thus, scientists still need to quantitatively evaluate and model the exact decay rates of DNA housed inside these protective cell membranes.

Despite these technical hurdles, the approach of harnessing the intact cells shed by organisms after they die brightly illuminates a highly promising path toward unlocking the full, high-resolution genetic story of our natural world with parallel development of single cell analysis on the cells from water shed by multicellular organisms.

Note: single cell analysis is commonly conducted on microbes which are unicellular organisms. However, single cell analysis on environment-medium derived multicellular organisms is very scarce for now. The main purpose of the current work was to determine a good timing / good location for cell collection to assure higher concentration of cells with high quality DNA to make the technical tests easier for the single cell analysis development on environmental samples.

Original source:

Yamanaka H, Hirohara T, Hoy MS, Chase DM, Duda JJ, Ostberg CO (2026) Live and dead fish shed different amounts of intact cells: Implications for advancing environmental DNA methodologies. Metabarcoding and Metagenomics 10: e177451. https://doi.org/10.3897/mbmg.10.177451

Revealing the Invisible: A New Baseline for Salish Sea Diatoms Answers a Global Call

A new checklist published in the Biodiversity Data Journal sheds light on the diversity of diatoms in the Salish Sea.

Invisible to the naked eye, phytoplankton play a critically important role at the base of marine food webs, yet their diversity continues to be underestimated. Recently, the UN Global Compact released “The Plankton Manifesto,” highlighting how these microscopic organisms are crucial for addressing the “triple planetary crisis” of climate change, biodiversity loss, and pollution. Diatoms, a major group of photosynthetic microalgae, are particularly powerful in driving roughly 20% of global photosynthesis and forming the very base of marine food webs.

Yet despite their monumental importance, microalgal ecosystems remain largely unexplored and poorly mapped. That is why a recent major scientific undertaking in the Northeast Pacific is so significant.

A First-of-its-Kind Baseline in the Salish Sea

sample locations from the Salish Sea
Map of sampling locations. A Distribution of sampling sites throughout the Salish Sea (green markers).Red frame: area of enlargement around Galiano Island (1B); B Sampling sites around Galiano Island (green markers). Map credit to Webber et al., 2026.

Diatom records of the Salish Sea bioregion have historically been fragmentary, dating back to early inventories in the 1800s, and with only scattered surveys filling the gap across the 20th and 21st centuries. As Andrew Simon, PhD student at the University of Alberta, president of IMERSS, and one of the study’s researchers, puts it:

The Salish Sea has long been studied for its rich marine biodiversity. Yet, until now, the history of research on its primary producers has been fragmented, and we have lacked a consolidated baseline record.

Now, for the first time, researchers have taken a significant step toward closing that gap. A team of Canadian researchers —  Mark Webber (University of Victoria; IMERSS), Elaine Humphrey (UVic; IMERSS), Arjan van Asselt (IMERSS), Alice Chang (UBC), Evan Morian (Hakai Institute; UBC), and Andrew Simon (IMERSS; University of Alberta) — has published a new checklist of 924 diatom taxa alongside a curated dataset of 11,469 records in the open-access journal Biodiversity Data Journal, providing a long-needed foundation for environmental monitoring across this region of the northeast Pacific Ocean.

  • Trigonium quinquelobatum
    Trigonium quinquelobatum, external valve view. SEM. Scale bar: 20 µm, Credit to Webber et al., 2026.
  • Cocconeis kerguelensis
    Cocconeis kerguelensis, SV. LM. Scale bar: 5 µm. Photo credit to Webber et al., 2026
  • Neocalyptrella robusta
    Neocalyptrella robusta, live, girdle view. LM. Scale bar: 50 µm, Credit to Webber et al., 2026

The findings include some curious discoveries. Several taxa are reported for the first time on the Pacific coast of North America, including Trigonium quinquelobatum, while others, such as Cocconeis kerguelensis (previously known only from the Indian Ocean) and Neocalyptrella robusta (previously confined to California), suggest a range expansion into cooler waters.

Ecologically, the genus Tabularia stands out as a dominant presence on eelgrass and macroalgae, despite having few species. In contrast, genera like Auliscus, Biddulphia, and Mastogloia are surprisingly scarce in the Salish Sea compared to other regions, leaving open questions about what limits their occurrence there.

This dataset also directly answers a key recommendation from the UN Plankton Manifesto, which urges the scientific community to strengthen plankton research and develop comprehensive plankton atlases to biomonitor the health of marine ecosystems.

Why Diatoms? Why Now?

Bacteriastrum hyalinium

Bacteriastrum hyalinium, valve view. LM. Scale bar 20 µm. Credit to Webber et al., 2026.

The Salish Sea —  the traditional territory of the Coast Salish peoples —  is home to roughly nine million people and is experiencing rapid growth in urbanization, industrial activity, and marine shipping.

Because diatom populations respond quickly to changes in water quality and environmental conditions, they serve as highly effective early-warning bioindicators for shifts in ecosystem health and pollution levels.

Without a clear picture of what the base of the food web looks like today, it is impossible to understand the impact of tomorrow’s environmental changes.

Attheya longicornis
Attheya longicornis, girdle view. SEM. Scale bar: 10 µm. Credit to Webber et al., 2026.

“We are fortunate to have had a dedicated group of academic researchers and community scientists contribute to this work over many years,” says Mark Webber, IMERSS’ resident diatomist. “Drawing from the literature, microscope analysis, and molecular sequencing, we now have a better picture of the diatoms present in the Salish Sea. Diatoms are vital to the health of countless organisms —  from shorebirds and shellfish to fish and mammals. This baseline provides a reference point for understanding changes that could ripple across the entire web of life.“

Local Research for Global Solutions

Actinoptychus adriaticus

Actinoptychus adriaticus var. pumila, exterior valve view. SEM. Scale bar: 5 µm. Credit to Webber et al., 2026.

The UN Plankton Manifesto stresses that understanding and managing plankton communities can unlock “Plankton-Based Solutions” to support fisheries, clean waters, and climate change mitigation.

Our work demonstrates how sustained collaboration between community scientists and research institutes can bridge these gaps, through partnering community expertise and observation with access to microscopy and molecular technologies,

the team concludes.

The new checklist and dataset will support researchers and policymakers in environmental assessments of the Salish Sea, as the team continues to refine and analyze the data to support ongoing regional research.

Original source

Webber M, Humphrey E, van Asselt A, Chang A, Morien E, Simon ADF (2026) Diatoms (Bacillariophyta) of the Salish Sea, Northeast Pacific: annotated checklist and new species reports. Biodiversity Data Journal 14: e189060. https://doi.org/10.3897/BDJ.14.e189060 

About Institute for Multidisciplinary Ecological Research in the Salish Sea (IMERSS)

IMERSS, or the Institute for Multidisciplinary Ecological Research in the Salish Sea, is a non-profit society based in Galiano Island, British Columbia, Canada. IMERSS joins scientific researchers, citizen scientists, and Indigenous communities to conduct multidisciplinary ecological research, monitor biodiversity and the environment, and communicate results to better understand and respond to change in the Salish Sea bioregion.

For more interesting research, follow  Biodiversity Data Journal,on Bluesky , Facebook and X.

A Simple Filter Swap Could Advance Marine eDNA Biomonitoring

A simple adjustment to water filtration methods can dramatically improve the detection of marine animal DNA when using advanced, PCR-free sequencing.

Over the past two decades, environmental DNA (eDNA) analysis has become a crucial tool for monitoring aquatic ecosystems. The most common method, metabarcoding, relies on PCR amplification of a smaller genetic region to identify specific taxa. However, PCR can lead to “significant taxonomic bias” because it often amplifies the DNA of different organisms unequally, making quantitative estimates difficult.

To avoid this, scientists have increasingly explored “shotgun sequencing“- an approach that sequences the DNA in a sample much more broadly – across the entire tree of life and across the genome. Unfortunately, in marine environments, shotgun sequencing is typically overwhelmed by microbial DNA, burying the genetic traces of less abundant macro-organisms such as animals.

Bigger Pores, Better Animal DNA Capture?

In a new study published in Metabarcoding and Metagenomics, researchers investigated if they could capture a higher proportion of eukaryotic (animal and plant) DNA simply by using filters with larger pore sizes.

Filter pore sizes are expected to influence results, since eDNA may be present in many different states, including but not limited to complete organisms, sloughed tissue, feces, free DNA, or gametes.

commented Dr. Adrián Gómez-Repollés, the lead author of the study.

To test this, the team collected 15 seawater samples from Skovshoved Harbour in Denmark and filtered them using pore sizes ranging from 0.2 µm to 8.0 µm. The results showed a stark contrast in the type of DNA captured based on the filter size.

Filters with smaller pore sizes (0.2 µm and 1.2 µm) retained a significantly greater proportion of bacterial reads than eukaryotic reads (63% vs. 28%); conversely, filters with larger pore sizes (5.0 µm and 8.0 µm) retained a significantly greater proportion of eukaryotic reads than bacterial reads (49% vs. 31%).

By switching to 5.0 µm or 8.0 µm filters, the researchers successfully reduced the dominance of bacteria. Of the 19 metazoan (animal) phyla detected using shotgun sequencing, all but one were found to be more abundant when using the larger pore sizes.

Looking to the Future of Biomonitoring

Taxonomic comparison at the kingdom and phylum levels of eukaryotes detected with shotgun sequencing and metabarcoding. A. Number and relative abundance (percentages) of taxonomically classified reads after rarefaction per sample and kingdom for shotgun sequencing; B. Number and relative abundance of taxonomically classified reads after rarefaction per sample and kingdom for metabarcoding. For both heatmaps, sample replicates are ordered along the y-axis by increasing pore size, starting with the enclosed filter type (EN) and continuing with the open pore filter type (OP). Pore sizes are in µm; C. Cladogram of the full set of eukaryotic phyla identified by shotgun sequencing and metabarcoding. Branch colors represent kingdoms (blue, Metazoa; red, Fungi; green, Viridiplantae; pink, other eukaryotes). Three surrounding rings indicate phyla shared between shotgun sequencing and metabarcoding (gray), phyla uniquely detected with metabarcoding (yellow), and phyla uniquely detected with shotgun sequencing (blue). Information in the rings is summarized in a Venn diagram in the upper left corner. Taxa marked with an asterisk are no longer categorized as a phylum. Credit to Gómez-Repollés et al., 2026

When compared alongside traditional 18S rDNA metabarcoding, the shotgun sequencing method successfully shared 39 of the 54 detected eukaryotic phyla, indicating a similar performance in detecting the presence of high-level taxonomic groups.

To test the potential of shotgun sequencing in applied biomonitoring even further, the researchers examined the results at genus-level for a number of well known marine animals such as fish, mussels, crustaceans and bristle worms. Here, they found both DNA matches to native Danish species but also to exotic taxa that were highly unlikely and probably due to the low level of resolution in shotgun sequencing, where genetic regions of low variation and coverage are sequenced.

Water sample collection. Photo by David Stanciu.

But, when they looked further into the results they observed that the local taxa consistently comprised a higher number of reads. The number of reads could thus be a simple way to initially separate authentic taxa from erroneous matches in eDNA studies based on shotgun sequencing, although the approach needs further testing. 

However, the authors note current limitations with the technology, primarily driven by incomplete public DNA reference databases. In the study, only 0.78% of the total shotgun reads could be definitively assigned to a superkingdom level. Another drawback was the lack of field controls to rule out cross-contamination or input from airborne DNA, coupled with a limited spatiotemporal design involving only a single sampling location. 

Despite these hurdles, shotgun sequencing and filters with larger pore sizes could potentially be a significant step forward for eDNA in marine biology. As global genomic databases continue to expand, “the taxonomic coverage and resolution of shotgun sequencing should improve, likely enhancing the potential of shotgun sequencing for future eDNA research“, says Philip Francis Thomsen, professor and senior author on the study. 

Original study:

Gómez-Repollés A, Sigsgaard EE, Jensen MR, Thomsen PF (2026) Filter pore size influences taxonomic composition of retained eDNA from seawater samples—evidence from shotgun sequencing. Metabarcoding and Metagenomics 10: e164232. https://doi.org/10.3897/mbmg.10.164232

Shining a light on dark taxa: new fungal phyla and classes revealed by eDNA long reads

Molecular analyses of soil and water commonly reveal large proportions of fungal taxa that cannot be assigned to any taxonomic or functional groups. Some of these so-called ‘dark taxa’ have been encoded alphanumerically, while others have remained completely overlooked.

Recent advances in long-read sequencing techniques have produced large amounts of high-quality rRNA marker gene data about eukaryotic organisms, but many of these taxa have remained unknown at the highest taxonomic levels: phylum and kingdom. 

Now, via a thorough analysis of the EUKARYOME long-read database, an international team led by Prof Leho Tedersoo (University of Tartu, Estonia) has discovered that a large proportion of the unknown eukaryotes belong to deep, hitherto undescribed fungal lineages.

By developing innovative approaches in taxonomy and performing rigorous phylogenetic analyses, the researchers described 30 novel fungal lineages from the order to phylum levels, including the type species of these groups. They published their findings in the open-access journal MycoKeys.

Names of new taxa were developed and voted for by all co-authors, with the names referring to type locality using the native language stems (Amerindian, Sámi, Estonian) prevailing. 

The authors also proposed the taxonomic terms “nucleotype” and “legitype” to refer to holotype-derived DNA samples and DNA sequences, respectively, which under certain circumstances (e.g., when holotype is lost) could also be used as types.

The taxonomic approach developed by Tedersoo et al. provides a means of describing and communicating unseen, potentially uncultivable microeukaryotic taxa.

Original source

Tedersoo L, Hosseyni Moghadam MS, Panksep K, Prins V, Anslan S, Mikryukov V, Bahram M, Abarenkov K, Kõljalg U, Esmaeilzadeh-Salestani K, Pawłowska J, Wurzbacher C, Ding Y, Alkahtani SH, Nilsson RH (2025) Thirty novel fungal lineages: formal description based on environmental samples and DNA. MycoKeys 124: 1-121. https://doi.org/10.3897/mycokeys.124.161674

Follow MycoKeys on Facebook and Bluesky.

Pensoft Publishers and the Southern eDNA Society join forces to advance eDNA research

The partnership aims to accelerate the development and application of eDNA technology and increase the visibility and impact of eDNA research.

Pensoft Publishers, a leading open-access publisher of scientific journals, has signed a Memorandum of Understanding with the Southern eDNA Society (SeDNAs) to foster collaboration and knowledge sharing in the field of environmental DNA (eDNA) research. By working together, the two organisations aim to accelerate the development and application of eDNA technology, ultimately contributing to a better understanding and conservation of our planet’s biodiversity.

The Southern eDNA Society is a recently established organisation of about 200 members that aims to facilitate communication, collaboration and knowledge transfer in the rapidly developing field of eDNA.

Pensoft is an innovative scholarly publisher of over 35 open-access, peer-reviewed journals across various disciplines, including the highly regarded Metabarcoding and Metagenomics (MBMG) journal. 

Featuring research on metabarcoding and metagenomics from both basic and applied perspectives, MBMG has a CiteScore of 5.4 and is indexed in Web of Science and Scopus. The journal is published on Pensoft’s end-to-end scholarly publishing platform ARPHA.

New open-access journal Metabarcoding & Metagenomics joins the lines of publisher Pensoft

The partnership will focus on enhancing the publication and dissemination of eDNA research, and increasing the visibility and impact of eDNA research, including by promoting eDNA conferences hosted by SeDNAs and research outputs from the eDNA community.

The Southern eDNA Society is excited to partner with the MBMG journal to advance our shared commitment to innovation, accessibility, and excellence in eDNA science. The exclusive discounted publication opportunities for SeDNAs members will help amplify their contributions to the global eDNA community.

Anastasija Zaiko, President of SeDNAs

To this end, both organisations will cross-promote each other’s activities, reaching a wider audience and fostering greater engagement within the eDNA community. Pensoft plans to support the dissemination of research presented at SeDNAs conferences, with the possibility of publishing special issues or topical collections in MBMG.

In addition, SeDNA members will receive a special discount on article processing charges in MBMG, which will hopefully encourage them to publish their work in the innovative open-access journal.

This memorandum of understanding builds upon a shared vision of advancing eDNA research and making scientific knowledge openly accessible. Pensoft has always expressed a commitment for open science and technological advancement and we are happy that we can take this step to support the growth and dissemination of eDNA science.

Prof. Lyubomir Penev, founder and CEO of Pensoft

The partnership between Pensoft, MBMG and SeDNAs is not the first of its kind for the scholarly publisher. Recently, another journal launched and published by Pensoft, Nature Conservation, used a similar memorandum to strengthen the collaboration between the publisher and the Infrastructure & Ecology Network Europe (IENE). IENE has so far published several special issues of research papers based on scientific work presented at the network’s conferences in Nature Conservation, with the most recent example, a collection of articles associated with the IENE 2022 International Conference, dating from last month. 

Biodiversity in a bird’s nest: DNA as a tool for bird conservation

Researchers employ eDNA to investigate ‘fascinating microcosms’ inside birds’ nests.

Researchers have found that advanced DNA technologies can get a detailed snapshot of insect diversity within a bird’s nest, showing everything from the bird’s last meal to disease-causing parasites.

A bird’s nest. Photo by Farrini

“Birds’ nests are fascinating microcosms, but until now, studies have only examined the living insects that can be seen crawling and flying around the nests,” says Valerie Levesque-Beaudin, lead author on the study and a leading expert in Diptera taxonomy at the Centre for Biodiversity Genomics (CBG) at the University of Guelph (U of G).

With newer DNA-based methods, researchers can pick up traces of environmental DNA to get a snapshot of all the species in these tiny ecosystems. “The analysis of nest contents and environmental DNA, or ‘eDNA’ as it’s called, via metabarcoding helps us to gain more insight into a bird’s diet, parasites, and other factors that could impact a bird’s health and breeding success,” says Levesque-Beaudin.

For the study, published in Metabarcoding and Metagenomics, researchers collected 20 birds’ nests from the 162-hectare Arboretum at U of G. They examined the nests using DNA barcoding to identify insects to species and DNA metabarcoding to look at the entire nest ecosystem.

Organisms leave traces of DNA behind as they move through the environment, and researchers can use metabarcoding to build a comprehensive picture of life in the nest. Metabarcoding pulls all DNA traces in a bulk sample – in this case, parts of dead insects, debris, and dust from birds’ nests. This method differs from DNA barcoding, where a single specimen – an insect in this case – is DNA sequenced to identify it to species level.

The CBG team used emergence traps for a first sweep of the nest’s contents followed by a second, deeper probe using DNA metabarcoding to identify all the species encountered in the nest. Researchers passed the nests through a sieve, collecting insect remains and the dust for DNA extraction. “We not only found insects making a living in the nest, but traces of prey, parasites, and many other things,” says Levesque-Beaudin. “The most unexpected was the amount of information gained on other birds’ species whose feathers were either used for nest building or whose nests were essentially overbuilt by the nesting species.”

A tent-like emergence trap is used to contain and collect insects living inside the nest debris. PHOTO: VALERIE LEVESQUE-BEAUDIN

“This approach has the potential to revolutionize how we study bird nests as a micro-ecosystem. It unravels connections between different ecological guilds within the nest and connections of the birds with their environment, which would otherwise remain hidden,” says Dr. Bettina Thalinger, senior author of the study.

The CBG’s Associate Director of Analytics, Dr. Dirk Steinke, says the study has positive implications for bird conservation efforts. He says his students have already begun looking at American Kestrels, a threatened bird of prey, to find out if there are clues in the nest communities via metabarcoding and if DNA can help scientists determine if lack of prey or increased parasitism could be among the causes of nestling mortalities.

Darwin’s finch, also called Galápagos finch. PHOTO BY CHRIS HO, CENTRE FOR BIODIVERSITY GENOMICS.

Galapagos finches are another species threatened by the avian vampire fly – a parasite that attacks nestlings – and treatments include pesticides. Steinke notes that one of his graduate students has begun using DNA metabarcoding in the finches’ nests to understand better the potential impact of pesticide treatment on the entire arthropod nest community.

Research article:

Levesque-Beaudin V, Steinke D, Böcker M, Thalinger B (2023) Unravelling bird nest arthropod community structure using metabarcoding. Metabarcoding and Metagenomics 7: e103279. https://doi.org/10.3897/mbmg.7.103279

News piece originally published by the Centre of Biodiversity Genomics. Republished with permission.

Simplified method to survey amphibians will aid conservation

Researchers developed a method to determine which amphibians inhabit a specific area. The new technique will resolve some of the issues with conventional methods, such as capture and observational surveys.

Ryukyu Sword Tailed Newt, or Firebellied Newt. Photo by Neil Dalphin via Creative Commons CC0.

An international collaborative research group of members from seven institutions has developed a method to determine which amphibians (frogs, newts and salamanders) inhabit a specific area. Their work was published in the open-access, peer-reviewed journal Metabarcoding and Metagenomics (MBMG).

To do so, the scientists amplified and analysed extra-organismal DNA (also known as environmental DNA or eDNA) found in the water. This DNA ends up in the water after being expelled from the amphibian’s body along with mucus and excrement. 

The research group included Postdoctoral Researcher Sakata K. Masayuki and Professor Minamoto Toshifumi (Kobe University), Associate Professor Kurabayashi Atsushi (Nagahama Institute of Bio-Science and Technology), Nakamura Masatoshi (IDEA Consultants, Inc.) and Associate Professor Nishikawa Kanto (Kyoto University). 

The newly developed technique will resolve some of the issues with conventional methods, such as capture and observational surveys, which require a specialist surveyor who can visually identify species. Conventional surveys are also prone to discrepancies due to environmental factors, such as climate and season.

The researchers hope that the new method will revolutionise species monitoring, as it will enable anyone to easily monitor the amphibians that inhabit an area by collecting water samples.  

While monitoring in general is crucial to conserve the natural ecosystems, the importance of surveying amphibians is even more pressing, given the pace of their populations’ decline.

Amongst major obstacles to amphibian monitoring, however, are the facts that they are nocturnal; their young (e.g. tadpoles) and adults live in different habitats; and that specialist knowledge is required to capture individuals and identify their species. These issues make it particularly difficult to accurately survey amphibians in a standardised way, and results of individual efforts often contradict each other.

On the other hand, eDNA analysis techniques have already been established in programmes targeted at monitoring fish species, where they are already commonplace. So, the researchers behind the present study joined forces to contribute towards the development of a similar standardised analysis method for amphibians.

First of all, the researchers designed multiple methods for analysing the eDNA of amphibians and evaluated their performance to identify the most effective method. Next, they conducted parallel monitoring of 122 sites in 10 farmlands across Japan using the developed eDNA analysis along with the conventional methods (i.e. capture surveys using a net and observation surveys). 

As a result, the newly developed method was able to detect all three orders of amphibians: Caudata (the newts and salamanders), Anura (the frogs), and Gymnophiona (the caecilians). 

Furthermore, this novel eDNA analysis method was able to detect more species across all field study sites than the conventional method-based surveys, indicating its effectiveness.

Research Background

Amphibian biodiversity is continuing to decline worldwide and collecting basic information about their habitats and other aspects via monitoring is vital for conservation efforts. Traditional methods of monitoring amphibians include visual and auditory observations, and capture surveys.

However, amphibians tend to be small in size and many are nocturnal. The success of surveys varies greatly depending on the climate and season, and specialist knowledge is required to identify species. Consequently, it is difficult to monitor a wide area and assess habitats. The last decade has seen the significant development of environmental DNA analysis techniques, which can be used to investigate the distribution of a species by analysing external DNA (environmental DNA) that is released into the environment along with an organism’s excrement, mucus and other bodily fluids. 

The fundamentals of this technique involve collecting water from the survey site and analysing the eDNA contained in it to find out which species inhabit the area. In recent years, the technique has gained attention as a supplement for conventional monitoring methods. Standardised methods of analysis have already been established for other species, especially fishes, and diversity monitoring using eDNA is becoming commonplace. 

However, eDNA monitoring of amphibians is still at the development stage. One reason for this is that the proposed eDNA analysis method must be suitable for the target species or taxonomic group, and there are still issues with developing and implementing a comprehensive method for detecting amphibians. If such a method could be developed, this would make it possible for monitoring to be conducted even by people who do not have the specialised knowledge to identify species nor surveying experience.

Hopefully, this would be established as a unified standard for large-scale monitoring surveys, such as those on a national scale. This research group’s efforts to develop and evaluate analysis methods will hopefully lay the foundations for eDNA analysis to become a common tool for monitoring amphibians, as well as fish. 

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Follow Metabarcoding and Metagenomics (MBMG) journal on Twitter and Facebook.

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Research article: 

Sakata MK, Kawata MU, Kurabayashi A, Kurita T, Nakamura M, Shirako T, Kakehashi R, Nishikawa K, Hossman MY, Nishijima T, Kabamoto J, Miya M, Minamoto T (2022) Development and evaluation of PCR primers for environmental DNA (eDNA) metabarcoding of Amphibia. Metabarcoding and Metagenomics 6: e76534. https://doi.org/10.3897/mbmg.6.76534

Extensive practical guide to DNA-based biodiversity assessment methods published as a ‘living’ document by DNAqua-Net COST Action

Between 2016 and 2021, over 500 researchers collaborated within the DNAqua-Net international network, funded by the European Union’s European Cooperation in Science and Technology programme (COST), with the goal to develop and advance biodiversity assessment methods based on analysis of DNA obtained from the environment (e.g. river water) or from unsorted collections of organisms. 

Such innovative methods are a real game changer when it comes to large-scale assessment of biodiversity and ecological monitoring, as collecting environmental samples that are sent to the lab for analysis is much cheaper, faster and non-invasive, compared with capturing and examining live organisms. However, large-scale adoption has been hindered by a lack of standardisation and official guidance. 

Recognising the urgent need to scale up ecological monitoring as we respond to the biodiversity and climate crises, the DNAqua-Net team published a guidance document for the implementation of DNA-based biomonitoring tools.

The guide considers four different types of samples: water, sediments, invertebrate collections and diatoms, and two primary analysis types: single species detection via qPCR and similar targeted methods; and assessment of biological communities via DNA metabarcoding. At each stage of the field and laboratory process the guide sets out the scientific consensus, as well as the choices that need to be made and the trade-offs they entail. In particular, the guide considers how the choices may be influenced by common practical constraints such as logistics, time and budget. Available in an Advanced Book format, the guidelines will be updated as the technology continues to evolve.

Leaders of DNAqua-Net are Prof. Dr. Florian Leese of the University of Duisburg-Essen (Germany) and Dr. Agnès Bouchez of the French National Institute for Agriculture, Food, and Environment (INRAE). The core writing team for the present guide book involves Dr. Micaela Hellström (MIX Research AB, Sweden), Dr. Kat Bruce (NatureMetrics Ltd., UK), Dr. Rosetta Blackman (University of Zurich and EAWAG, Switzerland), Dr. Sarah Bourlat (LIB/Museum Koenig, Germany), and Prof. Kristy Deiner (ETH Zurich and SimplexDNA AG, Switzerland).

“Back in 2016 we realised that all around the globe researchers are testing new eDNA methods, developing individual solutions and products. While this is excellent, we need to reach a consensus and provide this consensus to stakeholders from the applied sectors”, 

says Florian Leese.
This video was created as part of EU COST Action DNAqua-Net (CA15219) and shows how environmental DNA (eDNA) can be sampled and analysed from aquatic ecosystems. It shows the whole cycle from the start to final results. 
Credit: DNAqua-Net

The guide’s lead author Dr. Kat Bruce adds:

“The urgency of addressing the twin biodiversity and climate crises means that we need to accelerate the adoption of new technologies that can provide data and insights at large scales. In doing so, we walk a tricky line to agree on sufficiently standardised methods that can be usefully applied as soon as they add value, while still continuing to develop them further and innovate within the field. It was a daunting task to seek consensus from several hundred scientists working in a fast-moving field, but we found that our technology is based on a strong foundation of knowledge and there was a high level of agreement on the core principles – even if the details vary and different users make different choices depending on their environmental, financial or logistical constraints.”

Looking back on the last four years that culminated in the publication of a “living” research publication, Prof. Dr. Kristy Deiner says:

“The document took many twists and turns through more than ten versions and passionate discussions across many workshops and late night drinks. All in the days when we could linger at conferences without fear of the pandemic weighing on us. As we worked to find consensus, one thing was clear: we had a lot to say and a standard review paper was not going to cut it. With the knowledge and experience gathered across the DNAqua-Net, it made sense to not limit this flow of information, but rather to try and tackle it head on and use it to address the many questions we’ve all struggled with while developing DNA-based biodiversity survey methods.”

Now that the document – or at least its first version – is publicly available, the researchers are already planning for the next steps and challenges.

“The bottom line is we’ve come a long way in the last ten years. We have a buffet of methods for which many produce accurate, reliable and actionable data to the aid of biodiversity monitoring and conservation. While there is still much work to be done, the many unanswered questions are because the uptake is so broad. With this broad uptake comes novel challenges, but also new insights and a diversity of minds with new ideas to address them. As said this is planned to be a living document and we welcome continued inputs no matter how great or small,” says Deiner.

Dr. Micaela Hellström recalls:

“The book evolved over the four years of COST Action DNAqua-Net which made it possible for the many scientists and stakeholders involved to collaborate and exchange knowledge on an unprecedented scale. Our whole team is well aware of the urgent need to monitor biodiversity loss and to provide accurate species distribution information on large scales, to protect the species that are left. This was a strong driving force for all of us involved in the production of this document. We need consensus on how to coherently collect biodiversity data to fully understand changes in nature.”

“It was a great and intense experience to be a part of the five-person core writing team. In the months prior to submitting the document, we spent countless hours, weekends and late nights researching the field, communicating with researchers and stakeholders, and joining vivid Zoom discussions. As a result, the present book provides solid guidance on multiple eDNA monitoring methods that are – or will soon become – available as the field moves forward.” 

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The DNAqua-Net team invites fellow researchers and practitioners to provide their feedback and personal contributions using the contacts below.

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Original source:

Bruce K, Blackman R, Bourlat SJ, Hellström AM, Bakker J, Bista I, Bohmann K, Bouchez A, Brys R, Clark K, Elbrecht V, Fazi S, Fonseca V, Hänfling B, Leese F, Mächler E, Mahon AR, Meissner K, Panksep K, Pawlowski J, Schmidt Yáñez P, Seymour M, Thalinger B, Valentini A, Woodcock P, Traugott M, Vasselon V, Deiner K (2021) A practical guide to DNA-based methods for biodiversity assessment. Advanced Books. https://doi.org/10.3897/ab.e68634

One water bucket to find them all: Detecting fish, mammals, and birds from a single sample

Revolutionary environmental DNA analysis holds great potential for the future of biodiversity monitoring, concludes a new study.

Revolutionary environmental DNA analysis holds great potential for the future of biodiversity monitoring, concludes a new study

Collection of water samples for eDNA metabarcoding bioassessment.
Photo by Till-Hendrik Macher.

In times of exacerbating biodiversity loss, reliable data on species occurrence are essential, in order for prompt and adequate conservation actions to be initiated. This is especially true for freshwater ecosystems, which are particularly vulnerable and threatened by anthropogenic impacts. Their ecological status has already been highlighted as a top priority by multiple national and international directives, such as the European Water Framework Directive.

However, traditional monitoring methods, such as electrofishing, trapping methods, or observation-based assessments, which are the current status-quo in fish monitoring, are often time- and cost-consuming. As a result, over the last decade, scientists progressively agree that we need a more comprehensive and holistic method to assess freshwater biodiversity.

Meanwhile, recent studies have continuously been demonstrating that eDNA metabarcoding analyses, where DNA traces found in the water are used to identify what organisms live there, is an efficient method to capture aquatic biodiversity in a fast, reliable, non-invasive and relatively low-cost manner. In such metabarcoding studies, scientists sample, collect and sequence DNA, so that they can compare it with existing databases and identify the source organisms.

Furthermore, as eDNA metabarcoding assessments use samples from water, often streams, located at the lowest point, one such sample usually contains not only traces of specimens that come into direct contact with water, for example, by swimming or drinking, but also collects traces of terrestrial species indirectly via rainfalls, snowmelt, groundwaters etc. 

In standard fish eDNA metabarcoding assessments, these ‘bycatch data’ are typically left aside. Yet, from a viewpoint of a more holistic biodiversity monitoring, they hold immense potential to also detect the presence of terrestrial and semi-terrestrial species in the catchment.

In their new study, reported in the open-access scholarly journal Metabarcoding and Metagenomics, German researchers from the University of Duisburg-Essen and the German Environment Agency successfully detected an astonishing quantity of the local mammals and birds native to the Saxony-Anhalt state by collecting as much as 18 litres of water from across a two-kilometre stretch along the river Mulde.

After water filtration the eDNA filter is preserved in ethanol until further processing in the lab.
Photo by Till-Hendrik Macher.

In fact, it took only one day for the team, led by Till-Hendrik Macher, PhD student in the German Federal Environmental Agency-funded GeDNA project, to collect the samples. Using metabarcoding to analyse the DNA from the samples, the researchers identified as much as 50% of the fishes, 22% of the mammal species, and 7.4% of the breeding bird species in the region. 

However, the team also concluded that while it would normally take only 10 litres of water to assess the aquatic and semi-terrestrial fauna, terrestrial species required significantly more sampling.

Unlocking data from the increasingly available fish eDNA metabarcoding information enables synergies among terrestrial and aquatic biodiversity monitoring programs, adding further important information on species diversity in space and time. 

“We thus encourage to exploit fish eDNA metabarcoding biodiversity monitoring data to inform other conservation programs,”

says lead author Till-Hendrik Macher. 

“For that purpose, however, it is essential that eDNA data is jointly stored and accessible for different biodiversity monitoring and biodiversity assessment campaigns, either at state, federal, or international level,”

concludes Florian Leese, who coordinates the project.

Original source:

Macher T-H, Schütz R, Arle J, Beermann AJ, Koschorreck J, Leese F (2021) Beyond fish eDNA metabarcoding: Field replicates disproportionately improve the detection of stream associated vertebrate species. Metabarcoding and Metagenomics 5: e66557. https://doi.org/10.3897/mbmg.5.66557

Scientists took a rare chance to prove we can quantify biodiversity by ‘testing the water’

Recent study conducted at a UK fishery farm provides new evidence that DNA from water samples can accurately determine fish abundance and biomass

Organisms excrete DNA in their surroundings through metabolic waste, sloughed skin cells or gametes, and this genetic material is referred to as environmental DNA (eDNA).

As eDNA can be collected directly from water, soil or air, and analysed using molecular tools with no need to capture the organisms themselves, this genetic information can be used to report biodiversity in bulk. For instance, the presence of many fish species can be identified simultaneously by sampling and sequencing eDNA from water, while avoiding harmful capture methods, such as netting, trapping or electrofishing, currently used for fish monitoring.

While the eDNA approach has already been applied in a number of studies concerning fish diversity in different types of aquatic habitats: rivers, lakes and marine systems, its efficiency in quantifying species abundance (number of individuals per species) is yet to be determined. Even though previous studies, conducted in controlled aquatic systems, such as aquaria, experimental tanks and artificial ponds, have reported positive correlation between the DNA quantity found in the water and the species abundance, it remains unclear how the results would fare in natural environments.

However, a research team from the University of Hull together with the Environment Agency (United Kingdom), took the rare opportunity to use an invasive species eradication programme carried out in a UK fishery farm as the ultimate case study to evaluate the success rate of eDNA sampling in identifying species abundance in natural aquatic habitats. Their findings were published in the open-access, peer-reviewed journal Metabarcoding and Metagenomics.

“Investigating the quantitative power of eDNA in natural aquatic habitats is difficult, as there is no way to ascertain the real species abundance and biomass (weight) in aquatic systems, unless catching all target organisms out of water and counting/measuring them all,”

explains Cristina Di Muri, PhD student at the University of Hull.
Drained pond after fish translocation.
Photo by Dr. Watson H.V.

During the eradication, the original fish ponds were drained and all fish, except the problematic invasive species: the topmouth gudgeon, were placed in a new pond, while the original ponds were treated with a piscicide to remove the invasive fish. After the eradication, the fish were returned to their original ponds. In the meantime, all individuals were counted, identified and weighed from experts, allowing for the precise estimation of fish abundance and biomass.

“We then carried out our water sampling and ran genetic analysis to assess the diversity and abundance of fish genetic sequences, and compared the results with the manually collected data. We found strong positive correlations between the amount of fish eDNA and the actual fish species biomass and abundance, demonstrating the existence of a strong association between the amount of fish DNA sequences in water and the actual fish abundance in natural aquatic environments,”

reports Di Muri.
Environmental DNA sampling using water collection bottles
Photo by Dr. Peirson G.

The scientists successfully identified all fish species in the ponds: from the most abundant (i.e. 293 carps of 852 kg total weight) to the least abundant ones (i.e. one chub of 0.7 kg), indicating the high accuracy of the non-invasive approach.

“Furthermore, we used different methods of eDNA capture and eDNA storage, and found that results of the genetic analysis were comparable across different eDNA approaches. This consistency allows for a certain flexibility of eDNA protocols, which is fundamental to maintain results comparable across studies and, at the same time, choose the most suitable strategy, based on location surveyed or resources available,”

elaborates Di Muri.

“The opportunity of using eDNA analysis to accurately assess species diversity and abundance in natural environments will drive a step change in future species monitoring programmes, as this non-invasive, flexible tool is adaptable to all aquatic environments and it allows quantitative biodiversity surveillance without hampering the organisms’ welfare.”

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Original publication:

Di Muri C, Lawson Handley L, Bean CW, Li J, Peirson G, Sellers GS, Walsh K, Watson HV, Winfield IJ, Hänfling B (2020) Read counts from environmental DNA (eDNA) metabarcoding reflect fish abundance and biomass in drained ponds. Metabarcoding and Metagenomics 4: e56959. https://doi.org/10.3897/mbmg.4.56959