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Despite the ecological and societal importance of large rivers, fish sampling remains costly and limited to specific habitats (e.g., river banks). Using an eDNA metabarcoding approach, we regularly sampled 500 km of a large river (Rhône River). Comparisons with long-term electrofishing surveys demonstrated the ability of eDNA metabarcoding to qualitatively and quantitatively reveal fish assemblage structures (relative species abundance) but eDNA integrated a larger space than the classical sampling location. Combination of a literature review and field data showed that eDNA behaves in the water column like fine particulate organic matter. Its detection distance varied from a few km in a small stream to more than 100 km in a large river. To our knowledge, our results are the first demonstration of the capacity of eDNA metabarcoding to describe longitudinal fish assemblage patterns in a large river, and metabarcoding appears to be a reliable, cost-effective method for future monitoring.

Biodiversity monitoring delivers vital information to those making conservation decisions. Comprehensively measuring terrestrial biodiversity usually requires costly methods that can rarely be deployed at large spatial scales over multiple time periods, limiting conservation efficiency. Here we investigated the capacity of environmental DNA (eDNA) from stream water samples to survey terrestrial mammal diversity at multiple spatial scales within a large catchment. We compared biodiversity information recovered using an eDNA metabarcoding approach with data from a dense camera trap survey, as well as the sampling costs of both methods. Via the sampling of large volumes of water from the two largest streams that drained the study area, eDNA metabarcoding provided information on the presence and detection probabilities of 35 mammal taxa, 25% more than camera traps and for half the cost. While eDNA metabarcoding had limited capacity to detect felid species and provide individual-level demographic information, it is a cost-efficient method for large-scale monitoring of terrestrial mammals that can offer sufficient information to solve many conservation problems.

Repeated introductions and spread of invasive mosquito species (IMS) have been recorded on a large scale these last decades worldwide. In this context, members of the mosquito genus Aedes can present serious risks to public health as they have or may develop vector competence for various viral diseases. While the Tiger mosquito (Aedes albopictus) is a well-known vector for e.g. dengue and chikungunya viruses, the Asian bush mosquito (Ae. j. japonicus) and Ae. koreicus have shown vector competence in the field and the laboratory for a number of viruses including dengue, West Nile fever and Japanese encephalitis. Early detection and identification is therefore crucial for successful eradication or control strategies. Traditional specific identification and monitoring of different and/or cryptic life stages of the invasive Aedes species based on morphological grounds may lead to misidentifications, and are problematic when extensive surveillance is needed. In this study, we developed, tested and applied an environmental DNA (eDNA) approach for the detection of three IMS, based on water samples collected in the field in several European countries. We compared real-time quantitative PCR (qPCR) assays specific for these three species and an eDNA metabarcoding approach with traditional sampling, and discussed the advantages and limitations of these methods. Detection probabilities for eDNA-based approaches were in most of the specific comparisons higher than for traditional survey and the results were congruent between both molecular methods, confirming the reliability and efficiency of alternative eDNA-based techniques for the early and unambiguous detection and surveillance of invasive mosquito vectors. The ease of water sampling procedures in the eDNA approach tested here allows the development of large-scale monitoring and surveillance programs of IMS, especially using citizen science projects.

The precise knowledge of species distribution is a key step in conservation biology. However, species detection can be extremely difficult in many environments, specific life stages and in populations at very low density. The aim of this study was to improve the knowledge on DNA persistence in water in order to confirm the presence of the focus species in freshwater ecosystems. Aquatic vertebrates (fish: Siberian sturgeon and amphibian: Bullfrog tadpoles) were used as target species. In control conditions (tanks) and in the field (ponds), the DNA detectability decreases with time after the removal of the species source of DNA. DNA was detectable for less than one month in both conditions. The density of individuals also influences the dynamics of DNA detectability in water samples. The dynamics of detectability reflects the persistence of DNA fragments in freshwater ecosystems. The short time persistence of detectable amounts of DNA opens perspectives in conservation biology, by allowing access to the presence or absence of species e.g. rare, secretive, potentially invasive, or at low density. This knowledge of DNA persistence will greatly influence planning of biodiversity inventories and biosecurity surveys.

Information is scarce on how environmental and dispersal processes interact with biological features of the organisms, such as their habitat affinity, to influence patterns in biodiversity. We examined the role of habitat specialist vs. generalist species, and the spatial configuration, connectivity, and different environmental characteristics of river-floodplain habitats to get a more mechanistic understanding of alpha and beta diversity of fish metacommunities. We used environmental DNA metabarcoding to characterize species (taxa) richness and composition in two separate floodplains of the river Danube (Austria and Hungary) during two different hydrological conditions. Results showed that differences in the number of generalist and specialist species and their responses to connectivity and environmental gradients influenced patterns in alpha and beta diversity. Of the components of beta diversity, richness difference (nestedness) showed consistently higher values than replacement (turnover), mainly due to the decrease of specialist species along the connectivity gradient (i.e., from the mainstem to the most isolated oxbows). Variance in both alpha and beta diversity could be well predicted by a set of local and regional variables, despite high environmental variability, which characterizes river-floodplain ecosystems. Of these, the joint or shared variance fractions proved to be the most important, which indicates that the effects of local and regional processes cannot be unambiguously separated in these river-floodplain systems. Local scale environmental variables were more important determinants of both alpha and beta diversity in the low water period than in the high water period. These results indicate the differential role of local and regional processes in community organization during different hydrological conditions. Maintenance of both local and regional scale processes are thus important in the preservation of alpha and beta diversity of floodplain fish metacommunities, which should be considered by environmental management.

Innovative techniques, such as environmental DNA (eDNA) metabarcoding, are now promoting broader biodiversity monitoring at unprecedented scales, because of the reduction in time, presumably lower cost, and methodological efficiency. Our goal was to assess the efficiency of established inventory techniques (live-trapping grids, pitfall traps, camera trapping, mist netting) as well as eDNA for detecting Amazonian mammals. For terrestrial small mammals, we used 32 live-trapping grids based on Sherman and Tomahawk traps (total effort of 10,368 trap-nights); in addition to 16 pitfall traps (1,408 trap-nights). For bats, we used mist nets at 8 sites (4,800 net hours). For medium and large mammals, we used 72 camera trap stations (5,208 camera-days). We identified vertebrate and mammal taxa based on eDNA analysis (12S region, with V05 and Mamm01 markers) from water samples, including a total of 11 3-km transects for stagnant water sampling and seven small streams for running water sampling. A total of 106 mammal species were recorded. Building on sample-based rarefaction and extrapolation curves, both trapping grids and pitfall were successful, recording 91.16% and 82.1% of the expected species for these techniques (~22 and ~9 species), and 16.98% and 6.60% of the total recorded mammal species, respectively. Mist nets recorded 83.2% of the expected bat species (~48), and 34.91% of the total recorded species. Camera trapping recorded 99.2% of the predicted large- and medium-sized species (~31), and 33.02% of the total recorded species. eDNA recorded 75.4% of the expected mammal species for this technique (~68), and 47.0% of the total recorded species. eDNA resulted in a useful tool that saves on effort and reduces sampling costs. This study is among the first to show the large potential of eDNA metabarcoding for assessing Amazonian mammal communities, providing, in combination with conventional techniques, a rapid overview of mammal diversity with broad applications to monitoring, management and conservation. By including appropriate genetic markers and updated reference databases, eDNA metabarcoding method can be extended to the whole vertebrate community.

In the last few years, the study of environmental DNA (eDNA) has drawn attention for many reasons, including its advantages for monitoring and conservation purposes. So far, in aquatic environments, most of eDNA research has focused on the detection of single species using species-specific markers. Recently, species inventories based on the analysis of a single generalist marker targeting a larger taxonomic group (eDNA metabarcoding) have proven useful for bony fish and amphibian biodiversity surveys. This approach involves in situ filtering of large volumes of water followed by amplification and sequencing of a short discriminative fragment from the 12S rDNA mitochondrial gene. In this study, we went one step further by investigating the spatial representativeness (i.e. ecological reliability and signal variability in space) of eDNA metabarcoding for large-scale fish biodiversity assessment in a freshwater system including lentic and lotic environments. We tested the ability of this approach to characterize large-scale organization of fish communities along a longitudinal gradient, from a lake to the outflowing river. First, our results confirm that eDNA metabarcoding is more efficient than a single traditional sampling campaign to detect species presence, especially in rivers. Second, the species list obtained using this approach is comparable to the one obtained when cumulating all traditional sampling sessions since 1995 and 1988 for the lake and the river, respectively. In conclusion, eDNA metabarcoding gives a faithful description of local fish biodiversity in the study system, more specifically within a range of a few kilometers along the river in our study conditions, i.e. longer than a traditional fish sampling site.

Although we are currently experiencing worldwide biodiversity loss, local species richness does not always decline under anthropogenic pressure. This conservation paradox may also apply in protected areas but has not yet received conclusive evidence in marine ecosystems. Here, we survey fish assemblages in six Mediterranean no-take reserves and their adjacent fishing grounds using environmental DNA (eDNA) while controlling for environmental conditions. We detect less fish species in marine reserves than in nearby fished areas. The paradoxical gradient in species richness is accompanied by a marked change in fish species composition under different managements. This dissimilarity is mainly driven by species that are often overlooked by classical visual surveys but detected with eDNA: cryptobenthic, pelagic, and rare fishes. These results do not negate the importance of reserves in protecting biodiversity but shed new light on how under-represented species groups can positively react to fishing pressure and how conservation efforts can shape regional biodiversity patterns.

Assessing the impact of human activity on ecosystems often links local biodiversity to disturbances measured within the same locality. However, remote disturbances may also affect local biodiversity. Here, we used environmental DNA metabarcoding to evaluate the relationships between vertebrate biodiversity (fish and mammals) and disturbance intensity in two Amazonian rivers. Measurements of anthropic disturbance -here forest cover losses- were made from the immediate vicinity of the biodiversity sampling sites to up to 90 km upstream. The findings suggest that anthropization had a spatially extended impact on biodiversity. Forest cover losses of <11% in areas up to 30 km upstream from the biodiversity sampling sites were linked to reductions of >22% in taxonomic and functional richness of both terrestrial and aquatic fauna. This underscores the vulnerability of Amazonian biodiversity even to low anthropization levels. The similar responses of aquatic and terrestrial fauna to remote disturbances indicate the need for cross-ecosystem conservation plans that consider the spatially extended effects of anthropization. It is unclear how far the impact of deforestation can spread. Here the authors analyse freshwater eDNA data along two rivers in the Amazon forest, and find that low levels of deforestation are linked to substantial reductions of fish and mammalian diversity downstream.

Freshwater ecosystems are the most vulnerable worldwide and freshwater bivalves rank amongst the most threatened animals in the world. Surveying and monitoring freshwater bivalves are difficult tasks: they are difficult to find, hard to identify (taxonomic expertise is needed), and working underwater is technically challenging. It is therefore crucial to find more efficient methods to survey and monitor these species. Here, we present the first metabarcoding approach for freshwater bivalves and compare environmental DNA (eDNA) and traditional surveys. We describe two sets of primers (for Unionida and Venerida) developed for freshwater bivalves eDNA metabarcoding. These primers have been tested in the field, with about 300 studied sites. Results were compared to freshwater bivalves’ surveys using traditional methods, with eDNA always detecting more species than traditional surveys, especially when Sphaerids were taken into account. While our study initially focused on Western Palearctic freshwater bivalve species, our primers were confronted in silico with available sequences and have proven to be effective at a global scale. The results show that eDNA metabarcoding, with our developed primers, is a remarkable tool allowing for non-invasive surveys, detection of rare and inconspicuous species, absence data and overall freshwater bivalves routine monitoring.