An integrated spatio-temporal view of riverine biodiversity using environmental DNA metabarcoding.

Anthropogenically forced changes in global freshwater biodiversity demand more efficient monitoring approaches. Consequently, environmental DNA (eDNA) analysis is enabling ecosystem-scale biodiversity assessment, yet the appropriate spatio-temporal resolution of robust biodiversity assessment remains ambiguous. Here, using intensive, spatio-temporal eDNA sampling across space (five rivers in Europe and North America, with an upper range of 20-35 km between samples), time (19 timepoints between 2017 and 2018) and environmental conditions (river flow, pH, conductivity, temperature and rainfall), we characterise the resolution at which information on diversity across the animal kingdom can be gathered from rivers using eDNA. In space, beta diversity was mainly dictated by turnover, on a scale of tens of kilometres, highlighting that diversity measures are not confounded by eDNA from upstream. Fish communities showed nested assemblages along some rivers, coinciding with habitat use. Across time, seasonal life history events, including salmon and eel migration, were detected. Finally, effects of environmental conditions were taxon-specific, reflecting habitat filtering of communities rather than effects on DNA molecules. We conclude that riverine eDNA metabarcoding can measure biodiversity at spatio-temporal scales relevant to species and community ecology, demonstrating its utility in delivering insights into river community ecology during a time of environmental change.

Significant improvement in freshwater invertebrate biodiversity in all types of English rivers over the past 30 years.

There remains a persistent concern that freshwater biodiversity is in decline and being threatened by pollution. As the UK, and particularly England, is a densely populated nation with rivers of modest dilution capacity, this location is very suitable to examine how freshwater biodiversity has responded to human pressures over the past 30 years. A long-term dataset of 223,325 freshwater macroinvertebrate records from 1989 to 2018 for England was retrieved and examined. A sub-set of approximately 200 sites per English Region (1515 sites in total with 62,514 samples), with the longest and most consistent records were matched with predicted wastewater exposure, upstream land cover and terrain characteristics (latitude, altitude, slope gradient and flow discharge). To understand changes in macroinvertebrate diversity and sensitivity with respect to these parameters, the biotic indices of (i) overall family richness, (ii) Ephemeroptera, Plecoptera, Trichoptera (EPT) family richness, and (iii) the Biological Monitoring Working Party (BMWP) scores of NTAXA (number of scoring taxa) and (iv) ASPT (average score per taxon) were selected. A review of how close the BMWP scores come to those expected at minimally impacted reference sites was included. For all latitudes, altitudes, channel slope, river size, wastewater exposure levels, and differing proportions of upstream woodland, seminatural, arable and urban land cover, all diversity or sensitivity indices examined improved over this period, although this improvement has slowed in some cases post 2003. Mean overall family richness has increased from 15 to 25 family groups, a 66 % improvement. The improvement in mean EPT family richness (3 to 10 families, >300 % improvement), which are considered to be particularly sensitive to pollution, implies macroinvertebrate diversity has benefited from a national improvement in critical components of water quality.

Patterns of invertebrate functional diversity highlight the vulnerability of ecosystem services over a 45-year period.

Declines in invertebrate biodiversity1,2 pose a significant threat to key ecosystem services.3-5 Current analyses of biodiversity often focus on taxonomic diversity (e.g., species richness),6,7 which does not account for the functional role of a species. Functional diversity of species' morphological or behavioral traits is likely more relevant to ecosystem service delivery than taxonomic diversity, as functional diversity has been found to be a key driver of a number of ecosystem services including decomposition and pollination.8-12 At present, we lack a good understanding of long-term and large-scale changes in functional diversity, which limits our capacity to determine the vulnerability of key ecosystem services with ongoing biodiversity change. Here we derive trends in functional diversity and taxonomic diversity over a 45-year period across Great Britain for species supporting freshwater aquatic functions, pollination, natural pest control, and agricultural pests (a disservice). Species supporting aquatic functions showed a synchronous collapse and recovery in functional and taxonomic diversity. In contrast, pollinators showed an increase in taxonomic diversity, but a decline and recovery in functional diversity. Pest control agents and pests showed greater stability in functional diversity over the assessment period. We also found that functional diversity could appear stable or show patterns of recovery, despite ongoing changes in the composition of traits among species. Our results suggest that invertebrate assemblages can show considerable variability in their functional structure over time at a national scale, which provides an important step in determining the long-term vulnerability of key ecosystem services with ongoing biodiversity change.

Environmental DNA provides higher resolution assessment of riverine biodiversity and ecosystem function via spatio-temporal nestedness and turnover partitioning.

Rapidly assessing biodiversity is essential for environmental monitoring; however, traditional approaches are limited in the scope needed for most ecological systems. Environmental DNA (eDNA) based assessment offers enhanced scope for assessing biodiversity, while also increasing sampling efficiency and reducing processing time, compared to traditional methods. Here we investigated the effects of landuse and seasonality on headwater community richness and functional diversity, via spatio-temporal dynamics, using both eDNA and traditional sampling. We found that eDNA provided greater resolution in assessing biodiversity dynamics in time and space, compared to traditional sampling. Community richness was seasonally linked, peaking in spring and summer, with temporal turnover having a greater effect on community composition compared to localized nestedness. Overall, our assessment of ecosystem function shows that community formation is driven by regional resource availability, implying regional management requirements should be considered. Our findings show that eDNA based ecological assessment is a powerful, rapid and effective assessment strategy that enables complex spatio-temporal studies of community diversity and ecosystem function, previously infeasible using traditional methods.

Trait-based ecology at large scales: Assessing functional trait correlations, phylogenetic constraints and spatial variability using open data.

The growing use of functional traits in ecological research has brought new insights into biodiversity responses to global environmental change. However, further progress depends on overcoming three major challenges involving (a) statistical correlations between traits, (b) phylogenetic constraints on the combination of traits possessed by any single species, and (c) spatial effects on trait structure and trait-environment relationships. Here, we introduce a new framework for quantifying trait correlations, phylogenetic constraints and spatial variability at large scales by combining openly available species' trait, occurrence and phylogenetic data with gridded, high-resolution environmental layers and computational modelling. Our approach is suitable for use among a wide range of taxonomic groups inhabiting terrestrial, marine and freshwater habitats. We demonstrate its application using freshwater macroinvertebrate data from 35 countries in Europe. We identified a subset of available macroinvertebrate traits, corresponding to a life-history model with axes of resistance, resilience and resource use, as relatively unaffected by correlations and phylogenetic constraints. Trait structure responded more consistently to environmental variation than taxonomic structure, regardless of location. A re-analysis of existing data on macroinvertebrate communities of European alpine streams supported this conclusion, and demonstrated that occurrence-based functional diversity indices are highly sensitive to the traits included in their calculation. Overall, our findings suggest that the search for quantitative trait-environment relationships using single traits or simple combinations of multiple traits is unlikely to be productive. Instead, there is a need to embrace the value of conceptual frameworks linking community responses to environmental change via traits which correspond to the axes of life-history models. Through a novel integration of tools and databases, our flexible framework can address this need.

Executing multi-taxa eDNA ecological assessment via traditional metrics and interactive networks.

Current approaches to ecological assessment are limited by the traditional morpho-taxonomic methods presently employed and the inability to meet increasing demands for rapid assessments. Advancements in high throughput sequencing now enable rapid high-resolution ecological assessment using environmental DNA (eDNA). Here we test the ability of using eDNA-based ecological assessment methods against traditional assessment of two key indicator groups (diatoms and macroinvertebrates) and show how eDNA across multiple gene regions (COI, rbcL, 12S and 18S) can be used to infer interactive networks that link to ecological assessment criteria. We compared results between taxonomic and eDNA based assessments and found significant positive associations between macroinvertebrate (p < 0.001 R2 = 0.645) and diatom (p = 0.015, R2 = 0.222) assessment metrics. We further assessed the ability of eDNA based assessment to identify environmentally sensitive genera and found an order of magnitude greater potential for 18S, versus COI or rbcL, to determine environmental filtering of ecologically assessed communities. Lastly, we compared the ability of traditional metrics against co-occurrence network properties of our combined 18S, COI and rbcL indicator genera to infer habitat quality measures currently used by managers. We found that transitivity (network connectivity), linkage density and cohesion were significantly associated with habitat modification scores (HMS), whereas network properties were inconsistent with linking to the habitat quality score (HQS) metric. The incorporation of multi-marker eDNA network assessment opens up a means for finer scale ecological assessment, currently limited using traditional methods. While utilization of eDNA-based assessment is recommended, direct comparisons with traditional approaches are difficult as the methods are intrinsically different and should be treated as such with regards to future research. Overall, our findings show that eDNA can be used for effective ecological assessment while offering a wider range of scope and application compared to traditional assessment methods.

What Works? the Influence of Changing Wastewater Treatment Type, Including Tertiary Granular Activated Charcoal, on Downstream Macroinvertebrate Biodiversity Over Time.

The present study reviewed the impacts of wastewater on macroinvertebrates over 4 decades in a United Kingdom lowland river. This involved examining changes in chemicals, temperature, flow, and macroinvertebrate diversity from the 1970s until 2017 for a wastewater-dominated river downstream of Swindon in the United Kingdom (population ~ 220 000). When the wastewater treatment process changed from trickling filter to activated sludge in 1991, biological oxygen demand was nearly halved (90th percentile from 8.1 to 4.6 mg/L), ammonia peaks dropped more than 7-fold (90th percentile from 3.9 to 0.53 mg/L), whereas dissolved oxygen climbed consistently above 60% saturation (10th percentile from 49 to 64%) at a sampling point 2 km downstream of the wastewater treatment plant. A sustained increase in the number of macroinvertebrate species was evident from that point. River flow did not change, temperature rose slightly, and the major metal concentrations declined steadily over most of the monitoring period. Neither the introduction of phosphate stripping in 1999 nor the use of tertiary granular activated charcoal from 2008 to 2014 had strong positive effects on subsequent macroinvertebrate diversity. That the diversity still had not reached the ideal status by 2016 may be related to the modest habitat quality, agricultural pesticides, and limited recolonization potential in the catchment. The results indicate that urban wastewaters, with their chemical pollutants, are today probably not the biggest threat to the macroinvertebrate diversity of multiply stressed lowland rivers in the United Kingdom. Environ Toxicol Chem 2019;38:1820-1832. © 2019 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals, Inc. on behalf of SETAC.

Acidity promotes degradation of multi-species environmental DNA in lotic mesocosms.

Accurate quantification of biodiversity is fundamental to understanding ecosystem function and for environmental assessment. Molecular methods using environmental DNA (eDNA) offer a non-invasive, rapid, and cost-effective alternative to traditional biodiversity assessments, which require high levels of expertise. While eDNA analyses are increasingly being utilized, there remains considerable uncertainty regarding the dynamics of multispecies eDNA, especially in variable systems such as rivers. Here, we utilize four sets of upland stream mesocosms, across an acid-base gradient, to assess the temporal and environmental degradation of multispecies eDNA. Sampling included water column and biofilm sampling over time with eDNA quantified using qPCR. Our findings show that the persistence of lotic multispecies eDNA, sampled from water and biofilm, decays to non-detectable levels within 2 days and that acidic environments accelerate the degradation process. Collectively, the results provide the basis for a predictive framework for the relationship between lotic eDNA degradation dynamics in spatio-temporally dynamic river ecosystems.

Bending the rules: exploitation of allochthonous resources by a top-predator modifies size-abundance scaling in stream food webs.

Body mass-abundance (M-N) allometries provide a key measure of community structure, and deviations from scaling predictions could reveal how cross-ecosystem subsidies alter food webs. For 31 streams across the UK, we tested the hypothesis that linear log-log M-N scaling is shallower than that predicted by allometric scaling theory when top predators have access to allochthonous prey. These streams all contained a common and widespread top predator (brown trout) that regularly feeds on terrestrial prey and, as hypothesised, deviations from predicted scaling increased with its dominance of the fish assemblage. Our study identifies a key beneficiary of cross-ecosystem subsidies at the top of stream food webs and elucidates how these inputs can reshape the size-structure of these 'open' systems.

Phosphorus and nitrogen limitation and impairment of headwater streams relative to rivers in Great Britain: A national perspective on eutrophication.

This study provides a first national-scale assessment of the nutrient status of British headwater streams within the wider river network, by joint analysis of the national Countryside Survey Headwater Stream and Harmonised River Monitoring Scheme datasets. We apply a novel Nutrient Limitation Assessment methodology to explore the extent to which nutrients may potentially limit primary production in headwater streams and rivers, by coupling ternary assessment of nitrogen (N), phosphorus (P), and carbon (C) depletion, with N:P stoichiometry, and threshold P and N concentrations. P limitation was more commonly seen in the rivers, with greater prevalence of N limitation in the headwater streams. High levels of potential P and N co-limitation were found in the headwater streams, especially the Upland-Low-Alkalinity streams. This suggests that managing both P and N inputs may be needed to minimise risks of degradation of these sensitive headwater stream environments. Although localised nutrient impairment of headwater streams can occur, there were markedly lower rates of P and N impairment of headwater streams relative to downstream rivers at the national scale. Nutrient source contributions, relative to hydrological dilution, increased with catchment scale, corresponding with increases in the extent of agricultural and urban land-use. The estimated nutrient reductions needed to achieve compliance with Water Framework Directive standards, and to reach limiting concentrations, were greatest for the Lowland-High-Alkalinity rivers and streams. Preliminary assessments suggest that reducing P concentrations in the Lowland-High-Alkalinity headwater streams, and N concentrations in the Upland-Low-Alkalinity rivers, might offer greater overall benefits for water-quality remediation at the national scale, relative to the magnitude of nutrient reductions required. This approach could help inform the prioritisation of nutrient remediation, as part of a directional approach to water quality management based on closing the gaps between current and target nutrient concentrations.

Horizon scanning for invasive alien species with the potential to threaten biodiversity in Great Britain.

Invasive alien species (IAS) are considered one of the greatest threats to biodiversity, particularly through their interactions with other drivers of change. Horizon scanning, the systematic examination of future potential threats and opportunities, leading to prioritization of IAS threats is seen as an essential component of IAS management. Our aim was to consider IAS that were likely to impact on native biodiversity but were not yet established in the wild in Great Britain. To achieve this, we developed an approach which coupled consensus methods (which have previously been used for collaboratively identifying priorities in other contexts) with rapid risk assessment. The process involved two distinct phases: Preliminary consultation with experts within five groups (plants, terrestrial invertebrates, freshwater invertebrates, vertebrates and marine species) to derive ranked lists of potential IAS. Consensus-building across expert groups to compile and rank the entire list of potential IAS. Five hundred and ninety-one species not native to Great Britain were considered. Ninety-three of these species were agreed to constitute at least a medium risk (based on score and consensus) with respect to them arriving, establishing and posing a threat to native biodiversity. The quagga mussel, Dreissena rostriformis bugensis, received maximum scores for risk of arrival, establishment and impact; following discussions the unanimous consensus was to rank it in the top position. A further 29 species were considered to constitute a high risk and were grouped according to their ranked risk. The remaining 63 species were considered as medium risk, and included in an unranked long list. The information collated through this novel extension of the consensus method for horizon scanning provides evidence for underpinning and prioritizing management both for the species and, perhaps more importantly, their pathways of arrival. Although our study focused on Great Britain, we suggest that the methods adopted are applicable globally.

Using boreholes as windows into groundwater ecosystems.

Groundwater ecosystems remain poorly understood yet may provide ecosystem services, make a unique contribution to biodiversity and contain useful bio-indicators of water quality. Little is known about ecosystem variability, the distribution of invertebrates within aquifers, or how representative boreholes are of aquifers. We addressed these issues using borehole imaging and single borehole dilution tests to identify three potential aquifer habitats (fractures, fissures or conduits) intercepted by two Chalk boreholes at different depths beneath the surface (34 to 98 m). These habitats were characterised by sampling the invertebrates, microbiology and hydrochemistry using a packer system to isolate them. Samples were taken with progressively increasing pumped volume to assess differences between borehole and aquifer communities. The study provides a new conceptual framework to infer the origin of water, invertebrates and microbes sampled from boreholes. It demonstrates that pumping 5 m(3) at 0.4-1.8 l/sec was sufficient to entrain invertebrates from five to tens of metres into the aquifer during these packer tests. Invertebrates and bacteria were more abundant in the boreholes than in the aquifer, with associated water chemistry variations indicating that boreholes act as sites of enhanced biogeochemical cycling. There was some variability in invertebrate abundance and bacterial community structure between habitats, indicating ecological heterogeneity within the aquifer. However, invertebrates were captured in all aquifer samples, and bacterial abundance, major ion chemistry and dissolved oxygen remained similar. Therefore the study demonstrates that in the Chalk, ecosystems comprising bacteria and invertebrates extend from around the water table to 70 m below it. Hydrogeological techniques provide excellent scope for tackling outstanding questions in groundwater ecology, provided an appropriate conceptual hydrogeological understanding is applied.

Climate change impacts in multispecies systems: drought alters food web size structure in a field experiment.

Experimental data from intergenerational field manipulations of entire food webs are scarce, yet such approaches are essential for gauging impacts of environmental change in natural systems. We imposed 2 years of intermittent drought on stream channels in a replicated field trial, to measure food web responses to simulated climate change. Drought triggered widespread losses of species and links, with larger taxa and those that were rare for their size, many of which were predatory, being especially vulnerable. Many network properties, including size-scaling relationships within food chains, changed in response to drought. Other properties, such as connectance, were unaffected. These findings highlight the need for detailed experimental data from different organizational levels, from pairwise links to the entire food web. The loss of not only large species, but also those that were rare for their size, provides a newly refined way to gauge likely impacts that may be applied more generally to other systems and/or impacts.

Food web complexity and allometric scaling relationships in stream mesocosms: implications for experimentation.

1. Mesocosms are used extensively by ecologists to gain a mechanistic understanding of ecosystems based on the often untested assumption that these systems can replicate the key attributes of natural assemblages. 2. Previous investigations of stream mesocosm utility have explored community composition, but here for the first time, we extend the approach to consider the replicability and realism of food webs in four outdoor channels (4 m(2)). 3. The four food webs were similarly complex, consisting of diverse assemblages (61-71 taxa) with dense feeding interactions (directed connectance 0.09-0.11). Mesocosm food web structural attributes were within the range reported for 82 well-characterized food webs from natural streams and rivers. When compared with 112 additional food webs from standing freshwater, marine, estuarine and terrestrial environments, stream food webs (including mesocosms) had similar characteristic path lengths, but typically lower mean food chain length and exponents for the species-link relationship. 4. Body size (M) abundance (N) allometric scaling coefficients for trivariate taxonomic mesocosm food webs (-0.53 to -0.49) and individual size distributions (-0.60 to -0.58) were consistent and similar to those from natural systems, suggesting that patterns of energy flux between mesocosm consumers and resources were realistic approximations. 5. These results suggest that stream mesocosms of this scale can support replicate food webs with a degree of biocomplexity that is comparable to 'natural' streams. The findings highlight the potential value of mesocosms as model systems for performing experimental manipulations to test ecological theories, at spatiotemporal scales of relevance to natural ecosystems.

Ecological networks--beyond food webs.

1. A fundamental goal of ecological network research is to understand how the complexity observed in nature can persist and how this affects ecosystem functioning. This is essential for us to be able to predict, and eventually mitigate, the consequences of increasing environmental perturbations such as habitat loss, climate change, and invasions of exotic species. 2. Ecological networks can be subdivided into three broad types: 'traditional' food webs, mutualistic networks and host-parasitoid networks. There is a recent trend towards cross-comparisons among network types and also to take a more mechanistic, as opposed to phenomenological, perspective. For example, analysis of network configurations, such as compartments, allows us to explore the role of co-evolution in structuring mutualistic networks and host-parasitoid networks, and of body size in food webs. 3. Research into ecological networks has recently undergone a renaissance, leading to the production of a new catalogue of evermore complete, taxonomically resolved, and quantitative data. Novel topological patterns have been unearthed and it is increasingly evident that it is the distribution of interaction strengths and the configuration of complexity, rather than just its magnitude, that governs network stability and structure. 4. Another significant advance is the growing recognition of the importance of individual traits and behaviour: interactions, after all, occur between individuals. The new generation of high-quality networks is now enabling us to move away from describing networks based on species-averaged data and to start exploring patterns based on individuals. Such refinements will enable us to address more general ecological questions relating to foraging theory and the recent metabolic theory of ecology. 5. We conclude by suggesting a number of 'dead ends' and 'fruitful avenues' for future research into ecological networks.

Consumer-resource body-size relationships in natural food webs.

It has been suggested that differences in body size between consumer and resource species may have important implications for interaction strengths, population dynamics, and eventually food web structure, function, and evolution. Still, the general distribution of consumer-'resource body-size ratios in real ecosystems, and whether they vary systematically among habitats or broad taxonomic groups, is poorly understood. Using a unique global database on consumer and resource body sizes, we show that the mean body-size ratios of aquatic herbivorous and detritivorous consumers are several orders of magnitude larger than those of carnivorous predators. Carnivorous predator-prey body-size ratios vary across different habitats and predator and prey types (invertebrates, ectotherm, and endotherm vertebrates). Predator-prey body-size ratios are on average significantly higher (1) in freshwater habitats than in marine or terrestrial habitats, (2) for vertebrate than for invertebrate predators, and (3) for invertebrate than for ectotherm vertebrate prey. If recent studies that relate body-size ratios to interaction strengths are general, our results suggest that mean consumer-resource interaction strengths may vary systematically across different habitat categories and consumer types.