Measuring the state of aquatic environments using eDNA-upscaling spatial resolution of biotic indices.

Aquatic macroinvertebrates, including many aquatic insect orders, are a diverse and ecologically relevant organismal group yet they are strongly affected by anthropogenic activities. As many of these taxa are highly sensitive to environmental change, they offer a particularly good early warning system for human-induced change, thus leading to their intense monitoring. In aquatic ecosystems there is a plethora of biotic monitoring or biomonitoring approaches, with more than 300 assessment methods reported for freshwater taxa alone. Ultimately, monitoring of aquatic macroinvertebrates is used to calculate ecological indices describing the state of aquatic systems. Many of the methods and indices used are not only hard to compare, but especially difficult to scale in time and space. Novel DNA-based approaches to measure the state and change of aquatic environments now offer unprecedented opportunities, also for possible integration towards commonly applicable indices. Here, we first give a perspective on DNA-based approaches in the monitoring of aquatic organisms, with a focus on aquatic insects, and how to move beyond traditional point-based biotic indices. Second, we demonstrate a proof-of-concept for spatially upscaling ecological indices based on environmental DNA, demonstrating how integration of these novel molecular approaches with hydrological models allows an accurate evaluation at the catchment scale. This article is part of the theme issue 'Towards a toolkit for global insect biodiversity monitoring'.

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.

Foundation species stabilize an alternative eutrophic state in nutrient-disturbed ponds via selection on microbial community.

Eutrophication due to nutrient addition can result in major alterations in aquatic ecosystem productivity. Foundation species, individually and interactively, whether present as invasive species or as instruments of ecosystem management and restoration, can have unwanted effects like stabilizing turbid eutrophic states. In this study, we used whole-pond experimental manipulations to investigate the impacts of disturbance by nutrient additions in the presence and absence of two foundation species: Dreissena polymorpha (a freshwater mussel) and Myriophyllum spicatum (a macrophyte). We tracked how nutrient additions to ponds changed the prokaryotic and eukaryotic communities, using 16S, 18S, and COI amplicon sequencing. The nutrient disturbance and foundation species imposed strong selection on the prokaryotic communities, but not on the microbial eukaryotic communities. The prokaryotic communities changed increasingly over time as the nutrient disturbance intensified. Post-disturbance, the foundation species stabilized the prokaryotic communities as observed by the reduced rate of change in community composition. Our analysis suggests that prokaryotic community change contributed both directly and indirectly to major changes in ecosystem properties, including pH and dissolved oxygen. Our work shows that nutrient disturbance and foundation species strongly affect the prokaryotic community composition and stability, and that the presence of foundation species can, in some cases, promote the emergence and persistence of a turbid eutrophic ecosystem state.

Environmental DNA: The next chapter.

Molecular tools are an indispensable part of ecology and biodiversity sciences and implemented across all biomes. About a decade ago, the use and implementation of environmental DNA (eDNA) to detect biodiversity signals extracted from environmental samples opened new avenues of research. Initial eDNA research focused on understanding population dynamics of target species. Its scope thereafter broadened, uncovering previously unrecorded biodiversity via metabarcoding in both well-studied and understudied ecosystems across all taxonomic groups. The application of eDNA rapidly became an established part of biodiversity research, and a research field by its own. Here, we revisit key expectations made in a land-mark special issue on eDNA in Molecular Ecology in 2012 to frame the development in six key areas: (1) sample collection, (2) primer development, (3) biomonitoring, (4) quantification, (5) behaviour of DNA in the environment and (6) reference database development. We pinpoint the success of eDNA, yet also discuss shortfalls and expectations not met, highlighting areas of research priority and identify the unexpected developments. In parallel, our retrospective couples a screening of the peer-reviewed literature with a survey of eDNA users including academics, end-users and commercial providers, in which we address the priority areas to focus research efforts to advance the field of eDNA. With the rapid and ever-increasing pace of new technical advances, the future of eDNA looks bright, yet successful applications and best practices must become more interdisciplinary to reach its full potential. Our retrospect gives the tools and expectations towards concretely moving the field forward.

Evolutionary change in flight-to-light response in urban moths comes with changes in wing morphology.

Moths and other insects are attracted by artificial light sources. This flight-to-light behaviour disrupts their general activity focused on finding resources, such as mating partners, and increases predation risk. It thus has substantial fitness costs. In illuminated urban areas, spindle ermine moths Yponomeuta cagnagella were reported to have evolved a reduced flight-to-light response. Yet, the specific mechanism remained unknown, and was hypothesized to involve either changes in visual perception or general flight ability or overall mobility traits. Here, we test whether spindle ermine moths from urban and rural populations-with known differences in flight-to-light responses-differ in flight-related morphological traits. Urban individuals were found to have on average smaller wings than rural moths, which in turn correlated with a lower probability of being attracted to an artificial light source. Our finding supports the reduced mobility hypothesis, which states that reduced mobility in urban areas is associated with specific morphological changes in the flight apparatus.

Invasive species drive cross-ecosystem effects worldwide.

Invasive species are pervasive around the world and have profound impacts on the ecosystem they invade. Invasive species, however, can also have impacts beyond the ecosystem they invade by altering the flow of non-living materials (for example, nutrients or chemicals) or movement of organisms across the boundaries of the invaded ecosystem. Cross-ecosystem interactions via spatial flows are ubiquitous in nature, for example, connecting forests and lakes, grasslands and rivers, and coral reefs and the deep ocean. Yet, we have a limited understanding of the cross-ecosystem impacts invasive species have relative to their local effects. By synthesizing emerging evidence, here we demonstrate the cross-ecosystem impacts of invasive species as a ubiquitous phenomenon that influences biodiversity and ecosystem functioning around the world. We identify three primary ways by which invasive species have cross-ecosystem effects: first, by altering the magnitude of spatial flows across ecosystem boundaries; second, by altering the quality of spatial flows; and third, by introducing novel spatial flows. Ultimately, the strong impacts invasive species can drive across ecosystem boundaries suggests the need for a paradigm shift in how we study and manage invasive species around the world, expanding from a local to a cross-ecosystem perspective.

SWECO25: a cross-thematic raster database for ecological research in Switzerland.

Standard and easily accessible cross-thematic spatial databases are key resources in ecological research. In Switzerland, as in many other countries, available data are scattered across computer servers of research institutions and are rarely provided in standard formats (e.g., different extents or projections systems, inconsistent naming conventions). Consequently, their joint use can require heavy data management and geomatic operations. Here, we introduce SWECO25, a Swiss-wide raster database at 25-meter resolution gathering 5,265 layers. The 10 environmental categories included in SWECO25 are: geologic, topographic, bioclimatic, hydrologic, edaphic, land use and cover, population, transportation, vegetation, and remote sensing. SWECO25 layers were standardized to a common grid sharing the same resolution, extent, and geographic coordinate system. SWECO25 includes the standardized source data and newly calculated layers, such as those obtained by computing focal or distance statistics. SWECO25 layers were validated by a data integrity check, and we verified that the standardization procedure had a negligible effect on the output values. SWECO25 is available on Zenodo and is intended to be updated and extended regularly.

Groundwater is a hidden global keystone ecosystem.

Groundwater is a vital ecosystem of the global water cycle, hosting unique biodiversity and providing essential services to societies. Despite being the largest unfrozen freshwater resource, in a period of depletion by extraction and pollution, groundwater environments have been repeatedly overlooked in global biodiversity conservation agendas. Disregarding the importance of groundwater as an ecosystem ignores its critical role in preserving surface biomes. To foster timely global conservation of groundwater, we propose elevating the concept of keystone species into the realm of ecosystems, claiming groundwater as a keystone ecosystem that influences the integrity of many dependent ecosystems. Our global analysis shows that over half of land surface areas (52.6%) has a medium-to-high interaction with groundwater, reaching up to 74.9% when deserts and high mountains are excluded. We postulate that the intrinsic transboundary features of groundwater are critical for shifting perspectives towards more holistic approaches in aquatic ecology and beyond. Furthermore, we propose eight key themes to develop a science-policy integrated groundwater conservation agenda. Given ecosystems above and below the ground intersect at many levels, considering groundwater as an essential component of planetary health is pivotal to reduce biodiversity loss and buffer against climate change.

Multi-decadal improvements in the ecological quality of European rivers are not consistently reflected in biodiversity metrics.

Humans impact terrestrial, marine and freshwater ecosystems, yet many broad-scale studies have found no systematic, negative biodiversity changes (for example, decreasing abundance or taxon richness). Here we show that mixed biodiversity responses may arise because community metrics show variable responses to anthropogenic impacts across broad spatial scales. We first quantified temporal trends in anthropogenic impacts for 1,365 riverine invertebrate communities from 23 European countries, based on similarity to least-impacted reference communities. Reference comparisons provide necessary, but often missing, baselines for evaluating whether communities are negatively impacted or have improved (less or more similar, respectively). We then determined whether changing impacts were consistently reflected in metrics of community abundance, taxon richness, evenness and composition. Invertebrate communities improved, that is, became more similar to reference conditions, from 1992 until the 2010s, after which improvements plateaued. Improvements were generally reflected by higher taxon richness, providing evidence that certain community metrics can broadly indicate anthropogenic impacts. However, richness responses were highly variable among sites, and we found no consistent responses in community abundance, evenness or composition. These findings suggest that, without sufficient data and careful metric selection, many common community metrics cannot reliably reflect anthropogenic impacts, helping explain the prevalence of mixed biodiversity trends.

Freshwater fish diversity in the western Amazon basin shaped by Andean uplift since the Late Cretaceous.

South America is home to the highest freshwater fish biodiversity on Earth, and the hotspot of species richness is located in the western Amazon basin. The location of this hotspot is enigmatic, as it is inconsistent with the pattern observed in river systems across the world of increasing species richness towards a river's mouth. Here we investigate the role of river capture events caused by Andean mountain building and repeated episodes of flooding in western Amazonia in shaping the modern-day richness pattern of freshwater fishes in South America, and in Amazonia in particular. To this end, we combine a reconstruction of river networks since 80 Ma with a mechanistic model simulating dispersal, allopatric speciation and extinction over the dynamic landscape of rivers and lakes. We show that Andean mountain building and consequent numerous small river capture events in western Amazonia caused freshwater habitats to be highly dynamic, leading to high diversification rates and exceptional richness. The history of marine incursions and lakes, including the Miocene Pebas mega-wetland system in western Amazonia, played a secondary role.

Integrating citizen science and environmental DNA metabarcoding to study biodiversity of groundwater amphipods in Switzerland.

Groundwater is the physically largest freshwater ecosystem, yet one of the least explored habitats on earth, both because of accessing difficulties and the scarcity of the organisms inhabiting it. Here, we demonstrate how a two-fold approach provides complementary information on the occurrence and diversity of groundwater amphipods. Firstly, we used a citizen science approach in collaboration with municipal water providers who sampled groundwater organisms in their spring catchment boxes over multiple weeks, followed by DNA barcoding. Secondly, we collected four 10 L water samples at each site, in one sampling event, for environmental DNA (eDNA) metabarcoding. We found that citizen science was very effective in describing the distribution and abundance of groundwater amphipods. Although the single time-point of eDNA sampling did not detect as many amphipods, it allowed the assessment of the entire groundwater community, including microorganisms. By combining both methods, we found different amphipod species co-occurring with distinct sequences from the eDNA-metabarcoding dataset, representing mainly micro-eukaryotic species. We also found a distinct correlation between the diversity of amphipods and the overall biodiversity of groundwater organisms detected by eDNA at each site. We thus suggest that these approaches can be used to get a better understanding of subterranean biodiversity.

Fishing eDNA in One of the World's Largest Rivers: A Case Study of Cross-Sectional and Depth Profile Sampling in the Yangtze.

The world's largest rivers are home to diverse, endemic, and threatened fish species. However, their sheer sizes make large-scale biomonitoring challenging. While environmental DNA (eDNA) metabarcoding has become an established monitoring approach in smaller freshwater ecosystems, its suitability for large rivers may be challenged by the sheer extent of their cross sections (>1 km wide and tens of meters deep). Here, we sampled fish eDNA from multiple vertical layers and horizontal locations from two cross sections of the lower reach of the Yangtze River in China. Over half of the ASVs (amplicon sequence variants) were detected in only a single combination of the vertical layers and horizontal locations, with ∼7% across all combinations. We estimated the need to sample >100 L of water across the cross-sectional profiles to achieve ASV richness saturation, which translates to ∼60 L of water at the species level. No consistent pattern emerged for prioritizing certain depth and horizontal samples, yet we underline the importance of sampling and integrating different layers and locations simultaneously. Our study highlights the significance of spatially stratified sampling and sampling volumes when using eDNA approaches. Specifically, we developed and tested a scalable and broadly applicable strategy that advances the monitoring and conservation of large rivers.

Catchment-based sampling of river eDNA integrates terrestrial and aquatic biodiversity of alpine landscapes.

Monitoring of terrestrial and aquatic species assemblages at large spatial scales based on environmental DNA (eDNA) has the potential to enable evidence-based environmental policymaking. The spatial coverage of eDNA-based studies varies substantially, and the ability of eDNA metabarcoding to capture regional biodiversity remains to be assessed; thus, questions about best practices in the sampling design of entire landscapes remain open. We tested the extent to which eDNA sampling can capture the diversity of a region with highly heterogeneous habitat patches across a wide elevation gradient for five days through multiple hydrological catchments of the Swiss Alps. Using peristaltic pumps, we filtered 60 L of water at five sites per catchment for a total volume of 1800 L. Using an eDNA metabarcoding approach focusing on vertebrates and plants, we detected 86 vertebrate taxa spanning 41 families and 263 plant taxa spanning 79 families across ten catchments. For mammals, fishes, amphibians and plants, the detected taxa covered some of the most common species in the region according to long-term records while including a few more rare taxa. We found marked turnover among samples from distinct elevational classes indicating that the biological signal in alpine rivers remains relatively localised and is not aggregated downstream. Accordingly, species compositions differed between catchments and correlated with catchment-level forest and grassland cover. Biomonitoring schemes based on capturing eDNA across rivers within biologically integrated catchments may pave the way toward a spatially comprehensive estimation of biodiversity.

The recovery of European freshwater biodiversity has come to a halt.

Owing to a long history of anthropogenic pressures, freshwater ecosystems are among the most vulnerable to biodiversity loss1. Mitigation measures, including wastewater treatment and hydromorphological restoration, have aimed to improve environmental quality and foster the recovery of freshwater biodiversity2. Here, using 1,816 time series of freshwater invertebrate communities collected across 22 European countries between 1968 and 2020, we quantified temporal trends in taxonomic and functional diversity and their responses to environmental pressures and gradients. We observed overall increases in taxon richness (0.73% per year), functional richness (2.4% per year) and abundance (1.17% per year). However, these increases primarily occurred before the 2010s, and have since plateaued. Freshwater communities downstream of dams, urban areas and cropland were less likely to experience recovery. Communities at sites with faster rates of warming had fewer gains in taxon richness, functional richness and abundance. Although biodiversity gains in the 1990s and 2000s probably reflect the effectiveness of water-quality improvements and restoration projects, the decelerating trajectory in the 2010s suggests that the current measures offer diminishing returns. Given new and persistent pressures on freshwater ecosystems, including emerging pollutants, climate change and the spread of invasive species, we call for additional mitigation to revive the recovery of freshwater biodiversity.

Leaf-associated macroinvertebrate assemblage and leaf litter breakdown in headwater streams depend on local riparian vegetation.

Headwater streams harbor diverse macroinvertebrate communities and are hotspots for leaf litter breakdown. The process of leaf litter breakdown mediated by macroinvertebrates forms an important link between terrestrial and aquatic ecosystems. Yet, how the vegetation type in the local riparian zone influences leaf-associated macroinvertebrate assemblages and leaf litter breakdown rates is still not resolved. We investigated how leaf-associated macroinvertebrate assemblages and leaf litter fragmentation rates differ between forested and non-forested sites using experimental leaf litter bags in sixteen sites paired across eight headwater streams in Switzerland. Our results show that sensitive taxa of the invertebrate orders Ephemeroptera, Plecoptera and Trichoptera (EPT) and the functional group of shredders were strongly associated with forested sites with overall higher values of abundance, diversity, and biomass of EPTs in forested compared to non-forested sites. However, the importance of riparian vegetation differed between study regions, especially for shredders. Fragmentation rates, which are primarily the result of macroinvertebrate shredding, were on average three times higher in forested compared to non-forested sites. Our results demonstrate that not only the composition of the aquatic fauna but also the functioning of an essential ecosystem process depend on the vegetation type in the local riparian zone. Supplementary Information: The online version contains supplementary material available at 10.1007/s10750-022-05049-7.

A combination of machine-learning and eDNA reveals the genetic signature of environmental change at the landscape levels.

The current advances of environmental DNA (eDNA) bring profound changes to ecological monitoring and provide unique insights on the biological diversity of ecosystems. The very nature of eDNA data is challenging yet also revolutionizing how biological monitoring information is analysed. In particular, new metrics and approaches should take full advantage of the extent and detail of molecular data produced by genetic methods. In this perspective, machine learning algorithms are particularly promising as they can capture complex relationships between the multiple environmental pressures and the diversity of biological communities. We investigated the potential of a new generation of biomonitoring tools that implement machine-learning techniques to fully exploit eDNA datasets. We trained a machine learning model to discriminate between reference and impacted communities of freshwater macroinvertebrates and assessed its performances using a large eDNA dataset collected at 64 standard federal monitoring sites across Switzerland. We show that a model trained on eDNA is significantly better than a naive model and performs similarly to a model trained on traditional data. Our proof-of-concept shows that such a combination of eDNA and machine learning approaches has the potential to complement or even replace traditional environmental monitoring, and could be scaled along temporal or spatial dimensions.

Destabilizing Effects of Environmental Stressors on Aquatic Communities and Interaction Networks across a Major River Basin.

Human-driven environmental stressors are increasingly threatening species survival and diversity of river systems worldwide. However, it remains unclear how the stressors affect the stability changes across aquatic multiple communities. Here, we used environmental DNA (eDNA) data sets from a human-dominated river in China over 3 years and analyzed the stability changes in multiple communities under persistent anthropogenic stressors, including land use and pollutants. First, we found that persistent stressors significantly reduced multifaceted species diversity (e.g., species richness, Shannon's diversity, and Simpson's diversity) and species stability but increased species synchrony across multiple communities. Second, the structures of interaction networks inferred from an empirical meta-food web were significantly changed under persistent stressors, for example, resulting in decreased network modularity and negative/positive cohesion. Third, piecewise structural equation modeling proved that the persistent stress-induced decline in the stability of multiple communities mainly depended upon diversity-mediated pathways rather than the direct effects of stress per se; specifically, the increase of species synchrony and the decline of interaction network modularity were the main biotic drivers of stability variation. Overall, our study highlights the destabilizing effects of persistent stressors on multiple communities as well as the mechanistic dependencies, mainly through reducing species diversity, increasing species synchrony, and changing interaction networks.

Modelling environmental DNA transport in rivers reveals highly resolved spatio-temporal biodiversity patterns.

The ever-increasing threats to riverine ecosystems call for novel approaches for highly resolved biodiversity assessments across taxonomic groups and spatio-temporal scales. Recent advances in the joint use of environmental DNA (eDNA) data and eDNA transport models in rivers (e.g., eDITH) allow uncovering the full structure of riverine biodiversity, hence elucidating ecosystem processes and supporting conservation measures. We applied eDITH to a metabarcoding dataset covering three taxonomic groups (fish, invertebrates, bacteria) and three seasons for a catchment sampled for eDNA at 73 sites. We upscaled eDNA-based biodiversity predictions to approximately 1900 reaches, and assessed α- and ß-diversity patterns across seasons and taxonomic groups over the whole network. Genus richness predicted by eDITH was generally higher than values from direct eDNA analysis. Both predicted α- and ß-diversity varied depending on season and taxonomic group. Predicted fish α-diversity increased downstream in all seasons, while invertebrate and bacteria α-diversity either decreased downstream or were unrelated to network position. Spatial ß-diversity mostly decreased downstream, especially for bacteria. The eDITH model yielded a more refined assessment of freshwater biodiversity as compared to raw eDNA data, both in terms of spatial coverage, diversity patterns and effect of covariates, thus providing a more complete picture of freshwater biodiversity.

Groundwater environmental DNA metabarcoding reveals hidden diversity and reflects land-use and geology.

Despite being the most important source of liquid freshwater on the planet, groundwater is severely threatened by climate change, agriculture, or industrial mining. It is thus extensively monitored for pollutants and declines in quantity. The organisms living in groundwater, however, are rarely the target of surveillance programmes and little is known about the fauna inhabiting underground habitats. The difficulties accessing groundwater, the lack of expertise, and the apparent scarcity of these organisms challenge sampling and prohibit adequate knowledge on groundwater fauna. Environmental DNA (eDNA) metabarcoding provides an approach to overcome these limitations but is largely unexplored. Here, we sampled water in 20 communal spring catchment boxes used for drinking water provisioning in Switzerland, with a high level of replication at both filtration and amplification steps. We sequenced a portion of the COI mitochondrial gene, which resulted in 4917 ASVs, yet only 3% of the reads could be assigned to a species, genus, or family with more than 90% identity. Careful evaluation of the unassigned reads corroborated that these sequences were true COI sequences belonging mostly to diverse eukaryotic groups, not present in the reference databases. Principal component analyses showed a strong correlation of the community composition with the surface land-use (agriculture vs. forest) and geology (fissured rock vs. unconsolidated sediment). While incomplete reference databases limit the assignment of taxa in groundwater eDNA metabarcoding, we showed that taxonomy-free approaches can reveal large hidden diversity and couple it with major land-use drivers, revealing their imprint on chemical and biological properties of groundwater.

Experimental evolution of dispersal: Unifying theory, experiments and natural systems.

Dispersal is a central life history trait that affects the ecological and evolutionary dynamics of populations and communities. The recent use of experimental evolution for the study of dispersal is a promising avenue for demonstrating valuable proofs of concept, bringing insight into alternative dispersal strategies and trade-offs, and testing the repeatability of evolutionary outcomes. Practical constraints restrict experimental evolution studies of dispersal to a set of typically small, short-lived organisms reared in artificial laboratory conditions. Here, we argue that despite these restrictions, inferences from these studies can reinforce links between theoretical predictions and empirical observations and advance our understanding of the eco-evolutionary consequences of dispersal. We illustrate how applying an integrative framework of theory, experimental evolution and natural systems can improve our understanding of dispersal evolution under more complex and realistic biological scenarios, such as the role of biotic interactions and complex dispersal syndromes.