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1,076 results for “Metabarcoding”
Data from: 18S rRNA metabarcoding diet analysis of the predatory fish community across seasonal changes in prey availability
Predator-prey relationships are important ecological interactions, affecting biotic community composition and energy flow through a system, and are of interest to ecologists and managers. Morphological diet analysis has been the primary method used to quantify the diets of predators, but emerging molecular techniques using genetic data can provide more accurate estimates of relative diet composition. This study used sequences from the 18S V9 rRNA barcoding region to identify prey items in the gastrointestinal (GI) tracts of predatory fishes. Predator GI samples were taken from the Black River, Cheboygan Co., MI, USA (n=367 samples, 12 predator species) during periods of high prey availability, including the larval stage of regionally threatened lake sturgeon (Acipenser fulvescens Rafinesque 1817) in late May/early June of 2015 and of relatively lower prey availability in early July of 2015. DNA was extracted and sequenced from 355 samples (96.7%), and prey DNA was identified in 286 of the 355 samples (80.6%). Prey were grouped into 33 ecologically significant taxonomic groups based on the lowest taxonomic level sequences that could be identified using sequences available on GenBank. Changes in the makeup of diet composition, dietary overlap, and predator preference were analyzed comparing the periods of high and low prey abundance. Some predator species exhibited compositional changes in diet. Dietary overlap was slightly but significantly higher during the period of high prey abundance. There was little evidence for large changes in predator preference between the two periods. This study demonstrates the utility of molecular diet analysis in understanding predator-prey interactions in complex freshwater communities.
Data from: Reliable, verifiable and efficient monitoring of biodiversity via metabarcoding
To manage and conserve biodiversity, one must know what is being lost, where, and why, as well as which remedies are likely to be most effective. Metabarcoding technology can characterise the species compositions of mass samples of eukaryotes or of environmental DNA. Here, we validate metabarcoding by testing it against three high-quality standard data sets that were collected in Malaysia (tropical), China (subtropical) and the United Kingdom (temperate) and that comprised 55,813 arthropod and bird specimens identified to species level with the expenditure of 2,505 person-hours of taxonomic expertise. The metabarcode and standard data sets exhibit statistically correlated alpha- and beta-diversities, and the two data sets produce similar policy conclusions for two conservation applications: restoration ecology and systematic conservation planning. Compared with standard biodiversity data sets, metabarcoded samples are taxonomically more comprehensive, many times quicker to produce, less reliant on taxonomic expertise and auditable by third parties, which is essential for dispute resolution.
Data from: DNA metabarcoding unveils multi‐scale trophic variation in a widespread coastal opportunist
A thorough understanding of ecological networks relies on comprehensive information on trophic relationships among species. Since unpicking the diet of many organisms is unattainable using traditional morphology‐based approaches, the application of high‐throughput sequencing methods represents a rapid and powerful way forward. Here, we assessed the application of DNA‐metabarcoding with nearly universal primers for the mitochondrial marker cytochrome c oxidase I (COI) in defining the trophic ecology of adult brown shrimp, Crangon crangon, in six European estuaries. The exact trophic role of this abundant and widespread coastal benthic species is somewhat controversial, while information on geographical variation remains scant. Results revealed a highly opportunistic behaviour. Shrimp stomach contents contained hundreds of taxa (>1000 molecular operational taxonomic units), of which 291 were identified as distinct species, belonging to 35 phyla. Only twenty ascertained species had a mean relative abundance of more than 0.5%. Predominant species included other abundant coastal and estuarine taxa, including the shore crab Carcinus maenas and the amphipod Corophium volutator. Jacobs' selectivity index estimates based on DNA extracted from both shrimp stomachs and sediment samples were used to assess the shrimp's trophic niche indicating a generalist diet, dominated by crustaceans, polychaetes and fish. Spatial variation in diet composition, at regional and local scales, confirmed the highly flexible nature of this trophic opportunist. Furthermore, the detection of a prevalent, possibly endoparasitic fungus (Purpureocillium lilacinum) in the shrimp's stomach demonstrates the wide range of questions that can be addressed using metabarcoding, towards a more robust reconstruction of ecological networks.
Data from: DNA metabarcoding reveals the broad and flexible diet of a declining aerial insectivore
Aerial insectivores are highly mobile predators that feed on diverse prey items that have highly variable distributions. As such, investigating the diet, prey selection, and prey availability of aerial insectivores can be challenging. In this study, we used an integrated DNA barcoding method to investigate the diet and food supply of Barn Swallows, an aerial insectivore whose North American population has declined over the past 40 years. We tested the hypotheses that Barn Swallows are generalist insectivores when provisioning their young and select prey based on size. We predicted that the diets of nestlings would contain a range of insect taxa but would be biased towards large prey items and that the diet of nestlings would change as prey availability changed. We collected insects using Malaise traps at ten breeding sites and identified specimens using standard DNA barcoding. The sequences from these insect specimens were used to create a custom reference database of prey species and their relative sizes for our study area. We identified insect prey items from nestling fecal samples by using high-throughput DNA sequencing and comparing the sequences to our custom reference database. Barn Swallows fed nestlings prey items from 130 families representing 13 orders but showed selection for larger prey items that were predominantly from seven dipteran families. Nestling diet varied both within and between breeding seasons as well as between breeding sites. This dietary flexibility suggests that Barn Swallows are able to adjust their provisioning to changing prey availability on the breeding grounds when feeding their nestlings. Our study demonstrates the utility of custom reference databases for linking the abundance and size of insect prey in the habitat with prey consumed when employing molecular methods for dietary analysis.
Data from: Using DNA metabarcoding for simultaneous inference of common vampire bat diet and population structure
Metabarcoding diet analysis has become a valuable tool in animal ecology; however, co-amplified predator sequences are not generally used for anything other than to validate predator identity. Exemplified by the common vampire bat we demonstrate the use of metabarcoding to infer predator population structure alongside diet assessments. Growing populations of common vampire bats impact human, livestock and wildlife health in Latin America through transmission of pathogens, such as lethal rabies infections. Techniques to determine large scale variation in vampire bat diet and bat population structure would empower locality- and species-specific projections of disease transmission risks. However, previously used methods are not cost-effective and efficient for large scale applications. Using blood meal and faecal samples from common vampire bats from coastal, Andean and Amazonian regions of Peru, we showcase metabarcoding as a scalable tool to assess vampire bat population structure and feeding preferences. Dietary metabarcoding was highly effective, detecting vertebrate prey in 93.2% of the samples. Bats predominantly preyed on domestic animals, but fed on tapirs at one Amazonian site. In addition, we identified arthropods in 9.3% of samples, likely reflecting consumption of ectoparasites. Using the same data, we document mitochondrial geographic population structure in the common vampire bat in Peru. Such simultaneous inference of vampire bat diet and population structure can enable new insights into the interplay between vampire bat ecology and disease transmission risks. Importantly, the methodology can be extrapolated to metabarcoding diet studies of other animals to couple information on diet and population structure.
Data from: Metabarcoding and mitochondrial metagenomics of endogean arthropods to unveil the mesofauna of the soil
Biological communities inhabiting the soil are among the most diversified, complex and yet most poorly studied terrestrial ecosystems. The greatest knowledge gaps apply to the arthropod mesofauna (0·1–2 mm body size) because conventional morphological and molecular approaches are in many cases insufficient for the characterisation of these complex communities. The development of high-throughput sequencing (HTS) methodologies is required to solve current impediments and to further advance our understanding of below-ground biodiversity. We propose a flotation–Berlese–flotation (FBF) protocol for sampling and specimen processing to obtain 'clean' DNA extractions of arthropod mesofauna from the soil. In addition, we developed and tested HTS protocols for the characterisation of arthropod communities from these bulk DNA extractions using cox1 metabarcoding and shotgun metagenomic sequencing on the MiSeq Illumina platform. The FBF protocol provided DNA of soil arthropods from sufficiently large volumes of soil and free from contaminating bacteria and inhibitors. Metabarcoding and metagenomic sequencing on two deep soil samples from Iberian grasslands revealed >100 species of Acari and Collembola from 28 families. Genome assembly straight from shotgun sequencing of bulk specimens produced partial and full mitogenomes for 54 species with average length of >6000 bp. Metabarcoding and metagenomic sequencing resulted in closely congruent OTUs, but species numbers were highest with metabarcoding, while ∼73% of species were confirmed by matching shotgun sequence reads and ∼48% by contig assembly from those shotgun reads. In combination, the FBF protocol together with the PCR-based and shotgun sequencing pipelines addressed most of the challenges of studying soil arthropod mesofauna on the MiSeq Illumina platform. They are powerful, cost-efficient tools for characterising soil diversity in a phylogenetic and community ecology context. These methodological developments of HTS approaches for the study of mesofauna will accelerate ecological and evolutionary studies, biomonitoring of soil arthropods, and progress in both theoretical and applied soil science.
Data from: Environmental DNA metabarcoding studies are critically affected by substrate selection
Effective biomonitoring is critical for driving management outcomes that ensure long-term sustainability of the marine environment. In recent years environmental DNA (eDNA), coupled with metabarcoding methodologies, has emerged as a promising tool for generating biotic surveys of marine ecosystems, including those under anthropogenic pressure. However, more empirical data is needed on how to best implement eDNA field sampling approaches to maximise their utility for each specific application. The effect of the substrate chosen for eDNA sampling on the diversity of marine taxa detected by DNA metabarcoding has not yet been systematically analysed, despite aquatic systems being those most commonly targeted for eDNA studies. We investigated the effect of four commonly used eDNA substrates to explore taxonomic diversity: (i) surface water, (ii) marine sediment, (iii) settlement-plates and (iv) planktonic tows. With a focus on coastal ports, 332 eDNA samples from Australia (Indian and Southern oceans) and Kazakhstan (Caspian Sea) were collected and analysed by multi-assay DNA metabarcoding. Across study locations, between 30-52% of eukaryotic families detected were unique to a particular substrate and less than 6% of families were found in all four substrates. Taxonomic composition varied significantly depending on the substrate sampled implying that the suitability (and bias) of an eDNA substrate will depend on the focal taxa. These findings demonstrate that single substrate eDNA metabarcoding likely underestimates the total eukaryotic diversity. Future eDNA experimental design should consider incorporating multiple substrates or select substrate(s) best suited to the specific detection of target taxa.
Data from: ITS all right mama: Investigating the formation of chimeric sequences in the ITS2 region by DNA metabarcoding analyses of fungal mock communities of different complexities
The formation of chimeric sequences can create significant methodological bias in PCR-based DNA metabarcoding analyses. During mixed-template amplification of barcoding regions, chimera formation is frequent and well documented. However, profiling of fungal communities typically uses the more variable rDNA region ITS. Due to a larger research community, tools for chimera detection have been developed mainly for the 16S/18S markers. However, these tools are widely applied to the ITS region without verification of their performance. We examined the rate of chimera formation during amplification and 454 sequencing of the ITS2 region from fungal mock communities of different complexities. We evaluated the chimera detecting ability of two common chimera-checking algorithms: Perseus and UCHIME. Large proportions of the chimeras reported were false positives. No false negatives were found in the dataset. Verified chimeras accounted for only 0.2% of the total ITS2 reads, which is considerably less than what is typically reported in 16S and 18S metabarcoding analyses. Verified chimeric "parent sequences" had significantly higher percent identity to one another than to random members of the mock communities. Community complexity increased the rate of chimera formation. GC content was higher around the verified chimeric break points, potentially facilitating chimera formation through base pair mismatching in the neighboring regions of high similarity in the chimeric region. We conclude that the hypervariable nature of the ITS region seem to buffer the rate of chimera formation in comparison to other, less variable barcoding regions, due to shorter regions of high sequence similarity.
Data from: DNA barcoding and metabarcoding of standardized samples reveal patterns of marine benthic diversity
Documenting the diversity of marine life is challenging because many species are cryptic, small, and rare, and belong to poorly known groups. New sequencing technologies, especially when combined with standardized sampling, promise to make comprehensive biodiversity assessments and monitoring feasible on a large scale. We used this approach to characterize patterns of diversity on oyster reefs across a range of geographic scales comprising a temperate location [Virginia (VA)] and a subtropical location [Florida (FL)]. Eukaryotic organisms that colonized multilayered settlement surfaces (autonomous reef monitoring structures) over a 6-mo period were identified by cytochrome c oxidase subunit I barcoding (>2-mm mobile organisms) and metabarcoding (sessile and smaller mobile organisms). In a total area of ∼15.64 m2 and volume of ∼0.09 m3, 2,179 operational taxonomic units (OTUs) were recorded from 983,056 sequences. However, only 10.9% could be matched to reference barcodes in public databases, with only 8.2% matching barcodes with both genus and species names. Taxonomic coverage was broad, particularly for animals (22 phyla recorded), but 35.6% of OTUs detected via metabarcoding could not be confidently assigned to a taxonomic group. The smallest size fraction (500 to 106 μm) was the most diverse (more than two-thirds of OTUs). There was little taxonomic overlap between VA and FL, and samples separated by ∼2 m were significantly more similar than samples separated by ∼100 m. Ground-truthing with independent assessments of taxonomic composition indicated that both presence–absence information and relative abundance information are captured by metabarcoding data, suggesting considerable potential for ecological studies and environmental monitoring.
Supplementary material 12 from: Paez-Triana L, Herrera G, Vega L, Garcia-Corredor D, Pulido Medellín MO, Paniz-Mondolfi A, Muñoz M, Ramírez JD (2023) Metagenomic exploration of endosymbionts and pathogens in the tropical lineage of Rhipicephalus sanguineus sensu lato (s.l.) ticks in Colombia. Metabarcoding and Metagenomics 7: e109085. https://doi.org/10.3897/mbmg.7.109085
p-values obtained between departments and sexes within each department using the Wilcoxon test on the relative abundances of pathogens and endosymbionts
Supplementary material 3 from: Paez-Triana L, Herrera G, Vega L, Garcia-Corredor D, Pulido Medellín MO, Paniz-Mondolfi A, Muñoz M, Ramírez JD (2023) Metagenomic exploration of endosymbionts and pathogens in the tropical lineage of Rhipicephalus sanguineus sensu lato (s.l.) ticks in Colombia. Metabarcoding and Metagenomics 7: e109085. https://doi.org/10.3897/mbmg.7.109085
Significant differences in the relative abundances of assigned reads in Bacteria and Archaea between departments and sexes of the various genera found in the samples
Supplementary material 7 from: Paez-Triana L, Herrera G, Vega L, Garcia-Corredor D, Pulido Medellín MO, Paniz-Mondolfi A, Muñoz M, Ramírez JD (2023) Metagenomic exploration of endosymbionts and pathogens in the tropical lineage of Rhipicephalus sanguineus sensu lato (s.l.) ticks in Colombia. Metabarcoding and Metagenomics 7: e109085. https://doi.org/10.3897/mbmg.7.109085
Metadata and information regarding the genomes included in the analysis for endosymbiont and pathogen identification
Supplementary material 5 from: Paez-Triana L, Herrera G, Vega L, Garcia-Corredor D, Pulido Medellín MO, Paniz-Mondolfi A, Muñoz M, Ramírez JD (2023) Metagenomic exploration of endosymbionts and pathogens in the tropical lineage of Rhipicephalus sanguineus sensu lato (s.l.) ticks in Colombia. Metabarcoding and Metagenomics 7: e109085. https://doi.org/10.3897/mbmg.7.109085
Results of dereplication, as well as phylogenomics analysis and pangenome exploration of all samples (including longer MAGs)
Supplementary material 10 from: Paez-Triana L, Herrera G, Vega L, Garcia-Corredor D, Pulido Medellín MO, Paniz-Mondolfi A, Muñoz M, Ramírez JD (2023) Metagenomic exploration of endosymbionts and pathogens in the tropical lineage of Rhipicephalus sanguineus sensu lato (s.l.) ticks in Colombia. Metabarcoding and Metagenomics 7: e109085. https://doi.org/10.3897/mbmg.7.109085
p-values obtained between departments and sexes within each department using the Wilcoxon test on the relative abundances of each genus (Bacteria and Archaea)
Data and code for: "Foraging rates from metabarcoding: Predators have reduced functional responses in wild, diverse prey communities."
<p>Data and code for "Foraging rates from metabarcoding: Predators have reduced functional responses in wild, diverse prey communities."</p>
Supplementary material 3 from: Ahmed M, Slos D, Holovachov O (2024) Assessing the diversity of nematodes in the Store Mosse National Park (Sweden) using metabarcoding. Metabarcoding and Metagenomics 8: e111307. https://doi.org/10.3897/mbmg.8.111307
Proportions of total ASVs assigned to various nematode families including those unassigned at the family level across all samples
Metadata of sampling strategy during metabarcoding analysis (Journal publication supplement)
<p>The table presents supplementary materials and contains metadata of sampling strategy during metabarcoding analysis of four types of substrates in two ombrotrophic bog habitats, with other experimental and environmental parameters included in analyses. </p>
The precautionary principle and dietary DNA metabarcoding: commonly used abundance thresholds change ecological interpretation
<p>Dietary DNA metabarcoding enables researchers to identify and characterize trophic interactions with a high degree of taxonomic precision. It is also sensitive to sources of bias and contamination in the field and lab. One of the earliest and most common strategies for dealing with such sensitivities has been to filter resulting sequence data to remove low-abundance sequences before conducting ecological analyses based on the presence or absence of food taxa. Although this step is now often perceived to be both necessary and sufficient for cleaning up datasets, evidence to support this perception is lacking and more attention needs to be paid to the related risk of introducing other undesirable errors. Using computer simulations, we demonstrate that common strategies to remove low-abundance sequences can erroneously eliminate true dietary sequences in ways that impact downstream dietary inferences. Using real data from well-studied wildlife populations in Yellowstone National Park, we further show how these strategies can markedly alter the composition of individual dietary profiles in ways that scale-up to obscure ecological interpretations about dietary generalism, specialism, and niche partitioning. Although the practice of removing low-abundance sequences may continue to be a useful strategy to address a subset of research questions that focus on a subset of relatively abundant food resources, its continued widespread use risks generating misleading perceptions about the structure of trophic networks. Researchers working with dietary DNA metabarcoding data—or similar data such as environmental DNA, microbiomes, or pathobiomes—should be aware of potential drawbacks and consider alternative bioinformatic, experimental, and statistical solutions.</p>
Deciphering the diet of a wandering spider (Phoneutria boliviensis; Araneae: Ctenidae) by DNA metabarcoding of gut contents
<p>Arachnids are the most abundant land predators. Despite the importance of their functional roles as predators and the necessity to understand their diet for conservation, the trophic ecology of many arachnid species has not been sufficiently studied. In the case of the wandering spider, <i>Phoneutria boliviensis</i> F. O. Pickard-Cambridge, 1897, only field and laboratory observational studies on their diet exist. By using a DNA metabarcoding approach, we compared the prey found in the gut content of males and females from three distant Colombian populations of <i>P. boliviensis</i>. By DNA metabarcoding of the cytochrome <i>c</i> oxidase subunit I (COI), we detected and identified 234 prey items (individual captured by the spider) belonging to 96 operational taxonomic units (OTUs), as prey for this wandering predator. Our results broaden the known diet of <i>P. boliviensis</i> with at least 75 prey taxa not previously registered in fieldwork or laboratory experimental trials. These results suggest that <i>P. boliviensis</i> feeds predominantly on invertebrates (Diptera, Lepidoptera, Coleoptera and Orthoptera) and opportunistically on small squamates. Intersex and interpopulation differences were also observed. Assuming that prey preference does not vary between populations, these differences are likely associated with a higher local prey availability. Finally, we suggest that DNA metabarcoding can be used for evaluating subtle differences in the diet of distinct populations of <i>P. boliviensis, </i>particularly when predation records in the field cannot be established or quantified using direct observation.</p>
A national scale BioBlitz using citizen science and eDNA metabarcoding for monitoring coastal marine fish
<p>Marine biodiversity is threatened by human activities. To understand the changes happening in aquatic ecosystems and to inform management, detailed, synoptic monitoring of biodiversity across large spatial extents is needed. Such monitoring is challenging due to the time, cost, and specialized skills that this typically requires. In an unprecedented study, we here combined citizen science with eDNA metabarcoding to map coastal fish biodiversity at a national scale. We engaged 360 citizen scientists to collect filtered sea water samples from 100 sites across Denmark over two seasons (1 pm on September 29<sup>th</sup> 2019 and May 10<sup>th</sup> 2020), and by sampling at nearly the exact same time across all 100 sites, we obtained an overview of fish biodiversity largely unaffected by temporal variation. This would have been logistically impossible for the involved scientists without the help of volunteer citizens. We obtained a high return rate of 94% of the samples, and a total richness of 52 fish species, representing approximately 80% of coastal Danish fish species and approximately 25% of all Danish marine fish species. We retrieved distribution patterns matching known occurrence for both invasive, endangered, and cryptic species, and detected seasonal variation in accordance with known phenology. Dissimilarity of eDNA community compositions increased with distance between sites. Importantly, comparing our eDNA data with National Fish Atlas data (the latter compiled from a century of observations) we found positive correlation between species richness values and a congruent patterns of community compositions. These findings support the use of eDNA-based citizen science to detect patterns in biodiversity, and our approach is readily scalable to other countries, or even regional and global scales. We argue that future large-scale biomonitoring will benefit from using citizen science combined with emerging eDNA technology, and that such an approach will be important for data-driven biodiversity management and conservation.</p>
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