Metabarcoding and Metagenomics

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.”

***

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

DNA metabarcoding detects ecological stress within freshwater species

Metabarcoding allows scientists to extract DNA from the environment, in order to rapidly detect species inhabiting a particular habitat. While the method is a great tool that facilitates conservation activities, few studies have looked into its applicability in monitoring species’ populations and their genetic diversity, which could actually be critical to assess negative trends early on. The potential of the method is confirmed in a new study, published in the peer-reviewed scholarly journal Metabarcoding & Metagenomics.

In a new study, German scientists confirm that responses below species level can be inferred with DNA metabarcoding

Metabarcoding allows scientists to extract DNA from the environment (known as environmental DNA or eDNA), for example, river water or, as in the case of the study by the team from the University of Duisburg-Essen (Essen, Germany) within the German Barcode of Life project (GBOL II): Vera Zizka, Dr Martina Weiss and Prof Florian Leese, from individuals in bulk samples. Thus, they are able to detect what species inhabit a particular habitat.

However, while the method has already been known to be of great use in getting an approximate picture of local fauna, hence facilitating conservation prioritisation, few studies have looked into its applicability to infer responses below species level. That is, how the populations of a particular species fare in the environment of interest, also referred to as intraspecific diversity. Meanwhile, the latter could actually be a lot more efficient in ecosystem monitoring and, consequently, biodiversity loss mitigation.

The potential of the method is confirmed in a new study, published in the peer-reviewed scholarly journal Metabarcoding & Metagenomics. To do so, the researchers surveyed the populations of macroinvertebrate species (macrozoobenthos) in three German rivers: Emscher, Ennepe and Sieg, where each is subject to a different level of ecological disturbance. They were looking specifically at species reported at all of the survey sites by studying the number of different haplotypes (a set of DNA variations usually inherited together from the maternal parent) in each sample. The researchers point out that macrozoobenthos play a key role in freshwater ecosystem functionality and include a wide range of taxonomic groups with often narrow and specific demands with respect to habitat conditions.

“As the most basal level of biodiversity, genetic diversity within species is typically the first to decrease, and the last to regenerate, after stressor’s impact. It consequently provides a proxy for environmental impacts on communities long before, or even if never visible on species diversity level,”
explain the scientists.

Emscher is an urban stream in the Ruhr Metropolitan Area that has been used as an open sewage channel for the past hundred years, and is considered to be a very disturbed environment. Ennepe – regarded as moderately stressed – runs through both rural and urban sites, including ones with sewage treatment plant inflow. Meanwhile, Sieg is considered as a stable, near-natural river system with a good ecological and chemical status.

As a result, despite their original assumption that Sieg would support the most prominent diversity within populations of species sensitive to organic pollution, such as mayflies, stoneflies and caddisflies, the scientists reported no significant difference to the medium stressed river Ennepe. This was also true for overall biodiversity. On the other hand, the team discovered higher intraspecific diversity for species resilient to ecological disturbance like small worms and specialised crustaceans in the heavily disturbed Emscher. The latter phenomenon may be explained with low competition pressure for these species, their ability to use organic compounds as resources and, consequently, increased population growth.

“[T]his pioneer study shows that the extraction of intraspecific genetic variation, so-called ‘haplotypes’ from DNA metabarcoding datasets is a promising source of information to assess intraspecific diversity changes in response to environmental impacts for a whole metacommunity simultaneously,”
conclude the scientists.

However, the researchers also note limitations of their study, including the exclusion of specialist species that only occured at single sites. They prompt future studies to also carefully control for the individual number of specimens per species to quantify genetic diversity change specifically.

###

Original source:

Zizka VMA, Weiss M, Leese F (2020) Can metabarcoding resolve intraspecific genetic diversity changes to environmental stressors? A test case using river macrozoobenthos. Metabarcoding and Metagenomics 4: e51925. https://doi.org/10.3897/mbmg.4.51925

Pan-European sampling campaign sheds light on the massive diversity of freshwater plankton

In a major pan-European study, a research team from Germany have successfully extracted environmental DNA (eDNA) from as many as 218 lakes to refute a long-year belief that vital microorganisms do not differ significantly between freshwater bodies and geographic regions the way plants and animals do.

Their new-age approach to biodiversity studies resulted in the largest freshwater dataset along with a study published in the open access journal Metabarcoding and Metagenomics.

Surface freshwaters are of critical importance for terrestrial life and, in particular, human life and welfare. However, these vital ecosystems are severely understudied, as compared to terrestrial or oceanic biomes, and so are the microbial organisms living in them.

On the other hand, it is these invisible to the naked eye creatures, called protists, that are responsible for keeping our ecosystems running. Their diversity and their high metabolic rates maintain ecosystem stability. In fact, microbes are the major source of the worlds oxygen.

In 2012, the team of Prof. Jens Boenigk, University of Duisburg-Essen, undertook the sampling campaign to study the distribution pattern of microbial organisms on a continental scale and the impact of Europe’s climatic history on their present-day whereabouts.

They sampled freshwater lakes and ponds from sites in Norway, Sweden, Germany, Poland, the Czech Republic, Slovakia, Hungary, Romania, Austria, Italy, France, Spain and Switzerland. Site selection focused on the European orogens, specifically the Alps, the Pyrenees, the Apennine, the High Tatras, the southern Scandinavian mountains and the connecting flatlands.

Thanks to the excellent collaboration both within the team and with a number of scientific institutions across Europe, which gave their support as access points for re-stocking sampling equipment and immediate sample preservation, the campaign delivered groundbreaking results illuminating the hidden diversity of the microbial biosphere.

The scientists reported that plankton diversity was highly partitioned between lakes which bear distinct biological fingerprints. In particular, high mountain ranges imprinted the microbial communities on both regional and continental scale. Ecological factors, such as temperature and nutrient concentrations, are well accepted factors structuring plankton communities.

Beyond the high plankton diversity and the associated highly specific community composition in distinct lakes, the plankton community composition revealed signals of the past, i.e. since the last glaciation some 12,000 years ago.

While this expedition yielded many new scientific findings, the scientists note that these are only the first results of this continental survey.

“We are well aware that we have only just begun our exploration of the hidden diversity of plankton diversity,” they conclude.

###

Original source:

Boenigk J, Wodniok S, Bock C, Beisser D, Hempel C, Grossmann L, Lange A, Jensen M (2018) Geographic distance and mountain ranges structure freshwater protist communities on a European scale. Metabarcoding and Metagenomics 2: e21519. https://doi.org/10.3897/mbmg.2.21519

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

The new innovative academic journal makes use of the one-of-a-kind publishing platform ARPHA and its collaborative writing tool via the ARPHA-XML workflow

A new innovative open-access academic journal Metabarcoding and Metagenomics (MBMG) is launched to welcome novel papers from both basic and applied aspects.

Focusing on genetic approaches to study biodiversity across all ecosystems, MBMG covers a considerably large scope of research including environmental, microbial and applied metabarcoding and metagenomics (especially DNA-based bioassessment and -monitoring, quarantine, nature conservation, species invasions, eDNA surveillance), as well as associated topics, such as molecular ecology, DNA-based species delimitation and identification, and other emerging related fields. Submissions of bioinformatic approaches to MBMG (algorithms, software) are also encouraged.

Featuring novel article formats and data publishing workflows, MBMG is to reflect the rapid growth in the use of metabarcoding and metagenomics in life and environmental sciences.

Issued via ARPHA – the first ever publishing platform to support manuscripts all the way from authoring to peer review to publication and dissemination, designed by the academic publisher and technology provider Pensoft, the new journal is to host a wide range of outcomes from across the research cycle, including data, models, methods, workflows, software, perspectives, opinions, implementation strategies, as well as conventional research articles.

While the above-mentioned publication types are already available in other journals published on the ARPHA platform, such as Research Ideas and Outcomes (RIO), Biodiversity Data Journal and One Ecosystem, MBMG provides five extra domain-specific article types, namely: Emerging Technique, Applied Study, DNA Barcode Release, Primer Validation and Probe Validation.

The journal’s articles are to be available in three formats (PDF, XML, HTML) and full of semantic enhancements for better human- and machine-readability and discoverability. The XML-based workflow also ensures that content and data are available for extraction, indexing and re-use immediately after publication.

With Pensoft standing for transparent, reproducible and open science, the authors at MBMG are strongly encouraged to make all data publicly available either within the publication itself, or to link to external repositories. In their turn, the peer reviewers are also suggested to provide public access to their reviews and identities.

In time for the launch, MBMG has already gathered a team of experienced and renowned scientists from across the globe together on its editorial and advisory board.

“I am pleased to introduce the Metabarcoding and Metagenomics journal to the family of Pensoft,” says Prof. Lyubomir Penev, Founder and Managing Director at Pensoft. “With its exhaustive scope and advanced services and concept, I believe it fills fantastically a niche in our strong portfolio of mostly biodiversity- and ecology-themed journals.”

“Metabarcoding and metagenomics approaches are rapidly progressing and revolutionise research and its application alike,” Chief Editor Prof. Florian Leese states. “With the MBMG journal we provide an ideal platform to respond to this rapidly growing field, nucleate the emerging knowledge and stimulate further development.”

The first batch of research papers published in MBMG are now available on their new website.

“MBMG not only complements the range of journals in the field of molecular environmental life sciences, but also stands out as a novel outlet providing several unique features designed to help researchers to prepare for, and professionally deal with, the massive “deluge” of data,” reads the Editorial.

To celebrate the launch, MBMG starts with a tempting offer to potential authors: publishing will be completely free of charge during the beginning stages of the journal.

###

Follow Metabarcoding and Metagenomics on Twitter | Facebook.