scholarly journals Aquatic biofilms can act as natural environmental DNA samplers

2021 ◽  
Vol 4 ◽  
Author(s):  
Sinziana Rivera ◽  
Valentin Vasselon ◽  
Frederic Rimet ◽  
Agnès Bouchez
Keyword(s):  
Water Samples ◽  
Ecological Status ◽  
Fish Communities ◽  
Morphological Characteristics ◽  
Environmental Dna ◽  
Reference Database ◽  
Macroinvertebrate Communities ◽  
Taxonomic Assignment ◽  
Novel Approach ◽  
Single Matrix

Diatoms, macroinvertebrates and fish communities are widely used for the assessment of the ecological status of rivers and lakes. Metabarcoding studies of these communities are usually performed from “bulk” samples in the case of diatoms and macroinvertebrates; and from water samples in the case of fish. Recent studies, suggest that aquatic biofilms can physically act as environmental catchers of environmental DNA (eDNA) (e.g. Mariani et al. 2019). Thus, we propose an alternative metabarcoding approach to study macroinvertebrates and fishes directly from this matrix. The capacity of aquatic biofilms to catch macroinvertebrate eDNA was tested from a previous study in Mayotte Island were both biofilm samples and macroinvertebrate morphological inventories were available at same river sites (Rivera et al. 2021). First, macroinvertebrate specimens were identified based on their morphological characteristics. Second, DNA was extracted from biofilms, and macroinvertebrate communities were targeted using a standard COI barcode. The resulting morphological and molecular inventories were compared. Our results showed that both methods provided comparable structures and diversities for macroinvertebrate communities when using unassigned OTUs suggesting that macroinvertebrate DNA is present in biofilms and representative of the communities. However, after taxonomic assignment of OTUs, diversity and richness were no longer correlated. Indeed, many constraints were observed as the need for: a) more specific primers to avoid co-amplification of untargeted taxa inhabiting biofilms, b) primers targeting shorter barcodes to sequence more easily degraded eDNA that may be captured in biofilms, and c) a reference database well adapted to our tropical study sites. Finally, even if the results of this first study were encouraging, we wanted to test the biofilm approach on organisms that do not inhabit this environmental matrix in order to be able to distinguish between intra or extra-cellular DNA. Based on these observations, a second study looking for a fish eDNA signal in aquatic biofilms was performed. Environmental biofilm and water samples were collected in parallel at littoral sites at Lake Geneva. DNA was extracted from these samples, and fish communities were targeted using a standard 12S barcode. The molecular inventories derived from the biofilm and the water samples were compared. Both methods provide comparable floristic lists, providing a novel approach for ecological studies related to fish phenology using eDNA in biofilms. Our results open the door to the study of diatoms, macroinvertebrates and fish communities through metabarcoding from a single matrix reducing sampling efforts and costs.

scholarly journals A large-scale ecological assessment of Swiss rivers using environmental DNA for the monitoring of macroinvertebrates 

2021 ◽  
Vol 4 ◽  
Author(s):  
Jeanine Brantschen ◽  
Rosetta Blackman ◽  
Jean-Claude Walser ◽  
Florian Altermatt
Keyword(s):  
Random Forest ◽  
Community Composition ◽  
Large Scale ◽  
Anthropogenic Activities ◽  
Ecological Status ◽  
Environmental Dna ◽  
Biotic Index ◽  
Reference Database ◽  
Macroinvertebrate Communities ◽  
Monitoring Methods

Anthropogenic activities are changing the state of ecosystems worldwide, affecting community composition and often resulting in loss of biodiversity. Riverine ecosystems are among the most impacted ecosystems. Recording their current state with regular biomonitoring is important to assess the future trajectory of biodiversity. However, traditional monitoring methods for ecological assessments are costly and time-intense. Here, we compare environmental DNA (eDNA) to traditional kick-net sampling in a standardized framework of surface water quality assessment. We use surveys of macroinvertebrate communities to assess biodiversity and the biological state of riverine systems. Both methods were employed to monitor aquatic macroinvertebrate indicator groups at 92 sites across major Swiss river catchments. The eDNA data were taxonomically assigned using a customised reference database. All zero-radius Operational Taxonomic Units (zOTUs) mapping to one of the 142 traditionally used indicator taxon levels were used for subsequent diversity analyses (n = 205). At the site level, eDNA detected less indicator taxa than the kick-net method and alpha diversity correlated only weakly between the methods. However, the methods showed a strong congruence in the overall community composition (gamma diversity), as the same indicator groups were commonly detected. In order to set the community composition in relation to the biotic index, the ecological states of the sampling sites were predicted by a random forest approach. Using all zOTUs mapping to macroinvertebrate indicator groups (n = 693) as predictive features, the random forest models successfully predicted the ecological status of the sampled sites. The majority of the predictions (71%) resulted in the same classification like the kick-net based scores. Thus, the sampling of eDNA enabled the detection of indicator communities and provided valuable classifications of the ecological state, when combined with machine learning. Overall, eDNA based sampling has the potential to complement traditional surveys of macroinvertebrate communities in routine large-scale assessments in a non-invasive and scalable approach.

scholarly journals Environmental DNA (eDNA) metabarcoding surveys extend the range of invasion for non-indigenous freshwater species in Eastern Europe.

2021 ◽  
Author(s):  
Gert-Jan Jeunen ◽  
Tatsiana Lipinskaya ◽  
Helen Gajduchenko ◽  
Viktoriya Golovenchik ◽  
Michail Moroz ◽  
...  
Keyword(s):  
Surface Water ◽  
Water Samples ◽  
False Negative ◽  
Simultaneous Detection ◽  
Environmental Dna ◽  
Reference Database ◽  
Indigenous Species ◽  
Freshwater Species ◽  
Traditional Approaches ◽  
Surface Water Samples

Active environmental DNA (eDNA) surveillance through species-specific amplification has shown increased sensitivity in the detection of non-indigenous species (NIS) compared to traditional approaches. When many NIS are of interest, however, active surveillance decreases in cost- and time-efficiency. Passive surveillance through eDNA metabarcoding takes advantage of the complex DNA signal in environmental samples and facilitates the simultaneous detection of multiple species. While passive eDNA surveillance has previously detected NIS, comparative studies are essential to determine the ability of eDNA metabarcoding to accurately describe the range of invasion for multiple NIS versus alternative approaches. Here, we surveyed twelve sites, covering nine rivers across Belarus for NIS with three different techniques, i.e., an ichthyological, hydrobiological, and eDNA survey, whereby DNA was extracted from 500 mL surface water samples and amplified with two 16S rRNA primer assays targeting the fish and macro-invertebrate biodiversity. Nine non-indigenous fish and ten non-indigenous sediment-living macro-invertebrates were detected by traditional surveys, while seven NIS eDNA signals were picked up, including four fish, one aquatic and two sediment-living macro-invertebrates. Passive eDNA surveillance extended the range of invasion further north for two invasive fish and identified a new NIS for Belarus, the freshwater jellyfish Craspedacusta sowerbii. False-negative detections for the eDNA survey could be attributed to (i) preferential amplification of aquatic over sediment-living macro-invertebrates from surface water samples and (ii) an incomplete reference database. The evidence provided in this study recommends the implementation of both molecular-based and traditional approaches to maximize the probability of early detection of non-native organisms.

scholarly journals Environmental DNA enables detection of terrestrial mammals from forest pond water

2016 ◽  
Author(s):  
Masayuki Ushio ◽  
Hisato Fukuda ◽  
Toshiki Inoue ◽  
Kobayashi Makoto ◽  
Osamu Kishida ◽  
...  
Keyword(s):  
Water Samples ◽  
Cervus Nippon ◽  
Environmental Dna ◽  
Terrestrial Ecosystems ◽  
Illumina Miseq ◽  
Pcr Primers ◽  
Pond Water ◽  
Universal Primers ◽  
Taxonomic Assignment ◽  
Terrestrial Mammals

Terrestrial animals must have frequent contact with water to maintain their lives, implying that environmental DNA (eDNA) originating from terrestrial animals should be detectable from places containing water in terrestrial ecosystems. Aiming to detect the presence of terrestrial mammals using forest water samples, we applied a set of universal PCR primers (MiMammal, a modified version of fish universal primers) for metabarcoding mammalian eDNA. After verifying the primers’ usefulness in silico and using water samples from zoo cages of animals with known species compositions, we collected five 500-ml water samples from ponds in two cool-temperate forests in Hokkaido, northern Japan. Using eDNA extracted from the water samples, we constructed amplicon libraries using MiMammal primers for Illumina MiSeq sequencing. MiMammal metabarcoding yielded a total of 75,214 reads, which we then subjected to data pre-processing and taxonomic assignment. We thereby detected species of mammals common to the sampling areas, including deer (Cervus nippon), mouse (Mus musculus), vole (Myodes rufocanus), raccoon (Procyon lotor), rat (Rattus norvegicus) and shrew (Sorex unguiculatus). Previous applications of the eDNA metabarcoding approach have mostly been limited to aquatic/semiaquatic systems, but the results presented here show that the approach is also promising even in forest mammal biodiversity surveys.

scholarly journals PEMA v2: addressing metabarcoding bioinformatics analysis challenges

2021 ◽  
Vol 4 ◽  
Author(s):  
Haris Zafeiropoulos ◽  
Christina Pavloudi ◽  
Evangelos Pafilis
Keyword(s):  
High Performance ◽  
Bioinformatics Analysis ◽  
Marker Gene ◽  
Environmental Dna ◽  
Third Party ◽  
Reference Database ◽  
Marker Genes ◽  
Specific Reference ◽  
Taxonomic Assignment ◽  
Internal Joint

Environmental DNA (eDNA) and metabarcoding have launched a new era in bio- and eco-assessment over the last years (Ruppert et al. 2019). The simultaneous identification, at the lowest taxonomic level possible, of a mixture of taxa from a great range of samples is now feasible; thus, the number of eDNA metabarcoding studies has increased radically (Deiner and 2017). While the experimental part of eDNA metabarcoding can be rather challenging depending on the special characteristics of the different studies, computational issues are considered to be its major bottlenecks. Among the latter, the bioinformatics analysis of metabarcoding data and especially the taxonomy assignment of the sequences are fundamental challenges. Many steps are required to obtain taxonomically assigned matrices from raw data. For most of these, a plethora of tools are available. However, each tool's execution parameters need to be tailored to reflect each experiment's idiosyncrasy; thus, tuning bioinformatics analysis has proved itself fundamental (Kamenova 2020). The computation capacity of high-performance computing systems (HPC) is frequently required for such analyses. On top of that, the non perfect completeness and correctness of the reference taxonomy databases is another important issue (Loos et al. 2020). Based on third-party tools, we have developed the Pipeline for Environmental Metabarcoding Analysis (PEMA), a HPC-centered, containerized assembly of key metabarcoding analysis tools. PEMA combines state-of-the art technologies and algorithms with an easy to get-set-use framework, allowing researchers to tune thoroughly each study thanks to roll-back checkpoints and on-demand partial pipeline execution features (Zafeiropoulos 2020). Once PEMA was released, there were two main pitfalls soon to be highlighted by users. PEMA supported 4 marker genes and was bounded by specific reference databases. In this new version of PEMA the analysis of any marker gene is now available since a new feature was added, allowing classifiers to train a user-provided reference database and use it for taxonomic assignment. Fig. 1 shows the taxonomy assignment related PEMA modules; all those out of the dashed box have been developed for this new PEMA release. As shown, the RDPClassifier has been trained with Midori reference 2 and has been added as an option, classifying not only metazoans but sequences from all taxonomic groups of Eukaryotes for the case of the COI marker gene. A PEMA documentation site is now also available. PEMA.v2 containers are available via the DockerHub and SingularityHub as well as through the Elixir Greece AAI Service. It has also been selected to be part of the LifeWatch ERIC Internal Joint Initiative for the analysis of ARMS data and soon will be available through the Tesseract VRE.

scholarly journals Development of an environmental DNA method for monitoring fish communities: ground truthing in diverse lakes with characterised fish faunas

2018 ◽  
Author(s):  
Jianlong Li ◽  
Tristan W. Hatton-Ellis ◽  
Lori-Jayne Lawson Handley ◽  
Helen S. Kimbell ◽  
Marco Benucci ◽  
...  
Keyword(s):  
Water Samples ◽  
Fish Communities ◽  
Environmental Dna ◽  
Quantitative Information ◽  
12S Rrna ◽  
Similar Species ◽  
List Type ◽  
Large Lakes ◽  
Non Invasive ◽  
Wide Range

AbstractAccurate, cost-effective monitoring of fish is required to assess the quality of lakes under the European Water Framework Directive (WFD). Recent studies have shown that environmental DNA (eDNA) metabarcoding is an effective and non-invasive method, which can provide semi-quantitative information on fish communities in large lakes.This study further investigated the potential of eDNA metabarcoding as a tool for WFD status assessment by collecting and analysing water samples from eight Welsh lakes and six meres in Cheshire, England, with well described fish faunas. Water samples (N = 252) were assayed using two mitochondrial DNA regions (Cytb and 12S rRNA).eDNA sampling indicated the presence of very similar species in the lakes compared to those expected on the basis of existing and historical information. In total, 24 species were detected with a total of 111 species occurrences in the lakes studied using eDNA. Secondly, there was a significant positive correlation between expected faunas and eDNA data in terms of confidence of species occurrence (Spearman’s r = 0.74, df = 109, p <; 0.001). Thirdly, eDNA data can estimate relative abundance with the standard five-level classification scale (“DAFOR”). Lastly, four ecological fish communities were characterised using eDNA data which agrees with the pre-defined lake types according to environmental characteristics.Synthesis and applications. This study provides further evidence that eDNA metabarcoding could be a powerful and non-invasive monitoring tool for WFD purpose in a wide range of lake types, considerably outperforming other methods for community level analysis.

scholarly journals Assessing accuracy and completeness of GenBank for eDNA metabarcoding: towards a reliable marine fish reference database

2021 ◽  
Vol 4 ◽  
Author(s):  
Cristina Claver ◽  
Oriol Canals ◽  
Naiara Rodriguez-Ezpeleta
Keyword(s):  
Ribosomal Rna ◽  
Gap Analysis ◽  
Environmental Dna ◽  
Reference Database ◽  
Fish Diversity ◽  
Cyt B ◽  
Taxonomic Assignment ◽  
Reference Databases ◽  
Taxonomic Groups ◽  
High Level

Environmental DNA (eDNA) metabarcoding, the process of sequencing DNA collected from the environment for producing biodiversity inventories, is increasingly being applied to assess fish diversity and distribution in marine environments. Yet, the successful application of this technique deeply relies on accurate and complete reference databases used for taxonomic assignment. The most used markers for fish eDNA metabarcoding studies are the cytochrome C oxidase subunit 1 (COI), 16S ribosomal RNA (16S), the 12S ribosomal RNA (12S) and cytochrome b (cyt b) genes, whose sequences are usually retrieved from GenBank, the largest DNA sequence database that represents a worldwide public resource for genetic studies. Thus, the completeness and accuracy of GenBank is critical to derive reliable estimations from fish eDNA metabarcoding data. Here, we have i) compiled the checklist of European marine fishes, ii) performed a gap analysis of the four genes and, within COI and 12S, also of the most used barcodes for fish, and iii) developed a workflow to detect potentially incorrect records in GenBank. We found that from the 1965 species in the checklist (1761 Actinopterygii, 189 Elasmobranchii, 9 Holocephali, 4 Petromyzonti and 2 Myxini), about 70% have sequences for COI, whereas less have sequences for 12S, 16S and cyt b (45-55%). Among the species for which COI ad 12S sequences are available, about 60% and 40% have sequences covering the most used barcodes respectively. The analysis of pairwise distances between sequences revealed pairs belonging to the same species with significantly low similarity and pairs belonging to different high level taxonomic groups (class, order) with significantly large similarity. In light of this further confirmation of presence of a substantial number of incorrect records in GenBank, we propose a method for identifying and removing spurious sequences to create reliable and accurate reference databases for eDNA metabarcoding.

scholarly journals Environmental DNA metabarcoding uncovers environmental correlates of fish communities in spatially heterogeneous freshwater habitats

2021 ◽  
Vol 126 ◽  
pp. 107698
Author(s):  
Petr Blabolil ◽  
Lynsey R. Harper ◽  
Štěpánka Říčanová ◽  
Graham Sellers ◽  
Cristina Di Muri ◽  
...  
Keyword(s):  
Fish Communities ◽  
Environmental Dna ◽  
Environmental Correlates ◽  
Dna Metabarcoding ◽  
Freshwater Habitats

scholarly journals Environmental DNA diffusion of reef fishes along a distance gradient from four isolated islands of the Western Indian Ocean

2021 ◽  
Vol 4 ◽  
Author(s):  
Mélissa Jaquier ◽  
Camille Albouy ◽  
Wilhelmine Bach ◽  
Conor Waldock ◽  
Viriginie Marques ◽  
...  
Keyword(s):  
Distance Sampling ◽  
Environmental Dna ◽  
Western Indian Ocean ◽  
Reef Fishes ◽  
Species Distribution Modelling ◽  
Model Systems ◽  
Reference Database ◽  
Developmental Phase ◽  
Marine Environments ◽  
Habitat Types

Islands have traditionally served as model systems to study ecological and evolutionary processes (Warren et al. 2015) and could also represent a relevant system to study environmental DNA (eDNA). Isolated island reefs that are affected by climatic threats would particularly benefit from cost- and time-efficient biodiversity surveys to set priorities for their conservation. Among time efficiency methods, eDNA has emerged as a novel molecular metabarcoding technique to detect biodiversity from simple environmental samples even in remote marine environments. However, eDNA monitoring techniques for marine environments are at a developmental phase, with a few remaining unknowns related to DNA residence time and movement. In particular, the redistribution of eDNA, via ocean currents, could blur the composition signal and its association with local environmental conditions (Goldberg et al. 2016). Here, we investigated the detection variation of eDNA along a distance gradient across four islands in the French Scattered Islands. We collected 30 L of surface water per filter at an increasing distance from the islands reefs (0m, 250m, 500m, 750m). Using a metabarcoding protocol, we used the teleo primers to target a fraction of 12S mitochondrial DNA to detect Actinopterygii and Elasmobranchii. We then applied a sequence clustering approach to generate Molecular Taxonomic Units (MOTUs), which were assigned to a taxonomic group using a reference database. By assigning eDNA sequences to species using a public reference database, we classified species according to their preferred habitat types between benthic/demersal and pelagic. Our results show no significant relationship between distance and MOTUs richness for both habitat types. By using a Joint Species Distribution Modelling approach (JSDM, Hierarchical Modelling of Species Communities), we retained the multidimensional information captured by eDNA and detect species- and family-specific responses to distance (Fig. 1). We showed that benthic MOTUs were found in closer proximity to the reef, while typical pelagic MOTUs were found at greater distances from the reef. Hence, MOTU-level analyses coupled with JSDM were more informative that when aggregating it into coarser richness. Altogether, our eDNA distance sampling gradient detected an ecological signal of habitat selection by fish species, which suggest that eDNA could help understand the behavior of species and their distribution in marine environments at a fine spatial scale.

scholarly journals Overcoming limitations to environmental DNA studies: A coastal temperate reference sequence database for multiple chloroplast gene regions generated in a single assay.

2021 ◽  
Author(s):  
Nicole Foster ◽  
Kor-jent Dijk ◽  
Ed Biffin ◽  
Jennifer Young ◽  
Vicki Thomson ◽  
...  
Keyword(s):  
Dna Sequences ◽  
Dna Barcode ◽  
Environmental Dna ◽  
Reference Sequence ◽  
Reference Database ◽  
Chloroplast Gene ◽  
Coastal Plants ◽  
Reference Databases ◽  
Targeted Capture ◽  
Comprehensive Reference

A proliferation in environmental DNA (eDNA) research has increased the reliance on reference sequence databases to assign unknown DNA sequences to known taxa. Without comprehensive reference databases, DNA extracted from environmental samples cannot be correctly assigned to taxa, limiting the use of this genetic information to identify organisms in unknown sample mixtures. For animals, standard metabarcoding practices involve amplification of the mitochondrial Cytochrome-c oxidase subunit 1 (CO1) region, which is a universally amplifyable region across majority of animal taxa. This region, however, does not work well as a DNA barcode for plants and fungi, and there is no similar universal single barcode locus that has the same species resolution. Therefore, generating reference sequences has been more difficult and several loci have been suggested to be used in parallel to get to species identification. For this reason, we developed a multi-gene targeted capture approach to generate reference DNA sequences for plant taxa across 20 target chloroplast gene regions in a single assay. We successfully compiled a reference database for 93 temperate coastal plants including seagrasses, mangroves, and saltmarshes/samphire’s. We demonstrate the importance of a comprehensive reference database to prevent species going undetected in eDNA studies. We also investigate how using multiple chloroplast gene regions impacts the ability to discriminate between taxa.

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