Environmental DNA metabarcoding: Transforming how we survey animal and plant communities

Molecular Ecology, Journal Year: 2017, Volume and Issue: 26(21), P. 5872 - 5895

Published: Sept. 18, 2017

The genomic revolution has fundamentally changed how we survey biodiversity on earth. High-throughput sequencing ("HTS") platforms now enable the rapid of DNA from diverse kinds environmental samples (termed "environmental DNA" or "eDNA"). Coupling HTS with our ability to associate sequences eDNA a taxonomic name is called "eDNA metabarcoding" and offers powerful molecular tool capable noninvasively surveying species richness many ecosystems. Here, review use metabarcoding for animal plant richness, challenges in using approaches estimate relative abundance. We highlight applications freshwater, marine terrestrial environments, this broad context, distill what known about different sample types approximate space across time. provide guiding questions study design discuss workflow focus primers library preparation methods. additionally important criteria consideration bioinformatic filtering data sets, recommendations increasing transparency. Finally, looking future, emerging ecology, conservation, invasion biology, biomonitoring, can empower citizen science education.

Language: Английский

---

The ecology of environmental DNA and implications for conservation genetics

Conservation Genetics, Journal Year: 2015, Volume and Issue: 17(1), P. 1 - 17

Published: Sept. 8, 2015

Environmental DNA (eDNA) refers to the genetic material that can be extracted from bulk environmental samples such as soil, water, and even air. The rapidly expanding study of eDNA has generated unprecedented ability detect species conduct analyses for conservation, management, research, particularly in scenarios where collection whole organisms is impractical or impossible. While number studies demonstrating successful detection increased recent years, less research explored "ecology" eDNA—myriad interactions between extraorganismal its environment—and influence on detection, quantification, analysis, application conservation research. Here, we outline a framework understanding ecology eDNA, including origin, state, transport, fate material. Using this framework, review synthesize findings diverse environments, taxa, fields highlight important concepts knowledge gaps application. Additionally, identify frontiers conservation-focused see most potential growth, use estimating population size, genomic via inclusion other indicator biomolecules RNA proteins, automated sample consideration an expanded array creative samples. We discuss how more complete integral advancing these maximizing future applications

Language: Английский

---

Scrutinizing key steps for reliable metabarcoding of environmental samples

Methods in Ecology and Evolution, Journal Year: 2017, Volume and Issue: 9(1), P. 134 - 147

Published: July 1, 2017

Abstract Metabarcoding of environmental samples has many challenges and limitations that require carefully considered laboratory analysis workflows to ensure reliable results. We explore how decisions regarding study design, set‐up, bioinformatic processing affect the final results, provide guidelines for samples. evaluate performance four primer sets targeting COI 16S regions characterizing arthropod diversity in bat faecal samples, investigate metabarcoding results are affected by parameters including: (1) number PCR replicates per sample, (2) sequencing depth, (3) replicate strategy (i.e. either additively, combining sequences obtained from replicates, or restrictively, only retaining occur multiple each sample), (4) minimum copy be retained, (5) chimera removal, (6) similarity thresholds Operational Taxonomic Unit ( OTU ) clustering. Lastly, we measure within‐ between‐taxa dissimilarities when using public databases determine most appropriate clustering taxonomy assignment. Our show use reduces taxonomic biases increases coverage. profiles resulting set principally carried out sample filtered across them, sequence threshold threshold. also report considerable differences between sample. Sequencing depth dissimilarity unless strategies remove allegedly artefactual adjusted according analysed sequences. Finally, identity assignment differ markers. complex ideally requires investigation whether more than one same group is needed offset biases, balance detection with removal sequences, empirical selection tailored marker taxa.

Language: Английский

---

Counting with DNA in metabarcoding studies: How should we convert sequence reads to dietary data?

Molecular Ecology, Journal Year: 2018, Volume and Issue: 28(2), P. 391 - 406

Published: June 2, 2018

Abstract Advances in DNA sequencing technology have revolutionized the field of molecular analysis trophic interactions, and it is now possible to recover counts food sequences from a wide range dietary samples. But what do these mean? To obtain an accurate estimate consumer's diet should we work strictly with data sets summarizing frequency occurrence different taxa, or use relative number sequences? Both approaches are applied semi‐quantitative summaries, but often promoted as more conservative reliable option due taxa‐specific biases recovery sequences. We explore representative metabarcoding point out that summaries based on overestimate importance consumed small quantities (potentially including low‐level contaminants) sensitive count threshold used define occurrence. Our simulations indicate using read abundance ( RRA ) information provides view population‐level even moderate incorporated; however, impacting common taxa. when mean taxa samples small. The ideas presented here highlight need consider all sources bias justify methods interpret studies. encourage researchers continue addressing methodological challenges acknowledge unanswered questions help spur future investigations this rapidly developing area research.

Language: Английский

---

Ecosystem biomonitoring with eDNA: metabarcoding across the tree of life in a tropical marine environment

Scientific Reports, Journal Year: 2017, Volume and Issue: 7(1)

Published: Sept. 19, 2017

Abstract Effective marine management requires comprehensive data on the status of biodiversity. However, efficient methods that can document biodiversity in our oceans are currently lacking. Environmental DNA (eDNA) sourced from seawater offers a new avenue for investigating biota ecosystems. Here, we investigated potential eDNA to inform breadth present tropical environment. Directly sequencing using shotgun approach resulted only 0.34% 22.3 million reads assigning eukaryotes, highlighting inefficiency this method assessing eukaryotic diversity. In contrast, ‘tree life’ (ToL) metabarcoding and 20-fold fewer reads, could detect 287 families across major divisions eukaryotes. Our also show best performing ‘universal’ PCR assay recovered 44% eukaryotes identified all assays, need multiple assays catalogue Lastly, focusing fish genus Lethrinus , intra- inter-specific haplotypes samples, illustrating be used explore diversity beyond taxon identifications. Given sensitivity low cost advocate rapidly integrated into biomonitoring programs.

Language: Английский

---

Environmental DNA from Seawater Samples Correlate with Trawl Catches of Subarctic, Deepwater Fishes

PLoS ONE, Journal Year: 2016, Volume and Issue: 11(11), P. e0165252 - e0165252

Published: Nov. 16, 2016

Remote polar and deepwater fish faunas are under pressure from ongoing climate change increasing fishing effort. However, these communities difficult to monitor for logistic financial reasons. Currently, monitoring of marine fishes largely relies on invasive techniques such as bottom trawling, official reporting global catches, which can be unreliable. Thus, there is need alternative non-invasive qualitative quantitative oceanic surveys. Here we report environmental DNA (eDNA) metabarcoding seawater samples continental slope depths in Southwest Greenland. We collected at 188-918 m compared eDNA catch data trawling. used Illumina sequencing PCR products demonstrate that reads show equivalence obtained Twenty-six families were found with both trawling eDNA, while three only two Key commercial species Greenland the most abundant biomass catch, interpolation abundances between sampling sites showed good correspondence sizes. Environmental sequence assemblages correlated abundance Interestingly, shark (Somniosus microcephalus) high despite a single specimen being caught, demonstrating relevance approach large probably avoid trawls cases. Quantitative detection using remains tested further ascertain whether this technique able yield credible results routine application fisheries. Nevertheless, our study demonstrates proxy habitats. This relates directly applied fisheries well effects biodiversity-especially ecosystems.

Language: Английский

---

DNA metabarcoding—Need for robust experimental designs to draw sound ecological conclusions

Molecular Ecology, Journal Year: 2019, Volume and Issue: 28(8), P. 1857 - 1862

Published: April 1, 2019

DNA metabarcoding, especially when coupled with high-throughput sequencing, is currently revolutionizing our capacity to assess biodiversity across a full range of taxa and habitats, from soil microbes (e.g., Thompson et al., 2017) large marine fish Thomsen 2016), contemporary tens thousands year-old biological communities Willerslev 2003). The breadth potential applications immense spans surveys on the diversity or diet species native specific ecosystems bioindication (Pawlowski 2018). approach also cost-effective easy implement, which makes metabarcoding one tools choice 21st century for fundamental research future large-scale monitoring programs (reviewed in Bohan 2017; Creer 2016; Taberlet, Bonin, Zinger, & Coissac, 2018; Willerslev, 2015). However, as often case any emerging technology, we feel that rise occurring at pace manner loses sight challenges producing high-quality reproducible data (Baker, 2016). by essence multidisciplinary building upon many complementary expertises, including field theoretical knowledge, taxonomic expertise, molecular biology, bioinformatics, computational statistics. Combining all these within single studies necessary, not so much analyzing per se, but rather minimizing controlling possible biases can be introduced step experimental workflow—i.e., sampling analysis—and lead spurious ecological conclusions Bálint Nilsson 2019; Dickie Taberlet Whether starting material consists bulk samples (community DNA) and/or environmental (eDNA), rely deceptively simple succession core steps: (a) preservation material, (b), extraction, (c) PCR amplification taxonomically-informative genomic region, (d) sequencing amplicons, (e) sequence analysis using bioinformatic pipelines. Despite this apparent simplicity, each potentially introduce its own sources artifacts (Figure 1). For example, design might effective capturing processes under study, an undesired bias based detection. availability governed production rate, transport persistence, are largely dependent targeted organisms, their biomass, ecosystem considered. A correct assessment phenomenon require only implementation standardized standardized, randomized repeatable designs procedures (Dickie 2018), consideration dynamics underlying matrix (i.e., gut, faeces, water matrices tropical boreal organisms/ecosystems; Barnes Turner, Likewise, community study enriched purpose depending how sample collected filter size samples, removal roots soils), it transported/preserved, extracted (differential extraction efficiencies). well known important source biases, now fully revealed techniques. preferential certain over other ones due inappropriate primers provides such example (Clarke, Soubrier, Weyrich, Cooper, 2014; Deagle, Jarman, Pompanon, 2014). Primer both skew abundance profiles false negatives. produce negatives too through presence e.g., inhibitors, positives introduction replication errors polymerase formation chimeric fragments False workflow reagent contaminants (Salter 2014), extractions cross-contaminations. An even more insidious pertains occurrence "tag jumps", sometimes referred "mistagging", "tag-switching", "cross-talks" (Carlsen 2012; Edgar, Esling, Lejzerowicz, Pawlowski, 2015; Schnell, Bohmann, Gilbert, amplicons indeed tagged unique short nucleotide sequences added 5'-end "tags"), allow pooling PCRs run reducing costs. Each obtained then bioinformatically assigned back origin basis tags (Schnell preparation libraries tag particular fact recombined belonging another (Taberlet This introduces additional, non-negligible levels cross-contaminations, primarily involve most abundant have disproportionate impact low concentrations (Esling Murray, Coghlan, Bunce, Schnell Similarly, Illumina index located P5 adaptor subjected "index resulting cross-contaminations happens several individual pooled loaded same lane (Kircher, Sawyer, Meyer, 2012). Finally, instruments error rates (Schirmer above list problems clearly exhaustive, interested reader will find complete reviews elsewhere (e.g. Still, illustrates must considered carefully designing protocol interpreting results. crucial limit downstream analyses, ensure conclusion drawn authentic. There increasingly diverse field, laboratory Caporaso 2011; Valentini 2009) bioinformatics Boyer 2010; Dumbrell, Ferguson, Clark, 2016) aiming amount partial sampling, bias) contaminations, "tag/index errors) experiments. protocols does necessarily guarantee problem completely control. These continuously reconsidered, alongside emergence novel technologies provide new opportunities, challenges. Additionally, marker comes specificities, requires customization protocols. clustering threshold used form Molecular Operational Taxonomic Units relevant question addressed removing intraspecific variability level desired) critically depend specificities PCR/sequencing rates. Bioinformatics further fail exclude filtering thresholds relaxed, inflates estimates. they generate negatives, genuine metabarcode falsely flagged chimera, incorrect taxon incomplete reference databases (Alsos Riaz, Puillandre, problematic investigated strongly relies It therefore include types controls facilitate exclusion signal support reliability Amongst controls, conducting pilot experiments particularly helpful appropriate We recommend replicates multiple independent samples) technical extractions/PCR extract) included disentangle effect variances (Ficetola replications necessary because stochastic manner, concentration target low. essential analyze sufficient number negative PCR, steps, positive consisting mock communities, synthetic reflecting attributes products All sequenced along detection sporadic contaminations jumps while helping adjusting thresholds. Ultimately, token whole curation process (De Barba encourage careful itself, since steps curate study. Typically, given may deriving tag/index jumps. retained thus artifacts, depth. As different direct measurements better tuning considerations should precisely reported publications together illustrations statistics characterizing workflow, relevance quality underpinning conclusions. last, obvious control assessing plausibility composition priori knowledge system studied. Such derived sensing approaches visual observations. In case, exhaustive local specimens secure assignment Alsos When information unavailable, typically studying microorganisms, remains whether composed clades expected occur surveyed not, soils, sediments, gut environments harbour highly bacterial phyla 2017). users, readers, referees editors, realize mentioned issues remain overlooked. stance unsubstantiated claims undermine scientific advances if resolved. Inappropriate practices estimating richness fingerprint (Bent 2007), absence (Prosser, 2010), contaminant (Perez-Muñoz, Arrieta, Ramer-Tait, Walter, been repeatedly criticized microbial ecology, latter contribute rising debate about existence womb microbiota. Ancient has developed rigorous standards tackle related contamination, errors, reproducibility (Poinar 2000). believe users come age learnt past errors. At time guides best subject (Knight Pollock, Glendinning, Wisedchanwet, Watson, where costs rapidly decreasing, always mindful adage "better safe than sorry". note mean imply systematic use highest analytical reasonable nor universal remedy associated metabarcoding. Rather, researchers end-users adopt reflective decision-making experiment appraise results, ultimate aim prove robustness

Language: Английский

---

Annual time-series analysis of aqueous eDNA reveals ecologically relevant dynamics of lake ecosystem biodiversity

Nature Communications, Journal Year: 2017, Volume and Issue: 8(1)

Published: Jan. 18, 2017

Abstract The use of environmental DNA (eDNA) in biodiversity assessments offers a step-change sensitivity, throughput and simultaneous measures ecosystem diversity function. There remains, however, need to examine eDNA persistence the wild through temporal biota. Here, we metabarcoding two markers different lengths, derived from an annual time series aqueous lake shifts ecologically important group macroinvertebrates (Diptera: Chironomidae). analyses allow levels detection validation taxon richness community composition (β-diversity) time, with shorter fragments dominating community. Comparisons between eDNA, DNA, taxonomy UK species abundance data further show significant relationships estimates across disparate methodologies. Our results reveal dynamics validate utility for tracking seasonal at scale.

Language: Английский

---

Next-generation sequencing (NGS) for assessment of microbial water quality: current progress, challenges, and future opportunities

Frontiers in Microbiology, Journal Year: 2015, Volume and Issue: 6

Published: Sept. 25, 2015

Water quality is an emergent property of a complex system comprised interacting microbial populations and introduced chemical contaminants. Studies leveraging next-generation sequencing (NGS) technologies are providing new insights into the ecology microbially mediated processes that influence fresh water such as algal blooms, contaminant biodegradation, pathogen dissemination. In addition, methods targeting small subunit (SSU) rRNA hypervariable regions have allowed identification signature species serve bioindicators for sewage contamination in these environments. Beyond amplicon sequencing, metagenomic metatranscriptomic analyses communities environments reveal genetic capabilities interplay waterborne microorganisms, shedding light on mechanisms production biodegradation toxins other This review discusses challenges benefits applying NGS-based to research assessment. We will consider suitability biases inherent application NGS screening tool assessment biological risks discuss potential limitations direct quantitative interpretation data. Secondly, we examine case studies from recent literature where based been applied topics assessment, including development pollution source tracking, characterizing distribution toxin antibiotic resistance genes samples, investigating harmful pollutants threaten quality. Finally, provide short emerging platforms their applications next generation tools.

Language: Английский

---

Environmental DNA (eDNA) metabarcoding reveals strong discrimination among diverse marine habitats connected by water movement

Molecular Ecology Resources, Journal Year: 2018, Volume and Issue: 19(2), P. 426 - 438

Published: Dec. 21, 2018

While in recent years environmental DNA (eDNA) metabarcoding surveys have shown great promise as an alternative monitoring method, the integration into existing marine programs may be confounded by dispersal of eDNA signal. Currents and tidal influences could transport over distances, inducing false-positive species detection, leading to inaccurate biodiversity assessments and, ultimately, mismanagement environments. In this study, we determined ability distinguish localized signals obtained from four habitats within a small spatial scale (<5 km) subject significant along-shore water flow. Our survey detected 86 genera, 77 families across 11 phyla using three established assays targeting fish (16S rRNA gene), crustacean gene) eukaryotic (cytochrome oxidase subunit 1) diversity. Ordination cluster analyses for both taxonomic OTU data sets show distinct between sampled habitats, suggesting among was limited. Individual taxa with strong habitat preferences displayed accordance their respective habitat, whereas known less habitat-specific generated more ubiquitous signals. add evidence that environments detect broad range are spatially discrete. work also highlights refinement assay choice is essential realize full potential programs.

Language: Английский

---