Temperature, productivity, and habitat characteristics collectively drive lake food web structure.

While many efforts have been devoted to understand variations in food web structure among terrestrial and aquatic ecosystems, the environmental factors influencing food web structure at large spatial scales remain hardly explored. Here, we compiled biodiversity inventories to infer food web structure of 67 French lakes using an allometric niche-based model and tested how environmental variables (temperature, productivity, and habitat) influence them. By applying a multivariate analysis on 20 metrics of food web topology, we found that food web structural variations are represented by two distinct complementary and independent structural descriptors. The first is related to the overall trophic diversity, whereas the second is related to the vertical structure. Interestingly, the trophic diversity descriptor was mostly explained by habitat size (26.7% of total deviance explained) and habitat complexity (20.1%) followed by productivity (dissolved organic carbon: 16.4%; nitrate: 9.1%) and thermal variations (10.7%). Regarding the vertical structure descriptor, it was mostly explained by water thermal seasonality (39.0% of total deviance explained) and habitat depth (31.9%) followed by habitat complexity (8.5%) and size (5.5%) as well as annual mean temperature (5.6%). Overall, we found that temperature, productivity, and habitat characteristics collectively shape lake food web structure. We also found that intermediate levels of productivity, high levels of temperature (mean and seasonality), as well as large habitats are associated with the largest and most complex food webs. Our findings, therefore, highlight the importance of focusing on these three components especially in the context of global change, as significant structural changes in aquatic food webs could be expected under increased temperature, pollution, and habitat alterations.

A 34-year survey under phosphorus decline and warming: Consequences on stoichiometry and functional trait composition of freshwater macroinvertebrate communities.

Worldwide, freshwater systems are subjected to increasing temperatures and nutrient changes. Under phosphorus and nitrogen enrichment consumer communities are often thought to shift towards fast-growing and P-rich taxa, supporting the well-known link between growth rate and body stoichiometry. While these traits are also favoured under warming, the temperature effect on stoichiometry is less clear. As recently shown, there is a general link between functional traits and body stoichiometry, which makes the integration of stoichiometric traits a promising tool to help understanding the mechanisms behind taxonomic and functional community responses to nutrient changes and/or warming. Yet, such approaches have been scarcely developed at community level and on a long-term perspective. In this study, we investigated long-term responses in stoichiometry and functional trait composition of macroinvertebrate communities to nutrient changes (decreasing water P; increasing water N:P) and warming over a 34-year period in the Middle Loire River (France), testing the potentially opposing responses to these drivers. Both drivers should cause shifts in species composition, which will alter the overall community stoichiometry and functional composition following assumptions from ecological stoichiometry theory. We found that the macroinvertebrate community shifted towards P-poor taxa, causing significant trends in overall community stoichiometry which indicates long-term changes in the nutrient pool provided by these consumers (i.e. decrease in %N and %P, increase in N:P). Further, while the former high-P conditions favoured traits associated to detritus feeding and fast development (i.e. small maximum body size, short life duration), recent conditions favoured predators and slow-developing taxa. These results suggest nutrients to be a more important driver than temperature over this period. By providing a pivotal link between environmental changes and functional trait composition of communities, approaches based on stoichiometric traits offer sound perspectives to investigate ecological relationships between multiple drivers operating at various scales and ecosystem functioning.

Lessons from linking bio- and ecological traits to stoichiometric traits in stream macroinvertebrates.

Ecologists rely on various functional traits when investigating the functioning of ecological systems and its responses to global changes. Changing nutrient levels, for example, can affect taxa expressing different trait combinations in various ways, e.g., favoring small, fast-growing species under high phosphorus conditions. Stoichiometric traits, describing the elemental composition of organism body tissues, can help in understanding the mechanisms behind such functional shifts. So far, mainly life-history traits have been related to body stoichiometry (e.g., the growth rate hypothesis) on a limited number of taxa, and there is little knowledge of the general link between stoichiometric and other functional traits on a taxonomically large scale. Here, we highlight this link in the freshwater macroinvertebrates, testing predictions from underlying trait-based and Ecological Stoichiometry Theory (EST) in >200 taxa belonging to eight larger taxonomic groups. We applied a series of multivariate analyses on six of their stoichiometric traits (%C, %N, %P, C:N, C:P, and N:P) and 23 biological and ecological traits. We found significant relationships between stoichiometric traits and other types of traits when analyzing single-trait and multi-trait profiles. Patterns found within traits related to organism development or nutrient cycling were in line with our assumptions based on EST, e.g., traits describing predators were associated with high %N; traits suggesting a fast development (small maximum body size and high molting frequency) with high %P. Associations between ecological traits and body stoichiometry could be explained by the longitudinal stream gradient: Taxa preferring headwater habitats (i.e., high altitude, coarse substrate, and cold temperature) exhibited high %N and %P. Demonstrating the link between stoichiometric and both bio- and ecological traits on a large diversity of taxa underlines the potential of integrating stoichiometric traits into ecological analyses to improve our understanding of taxonomic and functional responses of communities-and ecosystems-to changing environmental conditions worldwide.

Evaluating the biological validity of European river typology systems with least disturbed benthic macroinvertebrate communities.

Humans have severely altered freshwater ecosystems globally, causing a loss of biodiversity. Regulatory frameworks, like the Water Framework Directive, have been developed to support actions that halt and reverse this loss. These frameworks use typology systems that summarize freshwater ecosystems into environmentally delineated types. Within types, ecosystems that are minimally impacted by human activities, i.e., in reference conditions, are expected to be similar concerning physical, chemical, and biological characteristics. This assumption is critical when water quality assessments rely on comparisons to type-specific reference conditions. Lyche Solheim et al. (2019) developed a pan-European river typology system, the Broad River Types, that unifies the national Water Framework Directive typology systems and is gaining traction within the research community. However, it is unknown how similar biological communities are within these individual Broad River Types. We used analysis of similarities and classification strength analysis to examine if the Broad River Types delineate distinct macroinvertebrate communities across Europe and whether they outperform two ecoregional approaches: the European Biogeographical Regions and Illies' Freshwater Ecoregions. We determined indicator and typical taxa for the types of all three typology systems and evaluated their distinctiveness. All three typology systems captured more variation in macroinvertebrate communities than random combinations of sites. The results were similar among typology systems, but the Broad River Types always performed worse than either the Biogeographic Regions or Illies' Freshwater Ecoregions. Despite reaching statistical significance, the statistics of analysis of similarity and classification strength were low in all tests indicating substantial overlap among the macroinvertebrate communities of different types. We conclude that the Broad River Types do not represent an improvement upon existing freshwater typologies when used to delineate macroinvertebrate communities and we propose future avenues for advancement: regionally constrained types, better recognition of intermittent rivers, and consideration of biotic communities.

The combination of chemical, structural, and functional indicators to evaluate the anthropogenic impacts on agricultural stream ecosystems.

Freshwater contamination by pesticides in agricultural landscapes is of increasing concern worldwide, with strong pesticide impacts on biodiversity, ecosystem functions, and ultimately human health (drinking water, fishing). In addition, the excessively large number of substances, as well as their low - and temporally variable - concentrations in water, make the chemical monitoring by grab sampling very demanding and not fully representative of the actual contamination. Tools that integrate temporal variations and that are ecologically relevant are clearly needed to improve the monitoring of freshwater contamination and assess its biological effects. Here, we studied pesticide contamination and its biological impacts in 10 stream sections (sites) belonging to 3 agricultural catchments in France. In each site, we deployed a combination of pesticide integrative samplers, biocenotic indicators based on benthic macroinvertebrates, and functional indicators based on leaf litter decomposition and associated fungal communities. The 3 approaches largely proved complementary: structural and functional indicators did not respond equally to different agricultural impacts such as pesticide contamination (as revealed by integrative samplers), nutrients, or oxygen depletion. Combining chemical, structural, and functional indicators thus seems an excellent strategy to provide a comprehensive picture of agricultural impacts on stream ecosystems.

A global perspective on the functional responses of stream communities to flow intermittence.

The current erosion of biodiversity is a major concern that threatens the ecological integrity of ecosystems and the ecosystem services they provide. Due to global change, an increasing proportion of river networks are drying and changes from perennial to non-perennial flow regimes represent dramatic ecological shifts with potentially irreversible alterations of community and ecosystem dynamics. However, there is minimal understanding of how biological communities respond functionally to drying. Here, we highlight the taxonomic and functional responses of aquatic macroinvertebrate communities to flow intermittence across river networks from three continents, to test predictions from underlying trait-based conceptual theory. We found a significant breakpoint in the relationship between taxonomic and functional richness, indicating higher functional redundancy at sites with flow intermittence higher than 28%. Multiple strands of evidence, including patterns of alpha and beta diversity and functional group membership, indicated that functional redundancy did not compensate for biodiversity loss associated with increasing intermittence, contrary to received wisdom. A specific set of functional trait modalities, including small body size, short life span and high fecundity, were selected with increasing flow intermittence. These results demonstrate the functional responses of river communities to drying and suggest that on-going biodiversity reduction due to global change in drying river networks is threatening their functional integrity. These results indicate that such patterns might be common in these ecosystems, even where drying is considered a predictable disturbance. This highlights the need for the conservation of natural drying regimes of intermittent rivers to secure their ecological integrity.

How diatom-, invertebrate- and fish-based diagnostic tools can support the ecological assessment of rivers in a multi-pressure context: Temporal trends over the past two decades in France.

The degradation of aquatic ecosystems, induced by worldwide intensification in the use of both land and aquatic resources, has highlighted the critical need for innovative methods allowing an objective quantification and ranking of anthropogenic pressure effects on aquatic organisms. Such diagnostic tools have a great potential for defining robust management responses to anthropogenic pressures. Our objective was to explore how the outputs of three diagnostic tools (based on benthic diatoms, macroinvertebrates and fishes) could be combined to (i) disentangle the temporal effects of multiple pressures over two decades and (ii) provide policy-relevant information for stream managers and decision makers. The diagnostic tools estimated, using taxonomy- and trait-based metrics, the impairment probabilities of biotic assemblages over time by different pressure categories, describing the alteration of water quality, hydromorphology and land use related to anthropogenic activities, in French streams (number of sites = 312). The main result shows that a large proportion of the time series exhibited no significant temporal patterns over the two decades (61.5% to 87.8%, depending on the used tests). Among time series exhibiting significant change, positive trends in impairment probabilities (i.e., degradation) were less frequent than negative ones, indicating a modest improvement in water quality at national scale over the study period. However, trends can be substantially different according to hydroecoregion and pressure category. The three biological compartments displayed convergent temporal responses according to the pressure category and regional context (e.g., lowland plains vs. mountains, pristine vs. agricultural regions). Altogether, this study proposes a unifying approach to integrate a vast amount of information in a single ecological diagnosis using an unparalleled database on natural and anthropized environments. Strengthening the synthesis of biological information provided by various biological compartments should be a priority before implementing evidence-based sustainable conservation and restoration actions.

Morphological vs. DNA metabarcoding approaches for the evaluation of stream ecological status with benthic invertebrates: Testing different combinations of markers and strategies of data filtering.

Macroinvertebrate assemblages are the most common bioindicators used for stream biomonitoring, yet the standard approach exhibits several time-consuming steps, including the sorting and identification of organisms based on morphological criteria. In this study, we examined if DNA metabarcoding could be used as an efficient molecular-based alternative to the morphology-based monitoring of streams using macroinvertebrates. We compared results achieved with the standard morphological identification of organisms sampled in 18 sites located on 15 French wadeable streams to results obtained with the DNA metabarcoding identification of sorted bulk material of the same macroinvertebrate samples, using read numbers (expressed as relative frequencies) as a proxy for abundances. In particular, we evaluated how combining and filtering metabarcoding data obtained from three different markers (COI: BF1-BR2, 18S: Euka02 and 16S: Inse01) could improve the efficiency of bioassessment. In total, 140 taxa were identified based on morphological criteria, and 127 were identified based on DNA metabarcoding using the three markers, with an overlap of 99 taxa. The threshold values used for sequence filtering based on the "best identity" criterion and the number of reads had an effect on the assessment efficiency of data obtained with each marker. Compared to single marker results, combining data from different markers allowed us to improve the match between biotic index values obtained with the bulk DNA versus morphology-based approaches. Both approaches assigned the same ecological quality class to a majority (86%) of the site sampling events, highlighting both the efficiency of metabarcoding as a biomonitoring tool but also the need for further research to improve this efficiency.

Comparison of markers for the monitoring of freshwater benthic biodiversity through DNA metabarcoding.

Metabarcoding of bulk or environmental DNA has great potential for biomonitoring of freshwater environments. However, successful application of metabarcoding to biodiversity monitoring requires universal primers with high taxonomic coverage that amplify highly variable, short metabarcodes with high taxonomic resolution. Moreover, reliable and extensive reference databases are essential to match the outcome of metabarcoding analyses with available taxonomy and biomonitoring indices. Benthic invertebrates, particularly insects, are key taxa for freshwater bioassessment. Nevertheless, few studies have so far assessed markers for metabarcoding of freshwater macrobenthos. Here we combined in silico and laboratory analyses to test the performance of different markers amplifying regions in the 18S rDNA (Euka02), 16S rDNA (Inse01) and COI (BF1_BR2-COI) genes, and developed an extensive database of benthic macroinvertebrates of France and Europe, with a particular focus on key insect orders (Ephemeroptera, Plecoptera and Trichoptera). Analyses on 1,514 individuals representing different taxa of benthic macroinvertebrates showed very different amplification success across primer combinations. The Euka02 marker showed the highest universality, while the Inse01 marker showed excellent performance for the amplification of insects. BF1_BR2-COI showed the highest resolution, while the resolution of Euka02 was often limited. By combining our data with GenBank information, we developed a curated database including sequences representing 822 genera. The heterogeneous performance of the different primers highlights the complexity in identifying the best markers, and advocates for the integration of multiple metabarcodes for a more comprehensive and accurate understanding of ecological impacts on freshwater biodiversity.

DISPERSE, a trait database to assess the dispersal potential of European aquatic macroinvertebrates.

Dispersal is an essential process in population and community dynamics, but is difficult to measure in the field. In freshwater ecosystems, information on biological traits related to organisms' morphology, life history and behaviour provides useful dispersal proxies, but information remains scattered or unpublished for many taxa. We compiled information on multiple dispersal-related biological traits of European aquatic macroinvertebrates in a unique resource, the DISPERSE database. DISPERSE includes nine dispersal-related traits subdivided into 39 trait categories for 480 taxa, including Annelida, Mollusca, Platyhelminthes, and Arthropoda such as Crustacea and Insecta, generally at the genus level. Information within DISPERSE can be used to address fundamental research questions in metapopulation ecology, metacommunity ecology, macroecology and evolutionary ecology. Information on dispersal proxies can be applied to improve predictions of ecological responses to global change, and to inform improvements to biomonitoring, conservation and management strategies. The diverse sources used in DISPERSE complement existing trait databases by providing new information on dispersal traits, most of which would not otherwise be accessible to the scientific community.

Environmental filtering and limiting similarity as main forces driving diatom community structure in Mediterranean and continental temporary and perennial streams.

Climatic extreme events such as droughts (unpredictable), dry periods (predictable) or even flush floods, threaten freshwater ecosystems worldwide. The filtering mechanisms of these events and their strength on communities, however, can be different among regions. While time-for-adaptation theory defines whether or not water scarcity can be considered as disturbance, the stress-dominance theory predicts an increase in importance of environmental filtering and a decrease in the role of biotic interactions in communities with increasing environmental stress. Here, we tested whether environmental filtering (leading to trait convergence) or limiting similarity (leading to trait divergence) is the main assembly rule shaping the structure and trait composition of benthic diatom assemblages in Mediterranean (Portuguese) and continental (Hungarian) temporary and perennial streams. We assumed that the trait composition of diatom assemblages in the two stream types would be less different in the Mediterranean than in the continental region (addressed to time-for-adaptation theory). We also hypothesized that trait composition would be shaped by environmental filtering in the Hungarian streams while by biotic interactions in Portuguese streams (addressed to stress-dominance theory). Our results supported our first hypothesis since traits, which associated primarily to temporary streams were found only in the continental region. Our findings, however, only partially proved the stress-dominance hypothesis. In the continental region, where drying up of streams were induced by unpredictable droughts, biotic interactions were the main assembly rules shaping community structure. In contrast, environmental filtering was nearly as important as limiting similarity in structuring trait composition in the Mediterranean region during the predictable dry phase with no superficial flow. These analyses also highlighted that drought events (both predictable and unpredictable ones) have a complex and strong influence on benthic diatom assemblages resulting even in irreversible changes in trait composition and thereby in ecosystem functioning.

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.

Modeling diadromous fish loss from historical data: Identification of anthropogenic drivers and testing of mitigation scenarios.

Diadromous fishes have drastically declined over the last century, especially in Europe. Several authors have highlighted the role of large dams in this decline, but in fact, its causes are potentially multiple and cumulative, including degradation of local environmental conditions and widespread fragmentation of hydrographic networks associated with the pervasive establishment of smaller barriers. Consequently, there is a need to improve the identification and prioritization of the drivers of diadromous species loss in order to identify and apply the most appropriate conservation and restoration measures. In this study, we used both historical sources (from mid-18th to early 20th century) and current data to quantify the long-term loss of diadromous taxa over 555 sites throughout the French river network. Then, we modeled the effects of several anthropogenic pressures (e.g. barriers, water quality, hydrological and river morphological alterations) on diadromous taxon loss. Lastly, we assessed the potential consequences of four different scenarios of anthropogenic pressure reduction. Due to uncertainties in historical sources, some species were grouped into taxa leading to a potential underestimation of actual species extinctions. Despite this limitation, our results showed that the decline in diadromous assemblages is widespread but with contrasting magnitudes depending on site locations. The maximum height and density of barriers appeared as the major factors of taxon loss. Over the scenarios tested, we observed that exclusively improving local conditions have much more limited effects than restoring river continuity. Focusing actions on large dam removal did not show the strongest responses compared to removing medium and small-sized barriers. For effective and sustainable restoration of diadromous fish assemblage, (1) historical occurrences of diadromous fishes should be used as an indicator for assessing recovery, and (2) undertaken measures must be adapted to each basin to target and limit the number of barriers to remove while allowing diadromous fish recovery.

A diagnosis-based approach to assess specific risks of river degradation in a multiple pressure context: Insights from fish communities.

In the context of increasing pressure on water bodies, many fish-based indices have been developed to evaluate the ecological status of rivers. However, most of these indices suffer from several limitations, which hamper the capacity of water managers to select the most appropriate measures of restoration. Those limitations include: (i) being dependent on reference conditions, (ii) not satisfactorily handling complex and non-linear biological responses to pressure gradients, and (iii) being unable to identify specific risks of stream degradation in a multi-pressure context. To tackle those issues, we developed a diagnosis-based approach using Random Forest models to predict the impairment probabilities of river fish communities by 28 pressure categories (chemical, hydromorphological and biological). In addition, the database includes the abundances of 72 fish species collected from 1527 sites in France, sampled between 2005 and 2015; and fish taxonomic and biological information. Twenty random forest models provided at least good performances when evaluating impairment probabilities of fish communities by those pressures. The best performing models indicated that fish communities were impacted, on average, by 7.34 ±â€¯0.03 abiotic pressure categories (mean ±â€¯SE), and that hydromorphological alterations (5.27 ±â€¯0.02) were more often detected than chemical ones (2.06 ±â€¯0.02). These models showed that alterations in longitudinal continuity, and contaminations by Polycyclic Aromatic Hydrocarbons were respectively the most frequent hydromorphological and chemical pressure categories in French rivers. This approach has also efficiently detected the functional impact of invasive alien species. Identifying and ranking the impacts of multiple anthropogenic pressures that trigger functional shifts in river biological communities is essential for managers to prioritize actions and to implement appropriate restoration programmes. Actually implemented in an R package, this approach has the capacity to detect a variety of impairments, resulting in an efficient assessment of ecological risks across various spatial and temporal scales.

Assessing anthropogenic pressure in the St. Lawrence River using traits of benthic macroinvertebrates.

This study aims to evaluate the anthropogenic pressure in the St. Lawrence River by assessing the relationships between chemical contamination of sediments and benthic community structure with the trait-based approach. Organic and inorganic contaminants as well as other sediment variables (sediment grain size, total organic carbon, nutrients, etc.) and benthic invertebrate assemblages were determined in 59 sites along the river. Biological and ecological traits of taxa were coded, taking into account regional climate and ecosystem conditions. The aims of this study were to (1) describe the relationships between traits and macroinvertebrate taxa and identify homogeneous clusters of taxa with the same combinations of functional traits, (2) describe spatial patterns in traits of macroinvertebrates in the St. Lawrence River, (3) link trait-based metrics and site groups to sediment quality and (4) define a trait-based strategy for diagnosing the ecological quality of the St. Lawrence River. Seven groups of taxa sharing similar trait-category attributes were defined. Moreover, four groups of sites were identified using the 'K-mean' non-hierarchical clustering approach. The 'IndVal' method enabled us to specifically defined trait categories corresponding to site groups on the basis of their indicator value. The relative abundances of taxa from five functional groups significantly varied among site groups. For example, some indicator traits such as multivoltine cycle, long life span, fixed clutches, tegumental respiration, asexual reproduction, and collector/gatherer feeding habit were associated to the most heavily polluted sites located in the Montreal harbour which showed the highest sediment concentrations in Pb, Zn and Cu. Three trait-based pressure-specific models were built, based on the random forest approach, for respectively (1) heavy metals, (2) BPCs and PAHs, and (3) TBTs occurring in the environment. These models could be applied to assess sediment quality using macroinvertebrate assemblages in a large Canadian river.

High stoichiometric food quality increases moulting organism vulnerability to pollutant impacts: An experimental test with Gammarus fossarum (Crustacea: Amphipoda).

Headwater organisms are most often simultaneously faced with multiple stressors such as low resource quality and pollutants. Higher food quality has been hypothesized to enhance the tolerance of organisms to pollutants, but the interactive effects of food quality and pollutants on species and ecosystems remain poorly studied. To better understand these interactive effects, we experimentally manipulated the phosphorus (P) content of two leaf litters with contrasted carbon quality (alder and maple). During four weeks, individuals of the detritivorous crustacean Gammarus fossarum were exposed to low levels of cadmium ([Cd] = 0, 0.35 or 0.70 µg L-1) while being fed with one of the leaf P treatments. When organisms were not exposed to Cd, their high survival rate was more driven by the carbon quality of the resource (litter species) than by its stoichiometric quality. In contrast, their number of moults and growth rates were primarily increased by the P content of resources. When exposed to Cd, G. fossarum survival rate was reduced, but this effect was largely magnified by a higher P level in resources. Our results showed that despite positive effects of resource stoichiometric quality on organism life history traits (growth, survival), a resource of high stoichiometric quality might be detrimental for organisms exposed to low and environmentally realistic levels of pollutants. Two non-exclusive hypotheses are proposed to explain these results. First, organisms fed on the highest quality resource exhibited the highest moulting frequencies (moults being the most critical life cycle step of arthropods), which could have rendered them more sensitive to pollutants. Secondly, the metabolism of organisms fed on higher quality resources was potentially enhanced, increasing the uptake of dissolved Cd by gammarids. This study suggests that species sensitivity to pollutants might be underestimated in ecosystems facing both nutrient constraint and pollutants.

Long-term trends in trait structure of riverine communities facing predation risk increase and trophic resource decline.

Many large European rivers have undergone multiple pressures that have strongly impaired ecosystem functioning at different spatial and temporal scales. Global warming and other environmental changes have favored the success of invasive species, deeply modifying the structure of aquatic communities in large rivers. Some exogenous species could alter trophic interactions within assemblages by increasing the predation risk for potential prey species (top-down effect) and limiting the dynamics of others via resource availability limitation (bottom-up effect). Furthermore, large transboundary rivers are complex aquatic ecosystems that have often been poorly investigated so that data for assessing long-term ecological trends are missing. In this study, we propose an original approach for investigating long-term combined effects of global warming, trophic resource decrease, predation risk, and water quality variations on the trait-based structure of macroinvertebrate and fish assemblages over 26 yr (1985-2011) and 427-km stretch of the river Meuse (France and Belgium). The study of temporal variations in biological, physiological, and ecological traits of macroinvertebrate and fish allowed identifying community trends and distinguishing impacts of environmental perturbations from those induced by biological alterations. We provide evidence, for this large European river, of an increase in water temperature (close to 1°C) and a decrease in phytoplankton biomass (-85%), as well as independent effects of these changes on both invertebrate and fish communities. The reduction of trophic resources in the water column by invasive molluscs has dramatically affected the density of omnivorous fish in favor of invertebrate feeders, while scrapers became the major feeding guild among invertebrates. Macroinvertebrate and fish communities have shifted from large-sized organisms with low fecundity to prolific, small-sized organisms, with early maturity, as a response to increased predation pressure.

Assessing anthropogenic pressures on streams: A random forest approach based on benthic diatom communities.

Benthic diatoms have been widely used to assess the ecological status of freshwater ecosystems, especially in the context of recent international water framework directive policies (e.g. the WFD). Despite diatom-based indices are known to respond fastly to water quality degradation, they are not designed to precisely identify the nature of pressures co-occurring in the environment. Based on large scale monitoring data, we aimed at building models able to estimate the risk of stream impairment by many types of anthropogenic pressures from taxonomy-based and trait-based characteristics of diatom assemblages. Random forest models were built to individually evaluate the impairment risk of diatom assemblages for six chemical and five hydromorphological or land-use related pressure categories. Eight models provided good impairment risk assessment (Area Under the Curve≥0.70). Under multi-pressure scenarios, models built for chemical pressures exhibited a better accuracy than hydromorphological or land-use related ones. Models were able to detect both ecological restoration and degradation, based on long-term surveys. These models have been implemented in a R user-friendly routine, to help stream managers to early identify degrading processes and prioritize management actions.

Assessing long-term effects of multiple, potentially confounded drivers in ecosystems from species traits.

Although species traits have the potential to disentangle long-term effects of multiple, potentially confounded drivers in ecosystems, this issue has received very little attention in the literature. We aimed at filling this gap by assessing the relative effects of hydroclimatic and water quality factors on the trait composition of invertebrate assemblages over 30 years in the Middle Loire River (France). Using a priori predictions on the long-term variation of trait-based adaptations over the three decades, we evaluated the ability of invertebrate traits to indicate the effects of warming, discharge reduction and water quality improvement. Hydroclimatic and water quality factors contributed to up to 65% of the variation in trait composition. More than 70% of the initial trait response predictions made according to observed long-term hydroclimatic changes were confirmed. They supported a general climate-induced trend involving adapted resistance and resilience strategies. A partial confounding effect of water quality improvement acting on trophic processes was also highlighted, indicating that improved water quality management can significantly help to reduce some adverse effects of climate change. This trait-based approach can have wider implications for investigating long-term changes driven by multiple, potentially confounded factors, as frequently encountered in the context of global change.

Investigating the emerging role of comparative proteomics in the search for new biomarkers of metal contamination under varying abiotic conditions.

This study aims at investigating the potential use of comparative proteomics as a multi-marker approach of metal contamination, taking into account the potential confounding effect of water temperature. The major objective was to identify combinations of proteins specifically responding to a given metal, even if included in a metal mixture. The diagnostic approach was performed via the comparative analysis of protein expression on spot mapping provided by adult males of Gammarus pulex (Amphipoda, Crustacea) respectively exposed to arsenate (As), cadmium (Cd) or a binary mixture of these metals (AsCd) at three realistic temperatures (5, 10 and 15°C). Proteomic expression analysis was performed by Differential in-Gel Electrophoresis (2D-DiGE), and completed by an adapted inferential statistical approach. Combinations of under/over-expressed protein spots discriminated the metal identity. However, none of these spots discriminated both the individual metal effect (As or Cd) and its effect in metal mixture (AsCd) whatever the tested temperature. Some limits of the two-dimensional analysis of protein spot maps in G. pulex have been highlighted: (i) the presence of contaminating peptides and/or abundant "déja-vu" proteins which can mask the responses of other proteins of interest or (ii) the presence of post-translational modifications. An optimization of the experimental design (especially during the sample preparation) has been described for future investigations. This study has also highlighted (i) the importance of precisely identifying the protein spots of interest to avoid erroneous interpretations in terms of action mechanisms of chemicals and (ii) the importance of working under controlled laboratory conditions with a temperature close to 10°C. In such conditions, we have demonstrated a higher impact of As than Cd on the energetic metabolism of Gammarus. This As impact is reduced in AsCd mixture confirming the antagonistic interaction of this binary mixture previously observed on G. pulex mortality at 10°C.