Emerging pollutants, such as pharmaceuticals and microplastics have become a pressing concern due to their widespread presence and potential impacts on ecological systems. To assess the ecosystem-level effects of these pollutants within a multi-stressor context, we simulated real-world conditions by exposing a near-natural multi-trophic aquatic food web to a gradient of environmentally relevant concentrations of fluoxetine and microplastics in large mesocosms over a period of more than three months. We measured the biomass and abundance of different trophic groups, as well as ecological functions such as nutrient availability and decomposition rate. To explore the mechanisms underlying potential community and ecosystem-level effects, we also performed behavioral assays focusing on locomotion parameters as a response variable in three species: Daphnia magna (zooplankton prey), Chaoborus flavicans larvae (invertebrate pelagic predator of zooplankton) and Asellus aquaticus (benthic macroinvertebrate), using water from the mesocosms. Our mesocosm results demonstrate that presence of microplastics governs the response in phytoplankton biomass, with a weak non-monotonic dose-response relationship due to the interaction between microplastics and fluoxetine. However, exposure to fluoxetine evoked a strong non-monotonic dose-response in zooplankton abundance and microbial decomposition rate of plant material. In the behavioral assays, the locomotion of zooplankton prey D. magna showed a similar non-monotonic response primarily induced by fluoxetine. Its predator C. flavicans, however, showed a significant non-monotonic response governed by both microplastics and fluoxetine. The behavior of the decomposer A. aquaticus significantly decreased at higher fluoxetine concentrations, potentially leading to reduced decomposition rates near the sediment. Our study demonstrates that effects observed upon short-term exposure result in more pronounced ecosystem-level effects following chronic exposure.

Upland rivers across Europe still exhibit undisturbed conditions and represent a treasure that we cannot afford to lose. We hypothesize that the combination of pristine and modified conditions could demonstrate biological responses along the stressor gradients. Thus, the response of aquatic macrophyte communities to anthropogenic stressors along upland rivers in Bulgaria was studied. Six stressors were selected out of 36 parameters grouped into hydromorphological, chemical variables and combined drivers (catchment land use). The stressors strongly affected species richness on the basis of biological type (bryophytes vs. vascular plants) and ecomorphological type (hydrophytes vs. helophytes). Hydrological alteration expressed by the change of the river\'s base flow and altered riparian habitats has led to a suppression of bryophytes and a dominance of riverbank plant communities. Seventy-five percent of mountain sites were lacking bryophytes, and the vegetation at semi-mountainous sites was dominated by vascular plants. It can be concluded that hydropeaking, organic and inorganic pollution, and discontinuous urban structures caused important modifications in the aquatic macrophyte assemblages. Macrophyte abundance and the biological and ecomorphological type of aquatic macrophytes reflect multi-stressor effects in upland rivers.

Human activities put stress on our oceans and with a growing global population, the impact is increasing. Stressors rarely act in isolation, with the majority of marine areas being impacted by multiple, concurrent stressors. Marine spatial cumulative impact assessments attempt to estimate the collective impact of multiple stressors on marine environments. However, this is difficult given how stressors interact with one another, and the variable response of ecosystems. As a result, assumptions and generalisations are required when attempting to model cumulative impacts. One fundamental assumption of the most commonly applied, semi-quantitative cumulative impact assessment method is that a change in modelled cumulative impact is correlated with a change in ecosystem condition. However, this assumption has rarely been validated with empirical data. We tested this assumption using a case study of seagrass in a large, inverse estuary in South Australia (Spencer Gulf). We compared three different seagrass condition indices, based on survey data collected in the field, to scores from a spatial cumulative impact model for the study area. One condition index showed no relationship with cumulative impact, whilst the other two indices had very small, negative relationships with cumulative impact. These results suggest that one of the most commonly used methods for assessing cumulative impacts on marine systems is not robust enough to accurately reflect the effect of multiple stressors on seagrasses; possibly due to the number and generality of assumptions involved in the approach. Future methods should acknowledge the complex relationships between stressors, and the impact these relationships can have on ecosystems. This outcome highlights the need for greater evaluation of cumulative impact assessment outputs and the need for data-driven approaches. Our results are a caution for marine scientists and resource managers who may rely on spatial cumulative impact assessment outputs for informing policy and decision-making.

Aquatic ecosystems are exposed to multiple stressors such as agricultural run-off (ARO) and climate-change related increase of temperature. We aimed to determine how ARO and the frequency of its input can affect shallow lake ecosystems through direct and indirect effects on primary producers and primary consumers, and whether warming can mitigate or reinforce the impact of ARO. We performed a set of microcosm experiments simulating ARO using a cocktail of three organic pesticides (terbuthylazine, tebuconazole, pirimicarb), copper and nitrate. Two experiments were performed to determine the direct effect of ARO on primary producers (submerged macrophytes, periphyton and phytoplankton) and on the grazing snail Lymnaea stagnalis, respectively. Three different ARO concentrations added as single doses or as multiple pulses at two different temperatures (22°C and 26°C) were applied. In a third experiment, primary producers and consumers were exposed together to allow trophic interactions. When functional groups were exposed alone, ARO had a direct positive effect on phytoplankton and a strong negative effect on L. stagnalis. When exposed together, primary producer responses were contrasting, as the negative effect of ARO on grazers led to an indirect positive effect on periphyton. Periphyton in turn exerted a strong control on phytoplankton, leading to an indirect negative effect of ARO on phytoplankton. Macrophytes showed little response to the stressors. Multiple pulse exposure increased the effect of ARO on L. stagnalis and periphyton when compared with the same quantity of ARO added as a single dose. The increase in temperature had only limited effects. Our results highlight the importance of indirect effects of stressors, here mediated by grazers and periphyton, and the frequency of the ARO input in aquatic ecosystems.

Benthic macroinvertebrate communities are used as indicators for anthropogenic stress in freshwater ecosystems. To better understand the relationship between anthropogenic stress and changes in macroinvertebrate community composition, it is important to understand how different stressors and species traits are associated, and how these associations influence variation in species occurrence and abundances. Here, we show the capacity of the multivariate technique of double constrained correspondence analysis (dc-CA) to analyse trait-environment relationships, and we compare it with the redundancy analysis method on community weighted mean values of traits (CWM-RDA), which is frequently used for this type of analysis. The analyses were based on available biomonitoring data for macroinvertebrate communities from the Danube River. Results from forward selection of traits and environmental variables using dc-CA analyses showed that aquatic stages, reproduction techniques, dispersal tactics, locomotion and substrate relations, altitude, longitudinal and transversal distribution, and substrate preferendum were significantly related to habitat characteristics, hydromorphological alterations and water quality measurements such as physico-chemical parameters, heavy metals, pesticides and pharmaceuticals. Environmental variables significantly associated with traits using the CWM-RDA method were generally consistent with those found in dc-CA analysis. However, the CWM-RDA does neither test nor explicitly select traits, while dc-CA tests and selects both traits and environmental variables. Moreover, the dc-CA analysis revealed that the set of environmental variables was much better in explaining the community data than the available trait set, a kind of information that can neither be obtained from CWM-RDA nor from RLQ (Environment, Link and Trait data), which is a close cousin of dc-CA but not regression-based. Our results suggest that trait-based analysis based on dc-CA may be useful to assess mechanistic links between multiple anthropogenic stressors and ecosystem health, but more data sets should be analysed in the same manner.

Contaminants of emerging concern and ocean changes are key environmental stressors for marine species with possibly synergistic, but still unexplored, deleterious effects. In the present study the influence of a simulated ocean acidification scenario (pH = 7.6) was investigated on metabolism and sub-lethal effects of carbamazepine, CBZ (1 µg/L), chosen as one of the most widely diffused pharmaceuticals in marine organisms. A multidisciplinary approach was applied on mussels, M. galloprovincialis, integrating measurement of drug bioaccumulation with changes in the whole transcriptome, responsiveness of various biochemical and cellular biomarkers including immunological parameters, lipid and oxidative metabolism, onset of genotoxic effects. Chemical analyses revealed a limited influence of hypercapnia on accumulation and excretion of CBZ, while a complex network of biological responses was observed in gene expression profile and functional changes at cellular level. The modulation of gamma-aminobutyric acid (GABA) pathway suggested similarities with the Mechanism of Action known for vertebrates: immune responses, cellular homeostasis and oxidative system represented the processes targeted by combined stressors. The overall elaboration of results through a quantitative Weight of Evidence model, revealed clearly increased cellular hazard due to interactions of CBZ with acidification compared to single stressors.

The Sürgü Stream, located in the Euphrates River basin of Turkey, is used for drinking water source, agricultural irrigation and rainbow trout production. Therefore, water quality of the stream is of great importance. In this study, multivariate statistical techniques (MSTs) and water quality index (WQI) were applied to assess water quality of the stream affected by multiple stressors such as untreated domestic sewage, effluents from fish farms, agricultural runoff and streambank erosion. For this, 16 water quality parameters at five sites along the stream were monitored monthly during one year. Most of parameters showed significant spatial variations, indicating the influence of anthropogenic activities. All parameters except TN (total nitrogen) showed significant seasonal differences due to high seasonality in WT (water temperature) and water flow. The spatial variations in the WQI were significant (p < 0.05) and the mean WQI values ranged from 87.6 to 95.3, indicating \"good\" to \"excellent\" water quality in the stream. Cluster analysis classified five sites into three groups, that is, clean region, low polluted region and very clean region. Stepwise temporal discriminant analysis (DA) identified that pH, WT, Cl-, SO42-, COD (chemical oxygen demand), TSS (total suspended solids) and Ca2+ are the parameters responsible for variations between seasons, and stepwise spatial DA identified that DO (dissolved oxygen), EC (electrical conductivity), NH4-N, TN (total nitrogen) and TSS are the parameters responsible for variations between the regions. Principal component analysis/factor analysis revealed that the parameters responsible for water quality variations were mainly associated with suspended solids (both natural and anthropogenic), soluble salts (natural) and nutrients and organic matter (anthropogenic).

Anthropogenic environmental changes, or \'stressors\', increasingly threaten biodiversity and ecosystem functioning worldwide. Multiple-stressor research is a rapidly expanding field of science that seeks to understand and ultimately predict the interactions between stressors. Reviews and meta-analyses of the primary scientific literature have largely been specific to either freshwater, marine or terrestrial ecology, or ecotoxicology. In this cross-disciplinary study, we review the state of knowledge within and among these disciplines to highlight commonality and division in multiple-stressor research. Our review goes beyond a description of previous research by using quantitative bibliometric analysis to identify the division between disciplines and link previously disconnected research communities. Towards a unified research framework, we discuss the shared goal of increased realism through both ecological and temporal complexity, with the overarching aim of improving predictive power. In a rapidly changing world, advancing our understanding of the cumulative ecological impacts of multiple stressors is critical for biodiversity conservation and ecosystem management. Identifying and overcoming the barriers to interdisciplinary knowledge exchange is necessary in rising to this challenge. Division between ecosystem types and disciplines is largely a human creation. Species and stressors cross these borders and so should the scientists who study them.

Ocean acidification (OA) and high light was found to negatively affect the Antarctic key species Phaeocystis antarctica, Fragilariopsis kerguelensis and Chaetoceros debilis. To unravel the underlying physiological response at the transcriptomic level, these species were grown under ambient and elevated pCO2 combined with low or high light. RNA sequencing revealed that the haptophyte was much more tolerant towards OA than the two diatoms as only these showed distinct OA-dependent gene regulation patterns. Under ambient pCO2, high light resulted in decreased glycolysis in P. antarctica. Contrastingly, upregulation of genes related to cell division and transcription as well as reduced expression of both cata- and anabolic carbon related pathways were seen in C. debilis. OA in combination with low light led to reduced respiration, but also surprisingly to higher expression of genes involved in light protection, transcription and translation in C. debilis. Though not affecting P. antarctica, OA combined with high light caused also photosensitivity in both diatoms. As additional response reallocation of carbon to lipids was found in C. debilis under these conditions. Overall, we conclude that P. antarctica is better adapted than the two diatoms to OA and high light.

Hosts face mortality from parasitic and environmental stressors, but interactions of parasitism with other stressors are not well understood, particularly for long-lived hosts. We monitored survival of flour beetles (Tribolium confusum) in a longitudinal design incorporating cestode (Hymenolepis diminuta) infection, starvation and exposure to the pesticide diatomaceous earth (DE). We found that cestode cysticercoids exhibit increasing morphological damage and decreasing ability to excyst over time, but were never eliminated from the host. In the presence of even mild environmental stressors, host lifespan was reduced sufficiently that extensive degradation of cysticercoids was never realized. Median host lifespan was 200 days in the absence of stressors, and 3-197 days with parasitism, starvation and/or DE. Early survival of parasitized hosts was higher relative to controls in the presence of intermediate concentrations of DE, but reduced under all other conditions tested. Parasitism increased host mortality in the presence of other stressors at times when parasitism alone did not cause mortality, consistent with an interpretation of synergy. Environmental stressors modified the parasite numbers needed to reveal intensity-dependent host mortality, but only rarely masked intensity dependence. The longitudinal approach produced observations that would have been overlooked or misinterpreted if survival had only been monitored at a single time point.