Environmental DNA Samples Research Articles

Environmental DNA metabarcoding as a tool for biodiversity assessment and monitoring: reconstructing established fish communities of north‐temperate lakes and rivers

AbstractAimTo evaluate the ability of precipitation‐based environmental DNA (eDNA) sample collection and mitochondrial 12S metabarcoding sequencing to reconstruct well‐studied fish communities in lakes and rivers. Specific objectives were to 1) determine correlations between eDNA species detections and known community composition based on conventional field sampling, 2) compare efficiency of eDNA to detect fish biodiversity among systems with variable morphologies and trophic states, and 3) determine if species habitat preferences predict eDNA detection.LocationUpper Great Lakes Region, North America.MethodsFish community composition was estimated for seven lakes and two Mississippi River navigation pools using sequence data from the mitochondrial 12S gene amplified from 10 to 50 water samples per waterbody collected in 50‐mL centrifuge tubes at a single time point. Environmental DNA (eDNA) was concentrated without filtration by centrifuging samples to reduce per‐sample handling time. Taxonomic detections from eDNA were compared to established community monitoring databases containing up to 40 years of sampling and a detailed habitat/substrate preference matrix to identify patterns of bias.ResultsMitochondrial 12S gene metabarcoding detected 15%–47% of the known species at each waterbody and 30%–76% of known genera. Non‐metric multidimensional scaling (NMDS) assessment of the community structure indicated that eDNA‐detected communities grouped in a similar pattern as known communities. Discriminant analysis of principal components indicated that there was a high degree of overlap in habitat/substrate preference of eDNA‐detected and eDNA‐undetected species suggesting limited habitat bias for eDNA sampling.Main conclusionsLarge numbers of small volume samples sequenced at the mitochondrial 12S gene can describe the coarse community structure of freshwater systems. However, additional conventional sampling and environmental DNA sampling may be necessary for a complete diversity census.

The Elusive Delta Hydrobe (Probythinella emarginata) in Kansas

The delta hydrobe (Caenogastropoda-Rissooidea: Hydrobiidae: Probythinella emarginata) was first collected in Kansas in 2001. Previously it was only known from the Pleistocene fossil record. Three live individuals were collected in Cedar Creek, Chase County during Kansas Department of Health and Environment macroinvertebrate surveys in June 2001. Subsequent surveys throughout Cedar Creek in 2010 yielded poor results and only two live individuals were collected. Surveys throughout selected streams in southeastern Kansas in 2011 yielded only one additional population in the Elk River, Montgomery County, Kansas. We resampled and re-evaluated the Cedar Creek and Elk River delta hydrobe populations in May through August 2020 using the same approach as previous surveys. No live individuals were collected. We searched additional perennial streams throughout the Verdigris River basin for the delta hydrobe. We collected few weathered shells but no live individuals were observed. We provide a comprehensive summary of all delta hydrobe shell collections in Kansas. This Pleistocene relic is elusive and likely imperiled in the state. Future surveys should use environmental DNA sampling approaches to more efficiently survey for presence of the delta hydrobe.

Investigating variability in microbial community composition in replicate environmental DNA samples down lake sediment cores.

Lake sediments are natural archives that accumulate information on biological communities and their surrounding catchments. Paleolimnology has traditionally focussed on identifying fossilized organisms to reconstruct past environments. In the last decade, the application of molecular methodologies has increased in paleolimnological studies, but further research investigating factors such as sample heterogeneity and DNA degradation are required. In the present study we investigated bacterial community heterogeneity (16S rRNA metabarcoding) within depth slices (1-cm width). Sediment cores were collected from three lakes with differing sediment compositions. Samples were collected from a variety of depths which represent a period of time of approximately 1,200 years. Triplicate samples were collected from each depth slice and bacterial 16S rRNA metabarcoding was undertaken on each sample. Accumulation curves demonstrated that except for the deepest (oldest) slices, the combination of three replicate samples were insufficient to characterise the entire bacterial diversity. However, shared Amplicon Sequence Variants (ASVs) accounted for the majority of the reads in each depth slice (max. shared proportional read abundance 96%, 86%, 65% in the three lakes). Replicates within a depth slice generally clustered together in the Non-metric multidimensional scaling analysis. There was high community dissimilarity in older sediment in one of the cores, which was likely due to the laminae in the sediment core not being horizontal. Given that most paleolimnology studies explore broad scale shifts in community structure rather than seeking to identify rare species, this study demonstrates that a single sample is adequate to characterise shifts in dominant bacterial ASVs.

PacBio sequencing of Glomeromycota rDNA: a novel amplicon covering all widely used ribosomal barcoding regions and its applicability in taxonomy and ecology of arbuscular mycorrhizal fungi

There is no consensus barcoding region for determination of arbuscular mycorrhizal fungal (AMF) taxa. To overcome this obstacle, we have developed an approach to sequence an AMF marker within the ribosome-encoding operon (rDNA) that covers all three widely applied variable molecular markers. Using a nested PCR approach specific to AMF, we amplified a part (c.2.5kb) of the rDNA spanning the majority of the small subunit rRNA (SSU) gene, the complete internal transcribed spacer (ITS) region and a part of the large subunit (LSU) rRNA gene. The PCR products were sequenced on the PacBio platform utilizing Single Molecule Real Time (SMRT) sequencing. Employing this method for selected environmental DNA samples, we were able to describe complex AMF communities consisting of various glomeromycotan lineages. We demonstrate the applicability of this new 2.5kb approach to provide robust phylogenetic assignment of AMF lineages without known sequences from pure cultures and to consolidate information about AMF taxon distributions coming from three widely used barcoding regions into one integrative dataset.

Environmental DNA sampling provides new management strategies for vernal pool branchiopods in California.

California’s vernal pools are declining ecosystems that support valuable native plant and animal diversity. Vernal pool branchiopods are particularly at risk from vernal pool habitat loss and conservation efforts have targeted their long-term protection through the establishment of preserves and conservation banks. These conservation strategies require repeated, perpetual monitoring of preserved habitat, which is currently carried out through dip-net surveys and visual identification of specimens. Dip-netting may be destructive and frequently requires some sacrifice of protected species. Environmental DNA offers a new, modern method to monitor many protected freshwater organisms. We designed qPCR-based species-specific assays for four of California’s vernal pool branchiopods: The Vernal Pool Fairy Shrimp Branchinecta lynchi (BRLY), the Midvalley Fairy Shrimp Branchinecta mesovallensis (BRME), and the Conservancy Fairy Shrimp Branchinecta conservatio (BRCO), and the Vernal Pool Tadpole Shrimp Lepidurus packardi (LEPA). We tested these assays using eDNA sampling protocols alongside traditional dip-net surveys to assess their viability as an alternative method to monitor vernal pool branchiopods. Based on occupancy modeling, each of our assays achieved a 95% or higher detection rate when using optimized sampling protocols.

Sucked in by eDNA – a promising tool for complementing riverine assessment of freshwater fish communities in Aotearoa New Zealand

ABSTRACT There is increasing interest in the health of waterways in Aotearoa New Zealand. The National Policy Statement for Freshwater Management 2020 (NPS-FM) places a greater onus on resource managers to report on the state of freshwater ecosystem health, including fish. Routine fish monitoring is time intensive and is principally conducted in ‘wadeable’ stream reaches. In comparison, environmental DNA (eDNA) sampling is a relatively recent and rapid technique that likely detects fish and other vertebrate and invertebrate taxa over a broader spatial scale. In this study, we present data for five wadeable streams comparing diversity derived from multi-year standardised electrofishing with eDNA samples collected from the same reaches. Further, we explore whether the number of eDNA monitoring ‘reads’ for species at a site may provide an approximate (semi-quantitative) indication of their ‘near field’ relative abundance based on one-pass electrofishing captures. Results indicated that while some aspects of the methodology require fine-tuning, eDNA sampling shows substantial promise for complimenting state of the environment (SOE) reporting for describing fish diversity in wadeable streams. The use of aquatic eDNA monitoring to provide a cost-effective indication of broader catchment scale biodiversity (e.g. birds and exotic pests) in wadeable and non-wadeable streams is also discussed.

Detection of Mycobacterium bovis in environmental samples using nested PCR

The detection of Mycobacterium bovis (M. bovis) in environmental samples with precision is imperative to control bovine tuberculosis (bTB) infections at the herd level, as residual M. bovis remains one of the major causes of recurring infections. In this study, a nested PCR method for the detection of M. bovis in environmental samples was applied to identify potential environmental reservoirs of the bacterium. A set of 200 environmental samples (167 fecal samples and 33 water samples) from 39 herds with a history of bTB outbreak was analyzed using a nested PCR method to detect residual M. bovis. Amplicon libraries of the IS6110 target gene fragment were amplified from M. bovis DNA using two established primer sets. A positive nested PCR result was observed in 69.5% of fecal samples and 66.7% of water samples, thus showing that residual M. bovis was present in the environmental samples of bTB-positive herds in a high proportion. This study is the first to demonstrate high levels of M. bovis DNA in environmental samples and to show that environmental reservoirs of this pathogen contribute to recurring outbreaks of bTB. Environmental monitoring of herds in which bTB outbreaks have occurred with high sensitivity and specificity is expected to help prevent the recurrence of potential bTB disease and improve the herd environment.

Integration of microfluidic sample preparation with PCR detection to investigate the effects of simultaneous DNA-Inhibitor separation and DNA solution exchange

In this paper, we applied a curved-channel microfluidic device to separate DNA from PCR-inhibitor-containing water and simultaneously wash them into clean water for detection using a portable PCR thermocycler. Environmental DNA (eDNA) sampling has become an effective surveying approach for detecting rare organisms. However, low concentration eDNA molecules may be masked by PCR inhibitors during amplification and detection, increasing the risk of false negatives. Therefore, technologies for on-site DNA separation and washing are urgently needed. Our device consisted of a half-circle microchannel with a DNA-inhibitor sample inlet, a clean buffer inlet, and multiple outlets. By using the flow-induced inertial forces, 10 μm DNA-conjugated microparticles were focused at the inner-wall of the curved microchannel while separation from 1 μm inhibitor-conjugated microparticles and DNA washing were achieved simultaneously with the Dean flow. We achieved singleplex focusing, isolation and washing of 10 μm particles at an efficiency of 94.5 ± 2.0%. In duplex experiments with 1 μm and 10 μm particles, larger particles were washed with an efficiency of 92.1 ± 1.6% and a purity of 79 ± 2%. By surface-functionalizing the microparticles with affinity groups against Atlantic salmon DNA and humic acid (HA), and processing samples of various concentrations in our device, we achieved an effective purification and detection of DNA molecules using the portable PCR thermocycler. Our method significantly decreased PCR quantitation cycles from Cq > 38 to Cq = 30.35 ± 0.5, which confirmed enhancement of PCR amplification. The proposed device takes a promising step forward in sample preparation towards an integrated device that can be used for simultaneous purification and solution exchange of DNA in point-of-need environmental monitoring applications.

Temporal and Spatial Analysis of Forward and Backward Microbial Contamination in a Mars Analog Mission

As human exploration missions to Mars are on the horizon, microbial cross-contamination remains a key issue to address. These issues can be approached today using advances in molecular metagenomics methods, which include rapid and sensitive sequencing platforms for characterizing microbial populations. Combined with analog missions, these methods provide powerful tools for assessing the challenges associated with planetary exploration. Here, we designed a protocol to monitor forward and backward contamination events and progression in an 11-days Mars analog mission in the Ramon crater in Israel. Forward contamination soil samples were collected daily from three sites–two sites in close proximity to the habitat and one isolated site. Backward contamination was determined in samples from nitrile gloves of six analog astronauts before and after extravehicular activities Temperature, relative humidity and soil composition data were also collected for all sites. Environmental DNA samples were extracted in the main habitat and 16S (bacterial) and 18S (eukaryotic, fungal) rRNA gene amplicons were sequenced and analyzed to study microbial population diversity and composition. Shannon Diversity index analysis and Principal Coordinates analysis (PCoA) of rRNA genes indicated that differences in the diversity and population composition were significant in sites closer to the habitat when compared to a reference site. These samples also demonstrated the introduction of human-associated taxa to the environment. Backward contamination consisted of bacterial taxa found on gloves upon return from EVA and also detected in soil, altogether 44 genera, indicating backward contamination events. To our knowledge, this is the first protocol to utilize advanced molecular technologies to investigate forward and backward contamination in a Mars analog mission.

Environmental DNA reveals links between abundance and composition of airborne grass pollen and respiratory health

Grass (Poaceae) pollen is the most important outdoor aeroallergen,1 exacerbating a range of respiratory conditions, including allergic asthma and rhinitis ("hay fever").2-5 Understanding the relationships between respiratory diseases and airborne grass pollen with a view to improving forecasting has broad public health and socioeconomic relevance. It is estimated that there are over 400 million people with allergic rhinitis6 and over 300 million with asthma, globally,7 often comorbidly.8 In the UK, allergic asthma has an annual cost of around US$ 2.8 billion (2017).9 The relative contributions of the >11,000 (worldwide) grass species (C. Osborne etal., 2011, Botany Conference, abstract) to respiratory health have been unresolved,10 as grass pollen cannot be readily discriminated using standard microscopy.11 Instead, here we used novel environmental DNA (eDNA) sampling and qPCR12-15 to measure the relative abundances of airborne pollen from common grass species during two grass pollen seasons (2016 and 2017) across the UK. We quantitatively demonstrate discrete spatiotemporal patterns in airborne grass pollen assemblages. Using a series of generalized additive models (GAMs), we explore the relationship between the incidences of airborne pollen and severe asthma exacerbations (sub-weekly) and prescribing rates of drugs for respiratory allergies (monthly). Our results indicate that a subset of grass species may have disproportionate influence on these population-scale respiratory health responses during peak grass pollen concentrations. The work demonstrates the need for sensitive and detailed biomonitoring of harmful aeroallergens in order to investigate and mitigate their impacts on human health.

Detection of fish pathogen Saprolegnia parasitica in environmental DNA samples by droplet digital PCR

Oomycetes are fungal-like microorganisms parasitic towards a large number of plant and animal species. Genera from order Saprolegniales, such as Saprolegnia and Aphanomyces, cause devastating infections of freshwater animals. Saprolegnia parasitica is a widely distributed oomycete pathogen that causes saprolegniosis, a disease responsible for significant economic losses in aquaculture, as well as declines of natural populations of fish and other freshwater organisms. Despite its negative impact, no monitoring protocol for S. parasitica has been established to date. Thus, we aimed to develop a droplet digital PCR (ddPCR) assay for the detection and quantification of S. parasitica in environmental DNA samples. Saprolegnia parasitica-specific primers were designed to target internal transcribed spacer region 2 (ITS 2), based on the alignment of ITS sequences of S. parasitica and a range of Saprolegnia spp., as well as other oomycetes. The specificity of primers was tested using genomic DNA of S. parasitica (as positive control) and DNA of non–S. parasitica oomycete isolates, as well as trout/crayfish DNA (as negative control). The primers specifically amplified a segment of the ITS region of oomycete pathogen S. parasitica, while no amplification (i.e. no positive droplets) was obtained for closely related Saprolegnia spp. (e.g. Saprolegnia sp. 1 and S. ferax) and other more distantly related oomycetes. Next, the limit of detection (LOD) of the assay was established by using serial dilutions of the S. parasitica genomic DNA. The determined sensitivity of the assay was high: LOD was 15 fg of pathogen’s genomic DNA per µL of the reaction mixture. Assay performance was further assessed with environmental DNA samples isolated from water from the trout farms and natural environments, as well as (ii) biofilm from the host surface (swab samples). Water samples were collected from 21 different locations in Croatia, while swab samples were collected from S. parasitica host/carrier species: (i) skin and eggs of the rainbow trout (Oncorhynchus mykiss Walbaum, 1792) and brown trout (Salmo trutta Linnaeus, 1758), and (ii) cuticle of signal crayfish (Pacifastacus leniusculus Dana, 1852) and narrow clawed crayfish (Pontastacus leptodactylus Eschscholtz, 1823). Samples were classified into agent levels A0 to A6, depending of the number of S. parasitica ITS copies per ng of total DNA. Saprolegnia parasitica was detected in 76 % of water samples (16/21) and the range of pathogen’s ITS copies in positive samples was between 0.02 and 14 copies/ng of total DNA (agent levels A1 to A3). Regarding the swab samples, S. parasitica load was significantly higher in diseased trout than in those with healthy appearance: 9375 vs 3.28 S. parasitica copies/ng of total swab DNA (average agent level A6 vs. A2, respectively). Despite the fact that none of the sampled crayfish had signs of infection, the pathogen was detected in all tested cuticle swabs. Swabs of P. leniusculus, a known S. parasitica host, had significantly higher S. parasitica load than swabs of P. leptodactylus, S. parasitica carrier: 390 vs 83 S. parasitica copies/ng (agent level A5 vs. A4, respectively). In conclusion, our results demonstrate the applicability of the newly developed ddPCR assay in monitoring and early detection of S. parasitica in aquaculture facilities and natural freshwater environments. This would help in a better understanding of S. parasitica ecology and its effects on the host populations.

RPA-based point-of-need detection assay for the invasive Mediterranean fanworm Sabella spallanzanii

The Mediterranean fanworm Sabella spallanzanii was first sighted in New Zealand in 2009 (most likely introduced via hull fouling) and has spread across multiple coastal locations. The species presents significant risks to ecological, economic, and societal values, and therefore is subject to targeted surveillance in 11 major ports and marinas, that were identified as high-risk marine biosecurity sites. Great effort and financial resources are put into bi-annual diving surveys that include removal of individuals to contain population growth and spread. In that context, sensitive molecular detection techniques gain great interest and are being increasingly tested for the fanworm detection in New Zealand and Australia. However, conventional molecular detection via PCR assays from environmental DNA (eDNA) samples requires specific laboratory resources and scientific expertise, which restricts the applicability of this approach by biosecurity practitioners or citizen scientists. In order to provide end-users with a fast, easy, and highly specific way to detect S. spallanzanii at the site of interest, a recombinase polymerase amplification assay (RPA) was designed for read-out with lateral flow detection. RPA generates amplification within 20 minutes at 39°C, which makes it possible to run assays hand-held. Because of this, RPA presents an excellent tool for the point-of-need application in targeted biosecurity surveillance. Here we present the results of the successful in silico and in vitro validation of the newly designed RPA assay for S. spallanzanii.

Evolution of a high-throughput environmental DNA sampling platform

With the rise of environmental DNA as a surveillance tool for aquatic species, a need has also arisen for professionally engineered research tools specifically designed for eDNA applications. We created the first portable, purpose-built eDNA sampling system in the form of a backpack smart-pump filtration apparatus and custom made eDNA filter packets for each sample. The eDNA-Sampler (previously ANDe) enables both point-location sampling and mobile sampling over a spatial distance, with the ability to standardize filtration parameters (e.g. flow rate, pressure, water volume, etc.). In this presentation we will describe the evolution of the eDNA sampling backpack and associated components, each designed to help streamline the eDNA sampling process and increase sampling efficiency. We have optimized the platform for mobile sampling by integrating GPS and data logging capabilities, in addition to modifying the chemistry of the eDNA filter packets to minimize the effort required for sample preservation. Results will be presented from a series of pilot studies in which the eDNA-Sampler capabilities were evaluated. Combined, the innovations described herein should help remove barriers to entry for potential eDNA practitioners and also improve overall eDNA data quality.

Estimating the genetic diversity of Pacific salmon and trout using multigene eDNA metabarcoding.

Genetic diversity underpins species conservation and management goals, and ultimately determines a species’ ability to adapt. Using freshwater environmental DNA (eDNA) samples, we examined mitochondrial genetic diversity using multigene metabarcode sequence data from four Oncorhynchus species across 16 sites in Oregon and northern California. Our multigene metabarcode panel included targets commonly used in population genetic NADH dehydrogenase 2 (ND2), phylogenetic cytochrome c oxidase subunit 1 (COI) and eDNA (12S ribosomal DNA) screening. The ND2 locus showed the greatest within‐species haplotype diversity for all species, followed by COI and then 12S rDNA for all species except Oncorhynchus kisutch. Sequences recovered for O. clarkii clarkii were either identical to, or one mutation different from, previously characterized haplotypes (95.3% and 4.5% of reads, respectively). The greatest diversity in O. c. clarkii was among coastal watersheds, and subsets of this diversity were shared with fish in inland watersheds. However, coastal streams and the Umpqua River watershed appear to harbour unique haplotypes. Sequences from O. mykiss revealed a disjunction between the Willamette watershed and southern watersheds suggesting divergent histories. We also identified similarities between populations in the northern Deschutes and southern Klamath watersheds, consistent with previously hypothesized connections between the two via inland basins. Oncorhynchus kisutch was only identified in coastal streams and the Klamath River watershed, with most diversity concentrated in the coastal Coquille watershed. Oncorhynchus tshawytscha was only observed at one site, but contained multiple haplotypes at each locus. The characterization of genetic diversity at multiple loci expands the knowledge gained from eDNA sampling and provides crucial information for conservation actions and genetic management.

Improved environmental DNA sampling scheme for Alabama sturgeon provides new insight into a species once presumed extinct

Detection of rare species can be challenging and time-consuming using conventional methods, but environmental DNA (eDNA) is becoming a commonly used tool for detection in conservation and management of species. This study demonstrates the utility of the precipitation method (precipitated and preserved in 3 M sodium acetate and 95% ethanol) for collection of eDNA to detect the seasonal distribution of the critically endangered Alabama sturgeon (Scaphirhynchus suttkusi). Surface and benthic water samples were collected across a wider geographic area than previously published for Alabama sturgeon eDNA. Surface and benthic samples both yielded detections and resulted in a similar proportion of positive detections to previous work. However, by sampling a greater portion of the distribution of the Alabama sturgeon, further insight was provided on potential sturgeon movement. The results of the precipitation method show that Alabama sturgeon detections increase during spawning months, and that the fish may be overwintering in the Tombigbee River. High detections from winter benthic samples suggest that habitat choice may play a role in detectability and highlight the need to consider natural history when designing environmental DNA studies. When designing environmental DNA collection for rare species, sampling design should factor in species ecology, habitat use, site characteristics, and specific questions driving the research.

Strategic considerations for invasive species managers in the utilization of environmental DNA (eDNA): steps for incorporating this powerful surveillance tool.

Invasive species surveillance programs can utilize environmental DNA sampling and analysis to provide information on the presence of invasive species. Wider utilization of eDNA techniques for invasive species surveillance may be warranted. This paper covers topics directed towards invasive species managers and eDNA practitioners working at the intersection of eDNA techniques and invasive species surveillance. It provides background information on the utility of eDNA for invasive species management and points to various examples of its use across federal and international programs. It provides information on 1) why an invasive species manager should consider using eDNA, 2) deciding if eDNA can help with the manager’s surveillance needs, 3) important components to operational implementation, and 4) a high-level overview of the technical steps necessary for eDNA analysis. The goal of this paper is to assist invasive species managers in deciding if, when, and how to use eDNA for surveillance. If eDNA use is elected, the paper provides guidance on steps to ensure a clear understanding of the strengths and limitation of the methods and how results can be best utilized in the context of invasive species surveillance.

The future of biodiversity monitoring and conservation utilizing environmental DNA

The future of biodiversity monitoring and conservation utilizing environmental DNA

Efficacy of environmental DNA sampling to detect the occurrence of endangered coho salmon (Oncorhynchus kisutch) in Mediterranean‐climate streams of California's central coast

AbstractAnalysis of environmental DNA (eDNA) has emerged as an important tool for investigating the occurrence and distribution of fish and other species in aquatic environments. However, in lotic systems, uncertainty remains about how environmental factors influence the downstream transport, degradation, and dilution of eDNA, and hence how detection probability varies with distance from the eDNA source. We conducted cage experiments and paired eDNA and snorkel surveys to evaluate the potential for eDNA methods to detect endangered coho salmon in Mediterranean‐climate streams of the Santa Cruz Mountains, California. Juvenile coho salmon at two densities were placed in cages, and water samples were collected and analyzed (qPCR) at seven locations from 10 to 1,000 m downstream. Although we detected coho salmon eDNA up to 1,000 m downstream of the cage, the vast majority of detections were within 200 m, and detections at greater distances occurred only at the higher fish density. In field collections from 21 stream reaches across the Santa Cruz Mountains, eDNA methods and snorkeling produced identical outcomes in terms of reach‐level detection of coho salmon. Probability of occurrence in a water sample and detection in a qPCR replicate both increased with observed fish density; however, even at the lowest densities, a protocol of three water samples and two qPCR replicates resulted in a >90% cumulative probability of detection. Overall, our studies suggest that detection probability decreased rapidly with distance in Santa Cruz Mountain streams, likely because low stream discharges, slow transport velocities, and strong interaction between the water column and stream substrate result in rapid degradation or settling of eDNA. Our findings thus support the use of eDNA methods for detecting rare species, but also indicate that local conditions strongly influence transport dynamics and hence need to be considered when developing survey designs.

Environmental DNA metabarcoding reveals winners and losers of global change in coastal waters.

Studies of the ecological effects of global change often focus on one or a few species at a time. Consequently, we know relatively little about the changes underway at real-world scales of biological communities, which typically have hundreds or thousands of interacting species. Here, we use COI mtDNA amplicons from monthly samples of environmental DNA to survey 221 planktonic taxa along a gradient of temperature, salinity, dissolved oxygen and carbonate chemistry in nearshore marine habitat. The result is a high-resolution picture of changes in ecological communities using a technique replicable across a wide variety of ecosystems. We estimate community-level differences associated with time, space and environmental variables, and use these results to forecast near-term community changes due to warming and ocean acidification. We find distinct communities in warmer and more acidified conditions, with overall reduced richness in diatom assemblages and increased richness in dinoflagellates. Individual taxa finding more suitable habitat in near-future waters are more taxonomically varied and include the ubiquitous coccolithophore Emiliania huxleyi and the harmful dinoflagellate Alexandrium sp. These results suggest foundational changes for nearshore food webs under near-future conditions.

The Taxon Hypothesis Paradigm-On the Unambiguous Detection and Communication of Taxa.

Here, we describe the taxon hypothesis (TH) paradigm, which covers the construction, identification, and communication of taxa as datasets. Defining taxa as datasets of individuals and their traits will make taxon identification and most importantly communication of taxa precise and reproducible. This will allow datasets with standardized and atomized traits to be used digitally in identification pipelines and communicated through persistent identifiers. Such datasets are particularly useful in the context of formally undescribed or even physically undiscovered species if data such as sequences from samples of environmental DNA (eDNA) are available. Implementing the TH paradigm will to some extent remove the impediment to hastily discover and formally describe all extant species in that the TH paradigm allows discovery and communication of new species and other taxa also in the absence of formal descriptions. The TH datasets can be connected to a taxonomic backbone providing access to the vast information associated with the tree of life. In parallel to the description of the TH paradigm, we demonstrate how it is implemented in the UNITE digital taxon communication system. UNITE TH datasets include rich data on individuals and their rDNA ITS sequences. These datasets are equipped with digital object identifiers (DOI) that serve to fix their identity in our communication. All datasets are also connected to a GBIF taxonomic backbone. Researchers processing their eDNA samples using UNITE datasets will, thus, be able to publish their findings as taxon occurrences in the GBIF data portal. UNITE species hypothesis (species level THs) datasets are increasingly utilized in taxon identification pipelines and even formally undescribed species can be identified and communicated by using UNITE. The TH paradigm seeks to achieve unambiguous, unique, and traceable communication of taxa and their properties at any level of the tree of life. It offers a rapid way to discover and communicate undescribed species in identification pipelines and data portals before they are lost to the sixth mass extinction.