scholarly journals Assessing the responses of Sphagnum micro-eukaryotes to climate changes using high throughput sequencing

PeerJ ◽  
2020 ◽  
Vol 8 ◽  
pp. e9821 ◽  
Author(s):  
Monika K. Reczuga ◽  
Christophe Victor William Seppey ◽  
Matthieu Mulot ◽  
Vincent E.J. Jassey ◽  
Alexandre Buttler ◽  
...  
Keyword(s):  
High Throughput ◽  
Ecosystem Functioning ◽  
High Throughput Sequencing ◽  
Ecosystem Respiration ◽  
Carbon Sources ◽  
Environmental Dna ◽  
Severe Drought ◽  
Food Web Interactions ◽  
Eukaryotic Diversity ◽  
Eukaryotic Microorganisms

Current projections suggest that climate warming will be accompanied by more frequent and severe drought events. Peatlands store ca. one third of the world’s soil organic carbon. Warming and drought may cause peatlands to become carbon sources through stimulation of microbial activity increasing ecosystem respiration, with positive feedback effect on global warming. Micro-eukaryotes play a key role in the carbon cycle through food web interactions and therefore, alterations in their community structure and diversity may affect ecosystem functioning and could reflect these changes. We assessed the diversity and community composition of Sphagnum-associated eukaryotic microorganisms inhabiting peatlands and their response to experimental drought and warming using high throughput sequencing of environmental DNA. Under drier conditions, micro-eukaryotic diversity decreased, the relative abundance of autotrophs increased and that of osmotrophs (including Fungi and Peronosporomycetes) decreased. Furthermore, we identified climate change indicators that could be used as early indicators of change in peatland microbial communities and ecosystem functioning. The changes we observed indicate a shift towards a more “terrestrial” community in response to drought, in line with observed changes in the functioning of the ecosystem.

scholarly journals High-throughput metabarcoding characterises fungal endophyte diversity in the phyllosphere of a barley crop

2021 ◽  
Author(s):  
Carla Bridget Milazzo ◽  
Katherine Grace Zulak ◽  
Mariano Jordi Muria-Gonzalez ◽  
Darcy Jones ◽  
Matthew Power ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Large Subunit ◽  
Fungal Endophytes ◽  
Environmental Dna ◽  
Fungal Endophyte ◽  
Bioinformatics Pipeline ◽  
Pests And Diseases ◽  
Key Factor ◽  
Agroecosystem Management

Over the last decade, the microbiome has received increasing attention as a key factor in macroorganism fitness. Sustainable pest management requires an understanding of the complex microbial endophyte communities existing symbiotically within plants and the way synthetic pesticides interact with them. Fungal endophytes are known to benefit plant growth and fitness and may deter pests and diseases. Recent advances in high-throughput sequencing (HTS) have enabled integrative microbiome studies especially in agricultural contexts. Here we profile the fungal endophyte community in the phyllosphere of two barley (Hordeum vulgare) cultivars exposed to two systemic foliar fungicides using metabarcoding, a HTS tool that constructs community profiles from environmental DNA (eDNA). We studied the fungal nuclear ribosomal large subunit (LSU) D2 and ITS2 DNA markers through a bioinformatics pipeline introduced here. We found 88 and 128 unique amplicon sequence variants (ASVs) using the D2 and ITS2 metabarcoding assays, respectively. With principal coordinate analysis (PCoA) and PERMANOVA, ASV diversity did not change in response to barley cultivar or fungicide treatment, however the community structure of unsprayed plants did change between two collection times eight days apart. The workflow described here can be applied to other microbiome studies in agriculture and we hope it encourages further research into crop microbiomes to improve agroecosystem management.

scholarly journals GenBank is a reliable resource for 21st century biodiversity research

2019 ◽  
Vol 116 (45) ◽  
pp. 22651-22656 ◽  
Author(s):  
Matthieu Leray ◽  
Nancy Knowlton ◽  
Shian-Lei Ho ◽  
Bryan N. Nguyen ◽  
Ryuji J. Machida
Keyword(s):  
Phylogenetic Analysis ◽  
Global Change ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Environmental Dna ◽  
Taxonomic Resolution ◽  
Biodiversity Research ◽  
Animal Diversity ◽  
Mitochondrial Sequences ◽  
The Many

Traditional methods of characterizing biodiversity are increasingly being supplemented and replaced by approaches based on DNA sequencing alone. These approaches commonly involve extraction and high-throughput sequencing of bulk samples from biologically complex communities or samples of environmental DNA (eDNA). In such cases, vouchers for individual organisms are rarely obtained, often unidentifiable, or unavailable. Thus, identifying these sequences typically relies on comparisons with sequences from genetic databases, particularly GenBank. While concerns have been raised about biases and inaccuracies in laboratory and analytical methods, comparatively little attention has been paid to the taxonomic reliability of GenBank itself. Here we analyze the metazoan mitochondrial sequences of GenBank using a combination of distance-based clustering and phylogenetic analysis. Because of their comparatively rapid evolutionary rates and consequent high taxonomic resolution, mitochondrial sequences represent an invaluable resource for the detection of the many small and often undescribed organisms that represent the bulk of animal diversity. We show that metazoan identifications in GenBank are surprisingly accurate, even at low taxonomic levels (likely <1% error rate at the genus level). This stands in contrast to previously voiced concerns based on limited analyses of particular groups and the fact that individual researchers currently submit annotated sequences to GenBank without significant external taxonomic validation. Our encouraging results suggest that the rapid uptake of DNA-based approaches is supported by a bioinformatic infrastructure capable of assessing both the losses to biodiversity caused by global change and the effectiveness of conservation efforts aimed at slowing or reversing these losses.

scholarly journals In silico assessment of primers for eDNA studies using PrimerTree and application to characterize the biodiversity surrounding the Cuyahoga River

2015 ◽  
Author(s):  
M.V. Cannon ◽  
J. Hester ◽  
A. Shalkhauser ◽  
E.R. Chan ◽  
K. Logue ◽  
...  
Keyword(s):  
High Throughput ◽  
Dna Sequences ◽  
Water Samples ◽  
High Throughput Sequencing ◽  
Biological Diversity ◽  
Pcr Amplification ◽  
Environmental Dna ◽  
Asian Carp ◽  
Broad Perspective ◽  
Wide Range

Analysis of environmental DNA (eDNA) enables the detection of species of interest from water and soil samples, typically using species-specific PCR. Here, we describe a method to characterize the biodiversity of a given environment by amplifying eDNA using primer pairs targeting a wide range of taxa and high-throughput sequencing for species identification. We tested this approach on 91 water samples of 40 mL collected along the Cuyahoga River (Ohio, USA). We amplified eDNA using 12 primer pairs targeting mammals, fish, amphibians, birds, bryophytes, arthropods, copepods, plants and several microorganism taxa and sequenced all PCR products simultaneously by high-throughput sequencing. Overall, we identified DNA sequences from 15 species of fish, 17 species of mammals, 8 species of birds, 15 species of arthropods, one turtle and one salamander. Interestingly, in addition to aquatic and semiaquatic animals, we identified DNA from terrestrial species that live near the Cuyahoga River. We also identified DNA from one Asian carp species invasive to the Great Lakes but that had not been previously reported in the Cuyahoga River. Our study shows that analysis of eDNA extracted from small water samples using wide-range PCR amplification combined with high-throughput sequencing can provide a broad perspective on biological diversity.

scholarly journals High-Throughput Sequencing of Environmental DNA as a Tool for Monitoring Eukaryotic Communities and Potential Pathogens in a Coastal Upwelling Ecosystem

2021 ◽  
Vol 8 ◽  
Author(s):  
Raquel Ríos-Castro ◽  
Alejandro Romero ◽  
Raquel Aranguren ◽  
Alberto Pallavicini ◽  
Elisa Banchi ◽  
...  
Keyword(s):  
High Throughput Sequencing ◽  
Coastal Upwelling ◽  
Marine Ecosystem ◽  
Environmental Dna ◽  
Functional Roles ◽  
Molecular Approaches ◽  
Eukaryotic Microorganisms ◽  
Causative Agents ◽  
Veterinary Importance ◽  
Water Fractions

The marine environment includes diverse microeukaryotic organisms that play important functional roles in the ecosystem. With molecular approaches, eukaryotic taxonomy has been improved, complementing classical analysis. In this study, DNA metabarcoding was performed to describe putative pathogenic eukaryotic microorganisms in sediment and marine water fractions collected in Galicia (NW Spain) from 2016 to 2018. The composition of eukaryotic communities was distinct between sediment and water fractions. Protists were the most diverse group, with the clade TSAR (Stramenopiles, Alveolata, Rhizaria, and Telonemida) as the primary representative organisms in the environment. Harmful algae and invasive species were frequently detected. Potential pathogens, invasive pathogenic organisms as well as the causative agents of harmful phytoplanktonic blooms were identified in this marine ecosystem. Most of the identified pathogens have a crucial impact on the aquacultural sector or affect to relevant species in the marine ecosystem, such as diatoms. Moreover, pathogens with medical and veterinary importance worldwide were also found, as well as pathogens that affect diatoms. The evaluation of the health of a marine ecosystem that directly affects the aquacultural sector with a zoonotic concern was performed with the metabarcoding assay.

scholarly journals Perspectives and benefits of high-throughput long-read sequencing in microbial ecology

2021 ◽  
Author(s):  
Leho Tedersoo ◽  
Mads Albertsen ◽  
Sten Anslan ◽  
Benjamin Callahan
Keyword(s):  
Microbial Ecology ◽  
High Throughput ◽  
Single Molecule ◽  
High Throughput Sequencing ◽  
Environmental Dna ◽  
Nanopore Sequencing ◽  
High Quality ◽  
Short Read ◽  
Sequencing Technologies ◽  
Long Read

Short-read, high-throughput sequencing (HTS) methods have yielded numerous important insights into microbial ecology and function. Yet, in many instances short-read HTS techniques are suboptimal, for example by providing insufficient phylogenetic resolution or low integrity of assembled genomes. Single-molecule and synthetic long-read (SLR) HTS methods have successfully ameliorated these limitations. In addition, nanopore sequencing has generated a number of unique analysis opportunities such as rapid molecular diagnostics and direct RNA sequencing, and both PacBio and nanopore sequencing support detection of epigenetic modifications. Although initially suffering from relatively low sequence quality, recent advances have greatly improved the accuracy of long read sequencing technologies. In spite of great technological progress in recent years, the long-read HTS methods (PacBio and nanopore sequencing) are still relatively costly, require large amounts of high-quality starting material, and commonly need specific solutions in various analysis steps. Despite these challenges, long-read sequencing technologies offer high-quality, cutting-edge alternatives for testing hypotheses about microbiome structure and functioning as well as assembly of eukaryote genomes from complex environmental DNA samples.

scholarly journals Assessment of Passive Traps Combined with High-Throughput Sequencing To Study Airborne Fungal Communities

2018 ◽  
Vol 84 (11) ◽  
Author(s):  
Jaime Aguayo ◽  
Céline Fourrier-Jeandel ◽  
Claude Husson ◽  
Renaud Ioos
Keyword(s):  
Dna Extraction ◽  
High Throughput ◽  
Fungal Diversity ◽  
High Throughput Sequencing ◽  
Linear Models ◽  
Environmental Dna ◽  
Illumina Miseq ◽  
Fungal Communities ◽  
Correct Identification ◽  
Spore Trap

ABSTRACTTechniques based on high-throughput sequencing (HTS) of environmental DNA have provided a new way of studying fungal diversity. However, these techniques suffer from a number of methodological biases which may appear at any of the steps involved in a metabarcoding study. Air is one of the most important environments where fungi can be found, because it is the primary medium of dispersal for many species. Looking ahead to future developments, it was decided to test 20 protocols, including different passive spore traps, spore recovery procedures, DNA extraction kits, and barcode loci. HTS was performed with the Illumina MiSeq platform targeting two subloci of the fungal internal transcribed spacer. Multivariate analysis and generalized linear models showed that the type of passive spore trap, the spore recovery procedure, and the barcode all impact the description of fungal communities in terms of richness and diversity when assessed by HTS metabarcoding. In contrast, DNA extraction kits did not significantly impact these results. Although passive traps may be used to describe airborne fungal communities, a study using specific real-time PCR and a mock community showed that these kinds of traps are affected by environmental conditions that may induce losses of biological material, impacting diversity and community composition results.IMPORTANCEThe advent of high-throughput sequencing (HTS) methods, such as those offered by next-generation sequencing (NGS) techniques, has opened a new era in the study of fungal diversity in different environmental substrates. In this study, we show that an assessment of the diversity of airborne fungal communities can reliably be achieved by the use of simple and robust passive spore traps. However, a comparison of sample processing protocols showed that several methodological biases may impact the results of fungal diversity when assessed by metabarcoding. Our data suggest that identifying these biases is of paramount importance to enable a correct identification and relative quantification of community members.

Barcoding the food chain: from Sanger to high-throughput sequencing

Genome ◽  
2016 ◽  
Vol 59 (11) ◽  
pp. 946-958 ◽  
Author(s):  
Joanne E. Littlefair ◽  
Elizabeth L. Clare
Keyword(s):  
Dna Barcoding ◽  
High Throughput ◽  
Food Chain ◽  
High Throughput Sequencing ◽  
Human Mobility ◽  
Environmental Dna ◽  
Universal Primers ◽  
Individual Species ◽  
Parasitic Infections ◽  
Forensic Evaluation

Society faces the complex challenge of supporting biodiversity and ecosystem functioning, while ensuring food security by providing safe traceable food through an ever-more-complex global food chain. The increase in human mobility brings the added threat of pests, parasites, and invaders that further complicate our agro-industrial efforts. DNA barcoding technologies allow researchers to identify both individual species, and, when combined with universal primers and high-throughput sequencing techniques, the diversity within mixed samples (metabarcoding). These tools are already being employed to detect market substitutions, trace pests through the forensic evaluation of trace “environmental DNA”, and to track parasitic infections in livestock. The potential of DNA barcoding to contribute to increased security of the food chain is clear, but challenges remain in regulation and the need for validation of experimental analysis. Here, we present an overview of the current uses and challenges of applied DNA barcoding in agriculture, from agro-ecosystems within farmland to the kitchen table.

scholarly journals From Rare Species Detection to Whole-Community Diversity Using High-Throughput Sequencing of Freshwater eDNA

2019 ◽  
Vol 3 ◽  
Author(s):  
Maria Judite Alves ◽  
Hugo Gante ◽  
Ana Veríssimo ◽  
Filipe Ribeiro ◽  
Carlos David Santos ◽  
...  
Keyword(s):  
High Throughput ◽  
Rare Species ◽  
High Throughput Sequencing ◽  
Environmental Dna ◽  
Community Diversity ◽  
Census Methods ◽  
Recent Approach ◽  
Fish Samples ◽  
Technological Developments

Understanding natural communities and ecosystems and the services they provide to humanity is highly dependent on knowledge about species composition and diversity through space and time. This is especially difficult in aquatic systems where traditional census methods provide species compositions that are usually truncated since rare species tend to go undetected. Detection of the rare species is important because they are either threatened or invasives at the earliest stage of invasion. One recent approach allowing detection of rare species uses environmental DNA (eDNA), present in water or soil, as traces of their existence. Here we propose to make use of recent technological developments in the area of high throughput sequencing to characterize freshwater fish communities and detect rare species, using a combination of eDNA metabarcoding and bulk eDNA metagenomics. A case-study will be conducted on the River Tagus (Portugal), which is inhabited by several rare fish species including both native and introduced taxa. In addition, the applicability of eDNA metagenomics for estimating the genetic diversity of populations will be assessed by comparing the results against those produced by traditional genetic screening of individual fish samples.

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