Best practices in metabarcoding of fungi: From experimental design to results

Molecular Ecology, Journal Year: 2022, Volume and Issue: 31(10), P. 2769 - 2795

Published: April 8, 2022

The development of high-throughput sequencing (HTS) technologies has greatly improved our capacity to identify fungi and unveil their ecological roles across a variety ecosystems. Here we provide an overview current best practices in metabarcoding analysis fungal communities, from experimental design through molecular computational analyses. By reanalysing published data sets, demonstrate that operational taxonomic units (OTUs) outperform amplified sequence variants (ASVs) recovering diversity, finding is particularly evident for long markers. Additionally, the full-length ITS region allows more accurate placement other eukaryotes compared ITS2 subregion. Finally, show specific methods compositional analyses reliable estimates shifts community structure. We conclude are especially promising integrating into full microbiome broader ecosystem functioning context, recovery novel lineages ancient organisms as well barcoding old specimens including type material.

Language: Английский

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Airborne environmental DNA for terrestrial vertebrate community monitoring

Current Biology, Journal Year: 2022, Volume and Issue: 32(3), P. 701 - 707.e5

Published: Jan. 6, 2022

Biodiversity monitoring at the community scale is a critical element of assessing and studying species distributions, ecology, diversity, movements, it key to understanding tracking environmental anthropogenic effects on natural ecosystems.1Navarro L.M. Fernández N. Guerra C. Guralnick R. Kissling W.D. Londoño M.C. Muller-Karger F. Turak E. Balvanera P. Costello M.J. et al.Monitoring biodiversity change through effective global coordination.Curr. Opin. Environ. Sustain. 2017; 29: 158-169Crossref Scopus (89) Google Scholar, 2Xu H. Cao Y. Yu D. M. He Gill Pereira H.M. Ensuring implementation post-2020 targets.Nat. Ecol. Evol. 2021; 5: 411-418Crossref PubMed (30) 3(2021). High time invest in biodiversity. Nat. 5, 263.Google 4Seibold S. Gossner M.M. Simons N.K. Blüthgen Müller J. Ambarli al.Arthropod decline grasslands forests associated with landscape-level drivers.Nature. 2019; 574: 671-674Crossref (391) Scholar Vertebrates terrestrial ecosystems are experiencing extinctions declines both population numbers sizes due increasing threats from human activities change.5Almond R.E.A. Grooten Petersen T. Living Planet Report 2020: Bending Curve Loss. WWF, 2020Google 6Ceballos G. Ehrlich P.R. Dirzo Biological annihilation via ongoing sixth mass extinction signaled by vertebrate losses declines.Proc. Natl. Acad. Sci. USA. 114: E6089-E6096Crossref (1031) 7Hughes J.B. Daily G.C. Population diversity: its extent extinction.Science. 1997; 278: 689-692Crossref (381) 8Gaston K.J. Fuller R.A. Commonness, depletion conservation biology.Trends 2008; 23: 14-19Abstract Full Text PDF (361) Terrestrial using existing methods generally costly laborious, although DNA (eDNA) becoming tool choice assess biodiversity, few sample types effectively capture diversity. We hypothesized that eDNA captured air could allow straightforward collection characterization communities. filtered three localities Copenhagen Zoo: stable, outside between outdoor enclosures, Rainforest House. Through metabarcoding airborne eDNA, we detected 49 spanning 26 orders 37 families: 30 mammal, 13 bird, 4 fish, 1 amphibian, reptile species. These spanned animals kept zoo, occurring zoo surroundings, used as feed zoo. The comprise range taxonomic families, sizes, behaviors, abundances. found shorter distance sampling device higher animal biomass increase probability detection. hereby show can offer fundamentally new way

Language: Английский

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A practical guide to DNA-based methods for biodiversity assessment

Pensoft Publishers eBooks, Journal Year: 2021, Volume and Issue: unknown

Published: Dec. 2, 2021

DNA-based methods for species detection and identification have revolutionised our ability to assess biodiversity in terrestrial, freshwater marine ecosystems. Starting from the seminal study that used eDNA detect invasive american bullfrogs France (Gentile Francesco Ficetola et al. 2008)), research conducted over last decade has demonstrated power of these approaches surveying a wide range groups. Early applications included use monitor Asian Carp USA (Jerde 2013). Following heavy scrutiny, method was eventually adopted, is still employed today by United States Geological Survey (USGS). A flurry followed, with tests designed many threatened including New zealand mudsnails (Goldberg 2013), crayfish (Geerts 2018), gammarids (R. Blackman 2017), great crested newts (Biggs 2015). The newt test been regulatory monitoring UK since 2014. During same time period, there proliferation studies high-throughput sequencing describe whole communities organisms mixed environmental samples, using an approach termed DNA metabarcoding (Taberlet 2012).

Language: Английский

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Airborne environmental DNA metabarcoding for the monitoring of terrestrial insects—A proof of concept from the field

Environmental DNA, Journal Year: 2022, Volume and Issue: 4(4), P. 790 - 807

Published: March 11, 2022

Abstract Biodiversity is in decline due to human‐induced pressures on ecosystems around the world. To be able counteract this alarming trend, it paramount closely monitor biodiversity at global scales. Because practically impossible with traditional methods, last decade has seen a strong push for new solutions. In aquatic ecosystems, monitoring of species from environmental DNA (eDNA) emerged as one most powerful tools our disposal, but terrestrial power eDNA so far been hampered by local scale samples. study, we report successful detection insects airborne samples taken field. We compare results two insect methods (1) light traps moth and (2) transect walks butterflies wild bees. Airborne metabarcoding revealed six classes arthropods, twelve order insects—including representatives four largest orders: Diptera (flies), Lepidoptera (butterflies moths), Coleoptera (beetles), Hymenoptera (bees, wasps, ants). did not detect all observed using suggest further directions development metabarcoding. also recovered nine vertebrates, including frogs, birds, mammals well 12 other phyla. potential become tool monitoring, many impactful applications pests, invasive, or endangered disease vectors.

Language: Английский

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Future of DNA-based insect monitoring

Trends in Genetics, Journal Year: 2023, Volume and Issue: 39(7), P. 531 - 544

Published: March 10, 2023

Language: Английский

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Environmental DNA: The next chapter

Molecular Ecology, Journal Year: 2024, Volume and Issue: 33(11)

Published: April 16, 2024

Abstract Molecular tools are an indispensable part of ecology and biodiversity sciences implemented across all biomes. About a decade ago, the use implementation environmental DNA (eDNA) to detect signals extracted from samples opened new avenues research. Initial eDNA research focused on understanding population dynamics target species. Its scope thereafter broadened, uncovering previously unrecorded via metabarcoding in both well‐studied understudied ecosystems taxonomic groups. The application rapidly became established research, field by its own. Here, we revisit key expectations made land‐mark special issue Ecology 2012 frame development six areas: (1) sample collection, (2) primer development, (3) biomonitoring, (4) quantification, (5) behaviour environment (6) reference database development. We pinpoint success eDNA, yet also discuss shortfalls not met, highlighting areas priority identify unexpected developments. In parallel, our retrospective couples screening peer‐reviewed literature with survey users including academics, end‐users commercial providers, which address focus efforts advance eDNA. With rapid ever‐increasing pace technical advances, future looks bright, successful applications best practices must become more interdisciplinary reach full potential. Our retrospect gives towards concretely moving forward.

Language: Английский

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Proficiency testing and cross-laboratory method comparison to support standardisation of diatom DNA metabarcoding for freshwater biomonitoring

Metabarcoding and Metagenomics, Journal Year: 2025, Volume and Issue: 9

Published: Jan. 10, 2025

DNA metabarcoding of benthic diatoms has been successfully applied for biomonitoring at the national scale and can now be considered technically ready routine application. However, protocols methods still vary between within countries, limiting their transferability comparability results. In order to overcome this, use diatom requires knowledge sources variability introduced by different steps procedure. Here, we examine how elements procedures contribute European laboratories. A set four experiments were performed focusing on extraction PCR amplification evaluate reproducibility laboratories currently scientific community. Under guidance a reference laboratory, 17 participants from 14 countries in parallel, using same fixed protocol own choice protocol. Experiments each participant standardised biofilm samples (river, lake mock community) investigate potential systematic random errors. Our results revealed successful amongst labs highly similar consistent ecological assessment outcome obtained regardless used participant. We propose an “all one but prove them all” strategy, suggesting that distinct community, as long consistency is proven following minimum standard requirements.

Language: Английский

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Strategies for sample labelling and library preparation in DNA metabarcoding studies

Molecular Ecology Resources, Journal Year: 2021, Volume and Issue: 22(4), P. 1231 - 1246

Published: Sept. 22, 2021

Abstract Metabarcoding of DNA extracted from environmental or bulk specimen samples is increasingly used to profile biota in basic and applied biodiversity research because its targeted nature that allows sequencing genetic markers many parallel. To achieve this, PCR amplification carried out with primers designed target a taxonomically informative marker within taxonomic group, sample‐specific nucleotide identifiers are added the amplicons prior sequencing. The latter enables assignment sequences back they originated from. Nucleotide can be during metabarcoding “library preparation”, is, when prepared for Different strategies this labelling exist. All have advantages, challenges limitations, some which lead misleading results, worst case compromise fidelity data. Given range questions addressed using metabarcoding, ensuring data generation robust fit chosen purpose critically important practitioners seeking employ assessments. Here, we present an overview three main workflows library preparation studies on Illumina platforms; one‐step PCR, two‐step tagged PCR. Further, distill key considerations researchers select appropriate strategy their specific study. Ultimately, by gaining insights into consequences different workflows, hope further consolidate power as tool assess across applications.

Language: Английский

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ONTbarcoder 2.0: rapid species discovery and identification with real‐time barcoding facilitated by Oxford Nanopore R10.4

Cladistics, Journal Year: 2023, Volume and Issue: 40(2), P. 192 - 203

Published: Dec. 2, 2023

Abstract Most arthropod species are undescribed and hidden in specimen‐rich samples that difficult to sort using morphological characters. For such samples, sorting putative with DNA barcodes is an attractive alternative, but needs cost‐effective techniques suitable for use many laboratories around the world. Barcoding portable inexpensive MinION sequencer produced by Oxford Nanopore Technologies (ONT) could be useful presorting because it requires little space inexpensive. However, similarly important user‐friendly reliable software analysis of ONT data. It here provided form ONTbarcoder 2.0 all commonly used operating systems includes a Graphical User Interface (GUI). Compared earlier version, has three key improvements related higher read quality obtained ONT's latest flow cells (R10.4), chemistry (V14 kits) basecalling model (super‐accuracy model). First, improved (R10.4) allows primers shorter indices than those previously needed (9 bp vs. 12–13 bp). This decreases primer cost can potentially improve PCR success rates. Second, now delivers real‐time barcoding complement sequencing. means first within minutes starting sequencing run; i.e. cell optimized terminating runs when most have already been obtained. The only input demultiplexing sheet data (raw or basecalled) generated either Mk1B Mk1C. Thirdly, we demonstrate availability R10.4 low‐cost Flongle option users who require 200–250 at time.

Language: Английский

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It's raining species: Rainwash eDNA metabarcoding as a minimally invasive method to assess tree canopy invertebrate diversity

Environmental DNA, Journal Year: 2022, Volume and Issue: 5(1), P. 3 - 11

Published: Nov. 22, 2022

Abstract Forest canopies are highly diverse ecosystems, but despite several decades of intense research, there remain substantial gaps in our knowledge their biodiversity and ecological interactions. One fundamental challenge canopy research is the limited accessibility ecosystem. Consequently, previous studies have relied on application either invasive methods such as chemical knockdown, or time‐consuming expensive setups walkways cranes. Therefore, time‐ cost‐efficient, ideally minimally yet comprehensive applications required to help close this gap. High‐throughput metabarcoding environmental DNA (eDNA) collected from water, soil, air provides a method for assessment, its potential monitoring has not been explored. Herein, we conducted eDNA washed off via rainwater explore research. We placed four 1 m 2 rain samplers beneath different trees (beech, oak, larch, pine) prior major event, filtered rainwater, performed cytochrome c oxidase subunit I (COI) gene profile invertebrate community. Additionally, identified all specimens present assess if only came physically rainwater. detected 50 species by metabarcoding, which 43 were water sample, thus likely representing true signals. Furthermore, observed distinct occurrence patterns corresponding trees, suggesting that host specificity can potentially be assessed using method. In conclusion, study proof principle rainwash offers tree canopies.

Language: Английский

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