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Fish monitoring is essential for assessing the effects of natural and anthropic stressors on marine ecosystems. In this context, environmental DNA (eDNA) metabarcoding appears to be a promising tool, due to its efficiency in species detection. However, before this method can be fully implemented in monitoring programs, more studies are needed to evaluate its ability to assess the composition of fish assemblages compared with traditional survey methods that have been used for decades. Here, we used both eDNA metabarcoding and Underwater Visual Census (UVC) to assess the taxonomic and functional diversity (presence-absence data) of Mediterranean fish communities. We collected eDNA samples and performed UVC strip transects inside and outside four Marine Protected Areas in the Mediterranean Sea. Samples for eDNA analysis were collected by filtering seawater simultaneously at the surface and the bottom, and DNA was amplified using a combination of three sets of primers. We found that eDNA alone made an outstanding characterisation of fish composition with the detection of 95 % of the 60 taxa identified in this study, whereas UVC recovered only 58 % of them. Functional diversity was better evaluated with eDNA than with UVC, with the detection of a greater breadth of functional traits. eDNA was even better at characterising functional than taxonomic diversity, providing reliable information on ecosystem functioning with little sampling effort. Together these results suggest that eDNA metabarcoding offers great potential for surveying complex marine ecosystems. Combining eDNA metabarcoding and UVC in integrated monitoring programs would therefore improve monitoring strategies and enhance our understanding of fish communities, a key step promoting their conservation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Understanding the influence of environmental pollutants on the assembly mechanisms of estuarine fish communities is vital for addressing conservation challenges in these biodiverse ecosystems. Although significant research has explored the toxic impacts of pollutants such as petroleum, heavy metals, and eutrophication on individual species and populations, their effects on community assembly processes and the differential responses of native versus non-native fish at the meta-community level remain inadequately understood. This study utilized environmental DNA (eDNA) metabarcoding to analyze fish community diversity across 28 subtropical estuaries in China, assessing how these pollutants affect community composition and assembly mechanisms. Results indicated that eDNA was 2.54 times more effective than traditional methods in species identification, while also enabling the detection of a higher number of non-native fish species and more diverse functional guilds within estuarine ecosystems. A significant distance decay pattern (p < 0.05) was observed among native fish, whereas non-native species exhibited non-significant patterns. Neutral and null models showed that non-native species had significantly higher migration rates (0.005939 vs 0.001757) and a greater contribution of stochastic processes (82.38 % vs 70.59 %) compared to native species. Additionally, distance-based redundancy analysis (db-RDA), variance partitioning analysis (VPA), and correlation analyses revealed that native species were strongly constrained by environmental factors, particularly oil, Hg, Zn, Pb, Cr 6+ , and NH 4 + , while non - native species displayed notable resilience to these pollutants. These findings highlight the potential for non-native species to disproportionately influence community dynamics and assembly through unrestricted random dispersal amid environmental disturbances. This research clarifies the contrasting ecological responses of native and non-native fish communities to anthropogenic pressures in estuarine environments, offering essential insights into ecosystem resilience and informing biodiversity conservation strategies in rapidly changing coastal ecosystems., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Nutrients introduced to the environment by finfish aquaculture pose environmental risks, which can be mitigated by robust environmental monitoring. Biological communities in soft sediments are good indicators of aquaculture derived environmental changes. Traditionally, monitoring programs have visually surveyed macrofauna communities. However, DNA metabarcoding is a potentially more efficient alternative. We compared alpha diversity, multivariate dispersion and taxonomic composition of macrofauna communities with metabarcoding derived bacterial and eukaryote communities along an organic enrichment gradient at a salmon farm in Tasmania, Australia. Additionally, we conducted a cost-benefit analysis comparing the approaches. All methods identified indicator taxa that changed in abundance over the enrichment gradient. Macrofauna analysis was the most sensitive method for detecting changes in alpha diversity, while metabarcoding was most sensitive for multivariate dispersion. Taxonomic composition of animal communities derived from the two methods differed drastically. Metabarcoding was cheaper than macrofauna for ≥93 samples and quicker for ≥14 samples., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 University of Tasmania. Published by Elsevier Ltd.. All rights reserved.)

Fish diversity, an important indicator of aquatic ecosystem health, is declining due to water pollution, overfishing, climate change, and invasive species. Effective surveying and monitoring are required to protect fish diversity. Here, a high-sensitivity environmental DNA (eDNA) metabarcoding technique was used to investigate fish diversity in the Danjiang River, Shaanxi Province, China. In total, 59 species were identified in eight orders, 19 families, and 40 genera. Cypriniformes and Perciformes were the main groups in the survey area, while Cyprinidae accounted for 50.85% of the total fish species. Rhinogobius similis (19%), Hemibarbus umbrifer (11%), Gnathopogon herzensteini (10%), Triplophysa stewarti (8%), and Zacco platypus (7%) were the dominant species. Eight rare and two exotic fish species were identified. Combined with analysis of historical data, the richness of fish identified using eDNA metabarcoding was significantly higher than that of fish captured in ground cages. Temperature, pH, and oxidation-reduction potential are the main environmental factors that affect the spatial distribution of fish communities. These results suggest that eDNA metabarcoding could be a new tool with broad application prospects; however, local databases must be improved. This study provides theoretical data and a methodological reference for protecting and managing fish diversity in the Qinling Mountains., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Harmful algal blooms (HABs) have emerged as a critical global environmental and ecological concern. Timely and accurate monitoring of the prevalent bloom-forming genera is crucial for HAB management. Conventional microscope-based methods are time-consuming, labor-intensive, and specialized expertise-dependent, often making them impractical for large-scale surveillance. Molecular methods, such as metabarcoding, provide efficient technical solutions; however, the lack of competent PCR primers and further field validation present obstacles to their wide use. Here, we successfully developed Aphanizomenon-specific primers and validated the application of environmental DNA (eDNA) metabarcoding for field-based monitoring of Aphanizomenon in 37 sites across lentic and lotic freshwater ecosystems in Beijing. The sensitivity and specificity tests of newly developed primers demonstrated high performance - comprehensive recovery of biodiversity in Aphanizomenon communities and high ratios (>95%) of Aphanizomenon sequences in datasets. We observed significant correlations between the sequence abundance derived from eDNA metabarcoding and the total cell density determined through microscopic identification across all the sampling sites, both in the spring (r = 0.8086, p < 0.0001) and summer (r = 0.7902, p < 0.0001), thus validating the utility of eDNA metabarcoding based on the newly developed primers for monitoring in the field. Further, we identified key environmental variables that were primary drivers responsible for the spatiotemporal distribution of Aphanizomenon abundance. These variables included temperature, total nitrogen, and dissolved oxygen in lentic ecosystems, and total phosphorus in lotic ecosystems. The method developed and validated here offers an accurate, efficient, and high-throughput tool for the monitoring of Aphanizomenon blooms in freshwater ecosystems., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Environmental DNA (eDNA) metabarcoding is an emerging tool for monitoring biological communities in aquatic ecosystems. The selection of bioinformatic pipelines significantly impacts the results of biodiversity assessments. However, there is currently no consensus on the appropriate bioinformatic pipelines for fish community analysis in eDNA metabarcoding. In this study, we compared three bioinformatic pipelines (Uparse, DADA2, and UNOISE3) using real and mock (constructed with 15/30 known fish) communities to investigate the differences in biological interpretation during the data analysis process in eDNA metabarcoding. Performance evaluation and diversity analyses revealed that the choice of bioinformatic pipeline could impact the biological results of metabarcoding experiments. Among the three pipelines, the operational taxonomic units (OTU)-based pipeline (Uparse) showed the best performance (sensitivity: 0.6250 ± 0.0166; compositional similarity: 0.4000 ± 0.0571), the highest richness (25-102) and minimal inter-group differences in alpha diversity. It suggested the OTU-based pipeline possessed superior capability in fish diversity monitoring compared to ASV/ZOTU-based pipeline. Additionally, the Bray-Curtis distance matrix achieved the highest discriminative effect in the PCoA (43.3%-53.89%) and inter-group analysis (P < 0.01), indicating it was better at distinguishing compositional differences or specific genera of fish community at different sampling sites than other distance matrices. These findings provide new insights into fish community monitoring through eDNA metabarcoding in estuarine environments., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Transitional waters are important habitats both for biodiversity and ecological functions, providing valuable natural resources and relevant ecosystem services. However, they are highly susceptible to climate changes and anthropogenic pressures responsible for biodiversity losses and require specific biomonitoring programs. Benthic macroinvertebrates are suitable as ecological indicators of transitional waters, being affected by biological, chemical, and physical conditions of the ecosystems about their life cycles and space-use behaviour. The advent of high-throughput sequencing technologies has allowed biodiversity investigations, at the molecular level, in multiple ecosystems and for different ecological guilds. Benthic macroinvertebrate communities' composition has been investigated, at the molecular level, mainly through DNA extracted from sediments in marine and riverine ecosystems. In this work, benthic macroinvertebrate communities are explored through eDNA metabarcoding from water samples in a Mediterranean transitional water ecosystem. This research highlighted the validity of eDNA metabarcoding as an efficient tool for the assessment of benthic macroinvertebrate community structure in transitional waters, unveiling the spatial heterogeneity of benthic macroinvertebrate communities correlated to the measured environmental gradients. The results suggest that peculiar features of transitional water ecosystems, such as shallow waters and limited currents, facilitate the assessment of benthic macroinvertebrate communities through environmental DNA analysis from surface water samples, opening for more rapid and accurate monitoring programs for these valuable ecosystems., (© 2024. The Author(s).)

Terrestrial invertebrates are highly important for the decomposition of dung from large mammals. Mammal dung has been present in many of Earth's ecosystems for millions of years, enabling the evolution of a broad diversity of dung-associated invertebrates that process various components of the dung. Today, large herbivorous mammals are increasingly introduced to ecosystems with the aim of restoring the ecological functions formerly provided by their extinct counterparts. However, we still know little about the ecosystem functions and nutrient flows in these rewilded ecosystems, including the dynamics of dung decomposition. In fact, the succession of insect communities in dung is an area of limited research attention also outside a rewilding context. In this study, we use environmental DNA metabarcoding of dung from rewilded Galloway cattle in an experimental set-up to investigate invertebrate communities and functional dynamics over a time span of 53 days, starting from the time of deposition. We find a strong signal of successional change in community composition, including for the species that are directly dependent on dung as a resource. While several of these species were detected consistently across the sampling period, others appeared confined to either early or late successional stages. We believe that this is indicative of evolutionary adaptation to a highly dynamic resource, with species showing niche partitioning on a temporal scale. However, our results show consistently high species diversity within the functional groups that are directly dependent on dung. Our findings of such redundancy suggest functional stability of the dung-associated invertebrate community, with several species ready to fill vacant niches if other species disappear. Importantly, this might also buffer the ecosystem functions related to dung decomposition against environmental change. Interestingly, alpha diversity peaked after approximately 20-25 days in both meadow and pasture habitats, and did not decrease substantially during the experimental period, probably due to preservation of eDNA in the dung after the disappearance of visiting invertebrates, and from detection of tissue remains and cryptic life stages., (© 2024 The Author(s). Journal of Animal Ecology published by John Wiley & Sons Ltd on behalf of British Ecological Society.)

Environmental DNA (eDNA) integrated with metabarcoding is a promising and powerful tool for species composition and biodiversity assessment in aquatic ecosystems and is increasingly applied to evaluate fish diversity. To date, however, no standardized eDNA-based protocol has been established to monitor fish diversity. In this study, we investigated and compared two filtration methods and three DNA extraction methods using three filtration water volumes to determine a suitable approach for eDNA-based fish diversity monitoring in the Pearl River Estuary (PRE), a highly anthropogenically disturbed estuarine ecosystem. Compared to filtration-based precipitation, direct filtration was a more suitable method for eDNA metabarcoding in the PRE. The combined use of DNeasy Blood and Tissue Kit (BT) and traditional phenol/chloroform (PC) extraction produced higher DNA yields, amplicon sequence variants (ASVs), and Shannon diversity indices, and generated more homogeneous and consistent community composition among replicates. Compared to the other combined protocols, the PC and BT methods obtained better species detection, higher fish diversity, and greater consistency for the filtration water volumes of 1 000 and 2 000 mL, respectively. All eDNA metabarcoding protocols were more sensitive than bottom trawling in the PRE fish surveys and combining two techniques yielded greater taxonomic diversity. Furthermore, combining traditional methods with eDNA analysis enhanced accuracy. These results indicate that methodological decisions related to eDNA metabarcoding should be made with caution for fish community monitoring in estuarine ecosystems.

Freshwater ecosystems have been threatened by complicated disturbances from both natural and anthropogenic variables, especially in dynamic and complex river basins. The environmental DNA (eDNA)-based approach provides a broader spectrum and higher throughput way of biomonitoring for biodiversity assessment compared with traditional morphological survey. Most eDNA metabarcoding studies have been limited to a few specific taxa/groups and habitat scopes. Here we applied the eDNA metabarcoding to characterize the structures and spatial variations of zooplankton and fish communities among different habitat types in a highly dynamic and complex freshwater ecosystem of the Daqing River basin (DRB). The results showed that varied species spectra of zooplankton and fish communities were identified and unique dominant species occurred across habitats. Additionally, markedly spatial distributions of biotic community structures were observed in areas with different habitat characteristics. Natural variables, including geographic distances and gradient ratio, as well as anthropogenic factors of chemical oxygen demand (COD) and organic chemicals demonstrated significant effects but different outcomes on the structures of zooplankton and fish communities. Moreover, the relative abundances of specific aquatic taxa were associated with the gradient of particular environmental variables. This case study verified the distribution patterns and differentiation mechanisms of biotic communities under habitat heterogeneity could be captured by application of eDNA biomonitoring. And habitat-specific and even species-specific environmental stressors would be diagnosed for improving management of complex river basins., (Copyright © 2021. Published by Elsevier Inc.)

Marine biodiversity can be surveyed using underwater visual censuses and recently with eDNA metabarcoding. Although a promising tool, eDNA studies have shown contrasting results related to its detection scale and the number of species identified compared to other survey methods. Also, its accuracy relies on complete reference databases used for taxonomic assignment and, as other survey methods, species detection may show false-negative and false-positive errors. Here, we compared results from underwater visual censuses and simultaneous eDNA metabarcoding fish surveys in terms of observed species and community composition. We also assess the effect of a custom reference database in the taxonomic assignment, and evaluate occupancy, capture and detection probabilities, as well as error rates of eDNA survey data. We amplified a 12S rRNA fish barcode from 24 sampling sites in the gulf of California. More species were detected with eDNA metabarcoding than with UVC. Because each survey method largely detected different sets of species, the combined approach doubled the number of species registered. Both survey methods recovered a known biodiversity gradient and a biogeographic break, but eDNA captured diversity over a broader geographic and bathymetric scale. Furthermore, the use of a modest-sized custom reference database significantly increased taxonomic assignment. In a subset of species, occupancy models revealed eDNA surveys provided similar or higher detection probabilities compared to UVC. The occupancy value of each species had a large influence on eDNA detectability, and in the false positive and negative error. Overall, these results highlight the potential of eDNA metabarcoding in complementing other established ecological methods for studies of marine fishes., (© 2021 John Wiley & Sons Ltd.)

As fish communities are a major concern in rivers ecosystems, we investigated if their environmental (e)DNA signals vary according to the sampling period or hydromorphological conditions. Three rivers were studied over a year using eDNA metabarcoding approach. The majority of the species (c. 80%) were detected all year round in two rivers having similar hydromorphological conditions, whereas in the river affected by an upstream lake waterflow, more species were detected sporadically (42%). For all the rivers, in more than 98% of the occasional detections, the reads abundance represented <0.4% of the total reads per site and per sampling session. Even if the majority of the fish communities remained similar over the year for each of the three rivers, specific seasonal patterns were observed. We studied if the waterflow or the reproduction period had an effect on the observed dynamics. Waterflow, which influences eDNA downstream transportation, had a global influence in taxonomic richness, while the fishes' reproductive period had only an influence on certain species. Our results may help selecting the best sampling strategy according to research objectives. To study fish communities at local scale, seasons of low waterflow periods are recommended. This particularly helps to restraint effects of external eDNA coming from connections with other aquatic environment (tributaries, lakes, wetlands, sewage effluents, etc.). To obtain a more integrative overview of the fish community living in a river basin, high waterflow or breeding seasons are preferable for enhancing species detection probability, especially for rare species., (© 2019 The Fisheries Society of the British Isles.)

Environmental DNA (eDNA) metabarcoding surveys enable rapid, noninvasive identification of taxa from trace samples with wide-ranging applications from characterizing local biodiversity to identifying food-web interactions. However, the technique is prone to error from two major sources: (a) contamination through foreign DNA entering the workflow, and (b) misidentification of DNA within the workflow. Both types of error have the potential to obscure true taxon presence or to increase taxonomic richness by incorrectly identifying taxa as present at sample sites, but multiple error sources can remain unaccounted for in metabarcoding studies. Here, we use data from an eDNA metabarcoding study designed to detect vertebrate species at waterholes in Australia's arid zone to illustrate where and how in the workflow errors can arise, and how to mitigate those errors. We detected the DNA of 36 taxa spanning 34 families, 19 orders and five vertebrate classes in water samples from waterholes, demonstrating the potential for eDNA metabarcoding surveys to provide rapid, noninvasive detection in remote locations, and to widely sample taxonomic diversity from aquatic through to terrestrial taxa. However, we initially identified 152 taxa in the samples, meaning there were many false positive detections. We identified the sources of these errors, allowing us to design a stepwise process to detect and remove error, and provide a template to minimize similar errors that are likely to arise in other metabarcoding studies. Our findings suggest eDNA metabarcoding surveys need to be carefully conducted and screened for errors to ensure their accuracy., (© 2020 John Wiley & Sons Ltd.)

As the largest semi-closed marine ecosystem in the world, the Black Sea has been heavily affected by human activities for a long time. Describing the biodiversity of multi-trophic biota in pelagic zone of the Black Sea and identifying the dominant environmental factors are prerequisites for protecting the sustainability of ecosystems. However, up to now, the taxonomic and distributional information about the Black Sea biota is not clear. Here, we employed a Tree-of-Life metabarcoding to analyze the biodiversity of eight communities in the Black Sea, investigated their biogeographical distribution, and further analyzed the influence of biological and abiotic factors on biota on large scales. We found that, (1) Over 8900 OTUs were detected in the Black Sea, of which 630 species were identified, covering the holistic biota from single-celled (bacteria 5620 OTUs 141 species; algae 1096 OTUs 185 species; protozoa 546 OTUs 146 species) to multicellular organisms (invertebrate metazoans 150 OTUs 34 species; fishes 1369 OTUs 76 species; large marine mammals 39 OTUs 5 species). (2) Higher trophic organisms (fishes and large mammals) distributed more evenly in space than the lower (microorganisms, protozoa and invertebrates). For lower trophic organisms, the vertical stratification was more obvious than the horizontal stratification (vertical p < 0.02, horizontal p < 0.05). (3) The bottom trophic organisms (bacteria and algae) of the food web significantly affected the distribution and composition of the others through biological interactions (Mantel p < 0.05). (4) At the level of abiotic factors, the effect of local species sorting on the composition of communities was 15% higher than that of mass dispersal effect. For the first time, this study monitored and profiled the holistic biodiversity in the pelagic zone of the Black Sea, and provided technological advances and preliminary knowledge for the ongoing Black Sea ecosystem protection efforts., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The environmental DNA (eDNA) metabarcoding represents a new promising tool for biomonitoring and environmental impact assessment. One of the main advantages of eDNA metabarcoding, compared to the traditional morphotaxonomy-based methods, is to provide a more holistic biodiversity information that includes inconspicuous morphologically non-identifiable taxa. Here, we use eDNA metabarcoding to survey marine biodiversity in the vicinity of the three offshore gas platforms in North Adriatic Sea (Italy). We isolated eDNA from 576 water and sediment samples collected at 32 sampling sites situated along four axes at increasing distances from the gas platforms. We obtained about 46 million eDNA sequences for 5 markers from nuclear 18S V1V2, 18S V4, 18S 37F and mitochondrial 16S and COI genes that cover a wide diversity of benthic and planktonic eukaryotes. Our results showed some impact of platform activities on benthic and pelagic communities at very close distance (<50 m), while communities for intermediate (125 m, 250 m, 500 m) and reference (1000 m, 2000 m) sites did not show any particular biodiversity changes that could be related to platforms activities. The most significant community change along the distance gradient was obtained with the 18S V1V2 marker targeting benthic eukaryotes, even though other markers showed similar trends, but to a lesser extent. These results were congruent with the AMBI index inferred from the eDNA sequences assigned to benthic macrofauna. We finally explored the relation between various physicochemical parameters, including hydrocarbons, on benthic community in the case of one of the platforms. Our results showed that these communities were not significantly impacted by most of hydrocarbons, but rather by macro-elements and sediment texture., (Copyright © 2019. Published by Elsevier Ltd.)

Current monitoring methods to assess benthic impacts of marine finfish aquaculture are based on complex biological indices and/or geochemistry data. The former requires benthic macrofauna morpho-taxonomic characterization that is time- and cost-intensive, while the latter provides rapid assessment of the organic enrichment status of sediments but does not directly measure biotic impacts. In this study, sediment samples were collected from seven stations at six salmon farms in British Columbia, Canada, and analyzed for geochemical parameters and by eDNA metabarcoding to investigate linkages between geochemistry and foraminifera. Sediment texture across farm sites ranged from sand to silty loam, while the maximum sediment pore-water sulphide concentration at each site ranged from 1,000 to 13,000 μM. Foraminifera alpha diversity generally increased with distance from cage edge. Adonis analyses revealed that farm site explained the most variation in foraminifera community, followed by sediment type, enrichment status, and distance from cage edge. Farm-specific responses were observed in diversity analyses, taxonomic difference analyses, and correlation analyses. Results demonstrated that species diversity and composition of foraminifera characterized by eDNA metabarcoding generated signals consistent with benthic biodiversity being impacted by finfish farming activities. This substantiates the validity of eDNA metabarcoding for augmenting current approaches to benthic impact assessments by providing more cost-effective and practicable biotic measures than traditional morpho-taxonomy. To capitalize on this potential, further work is needed to design a new nomogram that combines eDNA metabarcoding data and geochemistry data to enable accurate monitoring of benthic impacts of fish farming in a time- and cost-efficient way., (© 2019 John Wiley & Sons Ltd.)

Background: High-throughput amplicon sequencing of environmental DNA (eDNA metabarcoding) has become a routine tool for biodiversity survey and ecological studies. By including sample-specific tags in the primers prior PCR amplification, it is possible to multiplex hundreds of samples in a single sequencing run. The analysis of millions of sequences spread into hundreds to thousands of samples prompts for efficient, automated yet flexible analysis pipelines. Various algorithms and software have been developed to perform one or multiple processing steps, such as paired-end reads assembly, chimera filtering, Operational Taxonomic Unit (OTU) clustering and taxonomic assignment. Some of these software are now well established and widely used by scientists as part of their workflow. Wrappers that are capable to process metabarcoding data from raw sequencing data to annotated OTU-to-sample matrix were also developed to facilitate the analysis for non-specialist users. Yet, most of them require basic bioinformatic or command-line knowledge, which can limit the accessibility to such integrative toolkits. Furthermore, for flexibility reasons, these tools have adopted a step-by-step approach, which can prevent an easy automation of the workflow, and hence hamper the analysis reproducibility., Results: We introduce SLIM, an open-source web application that simplifies the creation and execution of metabarcoding data processing pipelines through an intuitive Graphic User Interface (GUI). The GUI interact with well-established software and their associated parameters, so that the processing steps are performed seamlessly from the raw sequencing data to an annotated OTU-to-sample matrix. Thanks to a module-centered organization, SLIM can be used for a wide range of metabarcoding cases, and can also be extended by developers for custom needs or for the integration of new software. The pipeline configuration (i.e. the modules chaining and all their parameters) is stored in a file that can be used for reproducing the same analysis., Conclusion: This web application has been designed to be user-friendly for non-specialists yet flexible with advanced settings and extensibility for advanced users and bioinformaticians. The source code along with full documentation is available on the GitHub repository ( https://github.com/yoann-dufresne/SLIM ) and a demonstration server is accessible through the application website ( https://trtcrd.github.io/SLIM/ ).

Different second-generation sequencing technologies may have taxon-specific biases when DNA metabarcoding prey in predator faeces. Our major objective was to examine differences in prey recovery from bat guano across two different sequencing workflows using the same faecal DNA extracts. We compared results between the Ion Torrent PGM and the Illumina MiSeq with similar library preparations and the same analysis pipeline. We focus on repeatability and provide an R Notebook in an effort towards transparency for future methodological improvements. Full documentation of each step enhances the accessibility of our analysis pipeline. We tagged DNA from insectivorous bat faecal samples, targeted the arthropod cytochrome c oxidase I minibarcode region and sequenced the product on both second-generation sequencing platforms. We developed an analysis pipeline with a high operational taxonomic unit (OTU) clustering threshold (i.e., ≥98.5%) followed by copy number filtering to avoid merging rare but genetically similar prey into the same OTUs. With this workflow, we detected 297 unique prey taxa, of which 74% were identified at the species level. Of these, 104 (35%) prey OTUs were detected by both platforms, 176 (59%) OTUs were detected by the Illumina MiSeq system only, and 17 (6%) OTUs were detected using the Ion Torrent system only. Costs were similar between platforms but the Illumina MiSeq recovered six times more reads and four additional insect orders than did Ion Torrent. The considerations we outline are particularly important for long-term ecological monitoring; a more standardized approach will facilitate comparisons between studies and allow faster recognition of changes within ecological communities., (© 2018 John Wiley & Sons Ltd.)

We evaluated benthic bacterial communities as bioindicators in environmental impact assessments of salmon aquaculture, a rapidly growing sector of seafood industry. Sediment samples (n=72) were collected from below salmon cages towards distant reference sites. Bacterial community profiles inferred from DNA metabarcodes were compared to reference data from standard macrofauna biomonitoring surveys of the same samples. Deltaproteobacteria were predominant in immediate vicinity of the salmon cages. Along the transect, significant shifts in bacterial community structures were observed with Gammaproteobacteria dominating the less-impacted sites. Alpha- and beta-diversity measures of bacterial communities correlated significantly with macrofauna diversity metrics and with five ecological status indices. Benthic bacterial communities mirror the reaction of macrofauna bioindicators to environmental disturbances caused by salmon farming. The implementation of bacterial eDNA metabarcoding in future Strategic Framework Directives is an alternative cost-effective high-throughput biomonitoring solution, providing a basis for management strategies in a matter of days rather than months., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Among vertebrates, herpetofauna has the highest proportion of declining species. Detection of environmental DNA (eDNA) is a promising method towards significantly increasing large-scale herpetological conservation efforts. However, the integration of eDNA results within a management framework requires an evaluation of the efficiency of the method in large natural environments and the calibration of eDNA surveys with the quantitative monitoring tools currently used by conservation biologists. Towards this end, we first developed species-specific primers to detect the wood turtle (Glyptemys insculpta) a species at risk in Canada, by quantitative PCR (qPCR). The rate of eDNA detection obtained by qPCR was also compared to the relative abundance of this species in nine rivers obtained by standardized visual surveys in the Province of Québec (Canada). Second, we developed multi-species primers to detect North American amphibian and reptile species using eDNA metabarcoding analysis. An occurrence index based on the distribution range and habitat type was compared with the eDNA metabarcoding dataset from samples collected in seven lakes and five rivers. Our results empirically support the effectiveness of eDNA metabarcoding to characterize herpetological species distributions. Moreover, detection rates provided similar results to standardized visual surveys currently used to develop conservation strategies for the wood turtle. We conclude that eDNA detection rates may provide an effective semiquantitative survey tool, provided that assay calibration and standardization is performed.