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Environmental risk assessment of metal mixtures is challenging due to the large number of possible mixtures and interactions. Mixture toxicity data cannot realistically be generated for all relevant scenarios. Therefore, methods for prediction of mixture toxicity from single-metal toxicity data are needed. We tested how well toxicity of Cu-Ni-Zn mixtures to Daphnia magna populations can be predicted based on the Dynamic Energy Budget theory with an individual-based model (DEB-IBM), assuming non-interactivity of metals on the physiological level. We exposed D. magna populations to Cu, Ni, and Zn and their mixture at a fixed concentration ratio. We calibrated the DEB-IBM with single-metal data and generated blind predictions of mixture toxicity (population size over time), with account for uncertainty. We compared the predictive performance of the DEB-IBM with respect to mixture effects on population density and population growth rates with that of two reference models applied on the population level, independent action and concentration addition. Our inferred physiological modes of action (pMoA) differed from literature-reported pMoAs, raising the question of whether this is a result of different model selection approaches, intraspecific variability, or whether different pMoAs might actually drive toxicity in a population context. Observed mixture effects were concentration- and endpoint-dependent. The independent action was overall more accurate than the concentration addition but concentration addition-predicted effects on population growth rate were slightly better. The DEB-IBM most accurately predicted effects on 6-week density, including antagonistic effects at high concentrations, which emerged from non-interactivity at the physiological level. Mixture effects on initial population growth rate appear to be more difficult to predict. To explain why model accuracy is endpoint-dependent, relationships between individual-level and population-level endpoints should be illuminated. Environ Toxicol Chem 2021;40:3034-3048. © 2021 SETAC.

The global consumption of human antidepressants has steadily increased over the last years. The most widely prescribed antidepressants are the selective serotonin reuptake inhibitors (SSRIs), which have been linked to various life-history effects in nontarget organisms. We investigated the effects of the SSRI citalopram hydrobromide on the life history of the copepod Nitocra spinipes. Slight but significant developmental delay effects were observed at nominal concentrations of 0.1 and 1 µg/L, with stronger effects occurring at measured concentrations of 178 µg/L and above. At 77 µg/L and above, a significant increase in adult body length and offspring production/brood was found, although the time between brood releases remained unaffected. The pre-adult surviving fraction was significantly reduced (by 44%) at 765 µg/L. For a mechanistic evaluation of these observations, we used a bioenergetics model for N. spinipes based on the dynamic energy budget theory. Toxicokinetic and toxicodynamic submodels were used to dynamically simulate the chemical uptake and elimination, as well as dose-response relationships for hypothetical physiological modes of action and survival over time. Although none of the commonly invoked physiological modes of action, acting on assimilation, maintenance, growth, or offspring production, could explain the observed combination of effects, a newly proposed physiological mode of action acting on the process of maturation delivered correct predictions in terms of each effect's direction. The model fits could be further improved by allowing for a gentler concentration-effect slope and by adding an auxiliary physiological mode of action acting on the reproduction efficiency. The quantitative explanations provided in the present study offer a starting point for exploratory simulation studies investigating the effects of SSRIs at higher ecological levels. Environ Toxicol Chem 2021;40:1928-1939. © 2021 SETAC.

Neonicotinoid insecticides have become of global concern for the aquatic environment. Harpacticoid copepods are among the organisms most sensitive to neonicotinoids. We exposed the brackish copepod Nitocra spinipes to 4 neonicotinoid insecticides (clothianidin, imidacloprid, thiacloprid, and thiamethoxam) to investigate acute toxicity on adults (96-h exposure) and effects on larval development (7-d exposure). We used these results in combination with publicly available ecotoxicity data to derive environmental quality standards (EQS). These EQS were ultimately used in a single-substance and mixture risk assessment for the Belgian part of the North Sea. Acute toxicity testing revealed that immobilization is a more sensitive endpoint than mortality, with 96-h median effect concentration (EC50) values of 6.9, 7.2, 25, and 120 µg L-1 for clothianidin, thiacloprid, imidacloprid, and thiamethoxam, respectively. In addition, the larval development tests resulted in 7-d no-observed-effect concentrations (NOECs) of 2.5, 2.7, 4.2, and >99 µg L-1 for clothianidin, thiacloprid, imidacloprid, and thiamethoxam, respectively. The derived saltwater annual average (AA-)EQS were 0.05, 0.0048, 0.002, and 0.016 µg L-1 for clothianidin, thiacloprid, imidacloprid, and thiamethoxam, respectively. Finally, the risk characterization revealed some exceedances of the AA-EQS in Belgian harbors for imidacloprid (number of exceedances, n = 2/4), for thiacloprid (n = 1/4), for thiamethoxam (n = 1/4), and for the mixture of the 4 neonicotinoids (n = 4/4), but not at the open sea. At the open sea site, the toxic unit sums relative to the AA-EQS were 0.72 and 0.22, suggesting no mixture risk, albeit with a relatively small margin of safety. Including short-term EC10 (96-h) values of N. spinipes for the AA-EQS derivation led to a refinement of the AA-EQS for clothianidin and thiamethoxam, suggesting their use for the AA-EQS derivation because one of the overarching goals of the definition of EQS is to protect species at the population level. Environ Toxicol Chem 2021;40:1353-1367. © 2021 SETAC.

Although easily collected in large numbers, the subsequent processing and identification of zooplankton have usually been a barrier to large-scale biodiversity assessments. Therefore, DNA barcoding has been increasingly used by non-taxonomists to identify specimens. Here, we studied the community composition of zooplankton in the Belgian part of the North Sea over the course of 1 year. We identified zooplankton using both a traditional approach based on morphological characteristics and by metabarcoding of a 650 bp fragment of the V4-V5 region of the 18S rRNA gene using nanopore sequencing. Using long rDNA sequences, we were able to identify several taxa at the species level, across a broad taxonomic scale. Using both methods, we compared community composition and obtained diversity metrics. Diversity indices were not significantly correlated. Metabarcoding allowed for comparisons of diversity and community composition, but not all groups were successfully sequenced. Additionally, some disparities existed between relative abundances of the most abundant taxa based on traditional counts and those based on sequence reads. Overall, we conclude that for zooplankton samples, metabarcoding is capable of detecting taxa with a higher resolution, regardless of developmental stage of the organism. Combination of molecular and morphological methods results in the highest detection and identification levels of zooplankton.

Mechanistic population models are gaining considerable interest in ecological risk assessment. The dynamic energy budget approach for toxicity (debtox) and the general unified threshold model for survival (guts) is a well-established theoretical framework that describes sub-lethal and lethal effects of a chemical stressor, respectively. However, there have been limited applications of these models for mixtures of chemicals, especially to predict long-term effects on populations. Here, we used DEBtox and GUTS in an Individual-Based Model (IBM) framework to predict both single and combined effects of copper and zinc on Daphnia magna populations. The model was calibrated based on standard chronic toxicity test results with the single substances. A mixture toxicity implementation based on the general independent action model for mixtures was developed and validated with data from a population experiment with copper and zinc mixtures. Population-level effects of exposure to individual metals were accurately predicted by DEB-IBM. The DEB-IBM framework also allowed to identify the potential mechanisms underlying these observations. Under independent action the DEB-IBM was able to predict the population dynamics observed in populations exposed to the single metals and their mixtures (R2 > 65% in all treatments). Our modelling shows that it is possible to extrapolate from single-substance effects at the individual level to mixture toxicity effects at the population level, without the need for mixture toxicity data at the individual level from standard mixture toxicity tests. The application of such modelling techniques can increase the ecological realism in risk assessment. This article is protected by copyright. All rights reserved.

Marine phytoplankton influence the composition of sea spray aerosols (SSAs) by releasing various compounds. The biogenic surfactant dipalmitoylphosphatidylcholine (DPPC) is known to accumulate in the sea surface microlayer, but its aerosolization has never been confirmed. We conducted a 1 year SSA sampling campaign at the Belgian coast and analyzed the SSA composition. We quantified DPPC at a median and maximum air concentration of 7.1 and 33 pg m-3, respectively. This discovery may be of great importance for the field linking ocean processes to human health as DPPC is the major component of human lung surfactant and is used as excipient in medical aerosol therapy. The natural airborne exposure to DPPC seems too low to induce direct human health effects but may facilitate the effects of other marine bioactive compounds. By analyzing various environmental variables in relation to the DPPC air concentration, using a generalized linear model, we established that wave height is a key environmental predictor and that it has an inverse relationship. We also demonstrated that DPPC content in SSAs is positively correlated with enriched aerosolization of Mg2+ and Ca2+. In conclusion, our findings are not only important from a human health perspective but they also advance our understanding of the production and composition of SSAs.

Since the early 2000s, biotic ligand models and related constructs have been a dominant paradigm for risk assessment of aqueous metals in the environment. We critically review 1) the evidence for the mechanistic approach underlying metal bioavailability models; 2) considerations for the use and refinement of bioavailability-based toxicity models; 3) considerations for the incorporation of metal bioavailability models into environmental quality standards; and 4) some consensus recommendations for developing or applying metal bioavailability models. We note that models developed to date have been particularly challenged to accurately incorporate pH effects because they are unique with multiple possible mechanisms. As such, we doubt it is ever appropriate to lump algae/plant and animal bioavailability models; however, it is often reasonable to lump bioavailability models for animals, although aquatic insects may be an exception. Other recommendations include that data generated for model development should consider equilibrium conditions in exposure designs, including food items in combined waterborne-dietary matched chronic exposures. Some potentially important toxicity-modifying factors are currently not represented in bioavailability models and have received insufficient attention in toxicity testing. Temperature is probably of foremost importance; phosphate is likely important in plant and algae models. Acclimation may result in predictions that err on the side of protection. Striking a balance between comprehensive, mechanistically sound models and simplified approaches is a challenge. If empirical bioavailability tools such as multiple-linear regression models and look-up tables are employed in criteria, they should always be informed qualitatively and quantitatively by mechanistic models. If bioavailability models are to be used in environmental regulation, ongoing support and availability for use of the models in the public domain are essential. Environ Toxicol Chem 2019;39:60-84. © 2019 SETAC.

In the present study, we aimed to test the protectiveness of the bioavailability-normalization procedure, with its associated hazardous concentrations for x% of the species (HCx), that is currently implemented to derive environmental threshold concentrations for nickel (Ni) in European environmental legislative frameworks. We exposed a natural plankton-dominated community to 3 constant Ni concentrations, that is, a control with no Ni added (background Ni of 1.2-4 µg/L) and the bioavailability-normalized HC5 and HC50 of 24 and 97 µg dissolved Ni/L, respectively, during a 56-d microcosm experiment under high dissolved organic carbon (DOC) conditions (DOC of 14 mg/L at test initiation). The effects of the bioavailability-normalized HC5 and HC50 values were evaluated at the levels of community structure (community composition and plankton group abundances), community functioning (measured as indirect physicochemical proxies for overnight respiration and carbon fluxes), and individual species abundances. The bioavailability-normalized HC50 treatment had clear effects (defined as effects occurring on at least 2 consecutive sampling days) on both the structure and functioning of the investigated aquatic community. Through its effect on community functioning (i.e., reduced pH and DOC), Ni also influenced its own bioavailability. Clear direct effects of Ni were observed for only 3 species (the Cyanobacteria Oscillatoria sp. 1 and the rotifers Asplanchna/Testidunela sp. and Trichocerca group similis). Most other effects occurring in the plankton community in the HC50 treatment were indirect and likely driven by the direct effect of Ni on the Cyanobacteria Oscillatoria sp. 1, which was the dominant phytoplankton species in the control microcosms. In contrast, the bioavailability-normalized HC5 did not induce clear effects on community structure and functioning endpoints: these were only affected on individual sampling days. Clear (direct) effects were observed for only 2 plankton species (the rotifer Trichocerca group similis and the Cyanobacteria Oscillatoria sp. 1), but their abundances recovered to control levels at the end of the study. In addition, a few species (1 phytoplankton and 3 zooplankton species) were affected in the HC5 treatment only on the last sampling day. It is uncertain whether these species would have shown clear effects over a longer exposure duration. Thus, our study shows that the bioavailability-normalized HC5 of Ni at high DOC induced clear effects on a few individual species. However, the overall conclusion is that the bioavailability-normalized HC5 of Ni as derived through the procedure that is currently implemented in European legislative frameworks protects against clear effects on community structure and function. Environ Toxicol Chem 2019;38:1923-1939. © 2019 SETAC.

Population models are increasingly being used to extrapolate individual-level effects of chemicals, including metals, to population-level effects. For metals, it is also important to take into account their bioavailability to correctly predict metal toxicity in natural waters. However, to our knowledge, no models exist that integrate metal bioavailability into population modeling. Therefore, our main aims were to 1) incorporate the bioavailability of copper (Cu) and zinc (Zn) into an individual-based model (IBM) of rainbow trout (Oncorhynchus mykiss), and 2) predict how survival-time concentration data translate to population-level effects. For each test water, reduced versions of the general unified threshold model of survival (GUTS-RED) were calibrated using the complete survival-time concentration data. The GUTS-RED individual tolerance (IT) showed the best fit in the different test waters. Little variation between the different test waters was found for 2 GUTS-RED-IT parameters. The GUTS-RED-IT parameter "median of distribution of thresholds" (mw ) showed a strong positive relation with the Ca2+ , Mg2+ , Na+ , and H+ ion activities. Therefore, mw formed the base of the calibrated GUTS bioavailability model (GUTS-BLM), which predicted 30-d x% lethal concentration (LCx) values within a 2-fold error. The GUTS-BLM was combined with an IBM, inSTREAM-Gen, into a GUTS-BLM-IBM. Assuming that juvenile survival was the only effect of Cu and Zn exposure, population-level effect concentrations were predicted to be 1.3 to 6.2 times higher than 30-d laboratory LCx values, with the larger differences being associated with higher interindividual variation of metal sensitivity. The proposed GUTS-BLM-IBM model can provide insight into metal bioavailability and effects at the population level and could be further improved by incorporating sublethal effects of Cu and Zn. Environ Toxicol Chem 2021;40:2764-2780. © 2021 SETAC.

Predicting metal sensitivities and metal mixture interactions for species within each trophic level is essential to understand the effects of metals at the ecosystem level. The present study was set up to explore the correlations of metal sensitivities among species and if these sensitivities or metal mixture interactions are related to growth or morphological traits. The toxicity of Ni, Cu, and Zn on algal growth was tested for 8 freshwater algal species when dosed singly and in combinations in phosphorus-limiting static systems. The metal sensitivities on specific growth rate (10% effect concentrations expressed as free ion activities) varied 2 to 3 orders of magnitude among species depending on metal. These sensitivities were unrelated (p > 0.05) to their specific growth rate (0.7-1.8 d-1 ) or cell volume (100 -103 m3 cell-1 ). Species-specific differences in one or more toxicokinetic and/or toxicodynamic (TKTD) processes are likely at the basis of this variation. The log-transformed metal sensitivities positively correlated (p

Driven by Regulation (EC) No. 1272/2008 and the European Water Framework Directive 2000/60/EC, we have re‐evaluated the available chronic freshwater ecotoxicity data for ionic silver (Ag) using strict data quality criteria. In addition, we generated new chronic ecotoxicity data for species potentially sensitive to Ag (the rotifer Brachionus calyciflorus, the cyanobacteria Anabaena flos‐aquae, and the aquatic plant Lemna minor) using Ag nitrate as the test substance. The 10% effect concentrations for the most sensitive endpoint per test species were 0.31 µg dissolved Ag/L for B. calyciflorus (population size), 0.41 µg dissolved Ag/L for A. flos‐aquae (growth rate), and 1.40 µg dissolved Ag/L for L. minor (root length). We included these values in the set of reliable chronic freshwater data, subsequently covering a total of 12 taxonomic groups and 15 species. Finally, we applied a species sensitivity distribution approach to the data set using various models. The best‐fitting model (Rayleigh distribution) resulted in a threshold value protective for 95% of the species of 0.116 µg dissolved Ag/L. This value is considered reliable and conservative in terms of species protection and can be used as a solid basis for setting thresholds for Ag in freshwater after application of an appropriate assessment factor. Furthermore, this value represents reasonable worst‐case conditions for bioavailability in European Union surface waters (low hardness and low dissolved organic carbon). Environ Toxicol Chem 2021;40:1678–1693. © 2021 European Precious Metals Federation. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

The increasing number of chemicals detected in the marine environment underlines the need for appropriate prioritization strategies prior to further testing and potential inclusion into monitoring programs. Here, a prioritization strategy is proposed for chemicals detected in the North Sea over the last decade, through the development of a Concern Index (CI) using exposure and toxicity data obtained from peer-review publications and the ToxCast database, respectively. A total of 158 chemicals were ranked and the most sensitive tested assay endpoints were identified. Additionally, similar analysis was performed for the classes of chemicals and Biological Process Targets (BPTs). By first ranking chemicals currently acknowledged for their high toxicity to the aquatic environment, i.e. naphthalene, salicylic acid and simazine, the obtained results not only reinforce the risk posed by these but also promote a confident extrapolation from mammalian in vitro toxicity data to fish. Furthermore, genes targeted by the most sensitive assays, related to basic cell maintenance processes and immune defense, are highly evolutionarily conserved across species. The identification of these assays further reinforces the importance of a shift from traditional toxicity endpoints to lower levels of biological organization, allowing the detection of adverse effects at lower concentrations.

The present study aimed at investigating interactive effects between dietary lipids and both short- and long-term exposures to a low, environmentally realistic, cadmium (Cd) concentration. Juvenile rainbow trout were fed four isolipidic diets (31.7 g/kg) enriched in either linoleic acid (LA, 18:2n-6), alpha-linolenic acid (ALA, 18:3n-3), eicosapentaenoic acid (EPA, 20:5n-3) or docosahexaenoic acid (DHA, 22:6n-3). From the 4th week of this 10-week experiment, the lipid level of the diet was increased (120.0 g/kg) and half of the fish fed each diet were aqueously exposed to Cd (0.3 μg/L) while the other half were not exposed to Cd (control). Fish were sampled and their liver was harvested for fatty acid profile, hepatic Cd and calcium concentrations, total glutathione level and gene expression assessment, either (i) after 4 weeks of feeding and 24 h of Cd contamination (day 29) (short-term Cd exposure) or (ii) after 10 weeks of feeding and 6 weeks of Cd contamination (day 70) (long-term Cd exposure). We found that both dietary lipids and Cd exposure influenced fatty acid homeostasis and metabolism. The hepatic fatty acid profile mostly reflected that of the diet (e.g. n-3/n-6 ratio) with some differences, including selective retention of specific long chain polyunsaturated fatty acids (LC-PUFAs) like DHA and active biotransformation of dietary LA and ALA into LC-PUFAs. Cd effects on hepatic fatty acid profiles were influenced by the duration of the exposure and the nutritional status of the fish. The effects of diet and Cd exposure on the fatty acid profiles were only sparsely explained by variation of the expression pattern of genes involved in fatty acid metabolism. The biological responses to Cd were also influenced by dietary lipids. Fish fed the ALA-enriched diet seemed to be the least affected by the Cd exposure, as they showed a higher detoxifying ability against Cd with an early upregulation of protective metallothionein a (MTa) and apoptosis regulator BCL2-Like1 (BCLx) genes, an increased long-term phospholipid synthesis and turnover and fatty acid bioconversion efficiency, as well as a lower long-term accumulation of Cd in their liver. In contrast, fish fed the EPA-enriched diet seemed to be the most sensitive to a long-term Cd exposure, with an impaired growth performance and a decreased antioxidant capacity (lower glutathione level). Our results highlight that low, environmentally realistic aqueous concentrations of Cd can affect biological response in fish and that these effects are influenced by the dietary fatty acid composition.

Organisms in the marine environment are being exposed to an increasing variety of chemicals. This research presents an effect-based monitoring method for the derivation of a margin of safety for environmentally realistic chemical mixtures. The method is based on a combination of passive sampling and ecotoxicity testing. First, passive sampling was performed using H2O-philic divinylbenzene Speedisks during 3 sampling campaigns between 2016 and 2018 at 4 sampling locations in the Belgian part of the North Sea. Next, we exposed the marine diatom Phaeodactylum tricornutum to Speedisk extracts that were reconstituted in HPLC-grade water and defined the MoS of each sample as the highest no-observed effect concentration, expressed as relative enrichment factor (REF). A REF was defined by comparing the concentrations of 89 personal care products, pesticides and pharmaceuticals in the biotest medium with those measured in water grab samples to relate exposure concentrations in the tests to environmental concentrations. Across eight marine samples, diatom growth inhibition was observed at REF ≥ 3.2 and margins of safety were found between REF 1.1–11.0. In addition, we found that reconstitution of extracts in HPLC-water was suitable to overcome the solvent-related challenges in biotesting that are usually associated with passive sampler extract spiking, whilst it still allowed REFs up to 44 in the biotest medium to be achieved. This method, however, likely covers mainly the polar fraction of environmentally realistic chemical mixtures and less the non-polar fraction. Nevertheless, for 5 out of 8 samples, the Margin of Safety (MoS) was found to be lower than 10, which represents the typically lowest possible assessment factor applied to no effects ecotoxicological data in conventional environmental risk assessments, suggesting ecological risks for these samples.

The harpacticoid copepod Nitocra spinipes is a commonly used test species in ecotoxicological studies and subject of multiple international testing guidelines. While we strive for a better understanding of toxicant-induced effects in this species, physiological models rooted in the Dynamic Energy Budget (DEB) theory can help to explain such effects on the level of energy allocations within the organism. In previous works that aimed to capture the copepod life history in a DEB model, diverging assumptions on the growth pattern of copepods were made. While some authors presumed von Bertalanffy growth, others presumed an upcurving in length versus time due to metabolic acceleration. In this study we parametrized the two typified DEB models “abp” (metabolic acceleration from birth to puberty) and “sbp” (standard von Bertalanffy growth from birth to puberty) on life history data of N. spinipes. Besides using data from literature, we also measured additional length-at-time data to aid the parameter estimation. As the body proportions of N. spinipes changed continuously throughout its development, we used the square root of the top view area as a length measure to scale with the cube root of structural volume in length-to-volume conversions. Experimental data were predicted well with both models according to goodness of fit criteria. Despite a slightly better data fit in abp, we cannot rule out sbp as implausible. Overall, we expect both models to perform equally well in future applications. More detailed data on N. spinipes and closely related species are needed to support or reject the presumption of metabolic acceleration in the life history of copepods.

Besides conventional scanning X-ray fluorescence imaging at synchrotron sources, full-field X-ray fluorescence (FF-XRF) imaging techniques that do not implicitly require spatial scanning of the sample have become available. FF-XRF has become achievable thanks to the development of a new type of energy dispersive CCD-based 2D detector, also referred to as a ‘color X-ray camera (CXC)’ or ‘SLcam’. We report on different imaging schemes for biological samples using FF-XRF imaging: (a) 2D ‘zoom’ imaging with pinhole optics using the ‘camera obscura’ principle; (b) 2D ‘fixed magnification’ imaging using magnifying polycapillary optics; and (c) 3D-FF-XRF imaging using an X-ray sheet beam or computed tomography (CT). The different FF-XRF imaging modes are illustrated using the crustacean Daphnia magna, a model organism for investigating the effects of metals on organism/ecosystem health, and foraminifera, a class of amoeboid protist. Detailed analytical characterization of the set-up is performed through analyzing various reference materials in order to determine limits of detection (LODs) and sensitivities. Experiments were performed using the BAMline at the BESSY synchrotron (Berlin, Germany) and using the P06 Hard X-ray Microprobe at the PETRAIII synchrotron (Hamburg, Germany).

Sea spray aerosols (SSAs) have profound effects on our climate and ecosystems. They also contain microbiota and biogenic molecules which could affect human health. Yet the exposure and effects of SSAs on human health remain poorly studied. Here, we exposed human lung cancer cells to extracts of a natural sea spray aerosol collected at the seashore in Belgium, a laboratory-generated SSA, the marine algal toxin homoyessotoxin and a chemical inhibitor of the mammalian target of rapamycin (mTOR) pathway. We observed significant increased expression of genes related to the mTOR pathway and Proprotein convertase subtilisin/kexin type 9 (PCSK9) after exposure to homoyessotoxin and the laboratory-generated SSA. In contrast, we observed a significant decrease in gene expression in the mTOR pathway and of PCSK9 after exposure to the natural SSA and the mTOR inhibitor, suggesting induction of apoptosis. Our results indicate that marine biogenics in SSAs interact with PCSK9 and the mTOR pathway and can be used in new potential pharmaceutical applications. Overall, our results provide a substantial molecular evidence base for potential beneficial health effects at environmentally relevant concentrations of natural SSAs.

Australian freshwaters have relatively low water hardness and different calcium (Ca) to magnesium (Mg) ratios compared with those in Europe. The hardness values of a substantial proportion of Australian freshwaters fall below the application boundary of the existing European nickel biotic ligand models (Ni BLMs) of 2 mg Ca/L. Toxicity testing was undertaken using Hydra viridissima to assess the predictive ability of the existing Ni BLM for this species in extremely soft waters. This testing revealed an increased competitive effect of Ca and Mg with Ni for binding to the biotic ligand in soft water (

It is generally assumed that as long as the majority of species experiences no direct adverse effects attributable to a single substance (i.e., potentially affected fraction [PAF]5%), no significant structural or functional effects at the community level are expected to occur. Whether this assumption holds for mixed metal contamination is not known. In the present study, we tested this by performing a microcosm experiment in which a naturally occurring freshwater planktonic community was exposed to a copper-nickel-zinc (Cu-Ni-Zn) mixture for 8 wk and various structural and functional community-level traits were assessed. In the low mixture concentration treatments (i.e., Ni-Zn mixtures, because there was no difference in Cu concentrations in these treatments with the control), community-level effects were relatively simple, only involving phytoplankton species groups. In the high mixture concentration treatments (Cu-Ni-Zn mixtures), community-level effects were more complex, involving several phytoplankton and zooplankton species groups. Multisubstance PAF (msPAF) values for all mixture treatments were calculated by applying the concentration addition model to bioavailability-normalized single-metal species sensitivity distributions (SSDs). Consistent effects on the structural traits community composition, abundance of zooplankton species groups, species diversity, and species richness and on the functional trait dissolved organic carbon (DOC) concentration (as a proxy for the microbial loop and pelagic food web interactions) were only observed at msPAF values0.05 (i.e., in the Cu-Ni-Zn mixture). However, consistent effects on the abundance of various phytoplankton species groups (structural traits) and on 2 measures of community respiration, overnight Δ dissolved oxygen (ΔDO) and ΔpH (functional traits), were already observed at msPAF values of ≤0.05 (i.e., in the Ni-Zn mixture). This indicates that the threshold msPAF value of 0.05 was not protective against metal mixture exposure for all community-level structural and functional endpoints in the present study. A possible explanation for this result is the mismatch between the species in the SSD and those in our microcosm community. Indeed, our data suggest that the presence of one single dominant and very Zn- and/or Ni-sensitive species in the investigated community (i.e., a cyanobacteria of the genus Oscillatoria), which is not represented in the SSD of these metals, was probably the driver of all observed effects at or below an msPAF of 0.05. Overall, the present results show that SSDs are not necessarily a good predictor of community-level effects for all types of communities and that the presence of dominant sensitive species may result in significant, consistent effects on certain structural and functional community-level endpoints at msPAF values ≤0.05, which is generally considered protective in many regulatory frameworks. Environ Toxicol Chem 2018;37:2380-2400. © 2018 SETAC.

Ecological interactions and abiotic stress factors may significantly affect species sensitivities to toxicants, and these are not incorporated in standard single-species tests. The present study tests whether a model, calibrated solely on single-species data, can explain abiotic stress factors in a two-species microcosm, a test applied to the effects of nutritional stress (phosphorus [P] limitation) on zinc (Zn) toxicity to Daphnia magna. A population model was developed based on P- and Zn-dependent algal and daphnid growth. Two separate two-species (phytoplankton Pseudokirchneriella subcapitata and consumer D. magna) microcosm experiments with P × Zn factorial combinations and a different pH (7.3 and 7.8) were set up to validate the model. The 21-d daphnid population size was considerably reduced by increased Zn and by decreased P supply, with a significant (p < 0.001) interaction between the 2 factors. The observed median effective concentration (EC50) of Zn on D. magna population size varied 12-fold (25 to 310 μg Zn L-1 ), with the lowest EC50 values found at the highest pH and high P treatments. For both experiments, Zn toxicity to D. magna was correctly predicted within a factor of 2 for EC50 values, and this is explained by the model through 1) a higher phytoplankton Zn sensitivity at higher pH, affecting food supply to D. magna, and 2) an increased algal P content at higher Zn, offering a nutritional benefit to daphnids that counteracts direct Zn toxicity under P limitation. The present study illustrates that indirect effects of Zn via producer-consumer relationships can outweigh the direct toxic effects and that models calibrated solely on single-species test data can help with interpreting these results in two-species systems. Environ Toxicol Chem 2018;37:2153-2164. © 2018 SETAC.

Under natural conditions, organisms can experience a variety of abiotic (e.g., temperature, pH) and biotic (e.g., species interactions) factors, which can interact with toxicant effects. By ignoring species interactions conventional ecotoxicological studies (i.e., single-species tests) oversimplify the actual field situation. We investigated whether temperature and interspecific competition affected the effects of zinc (Zn) on a Daphnia longispina population. The D. longispina populations were exposed in a full factorial design to 3 different Zn treatments (background, 29, and 110 μg Zn/L), 2 different temperature regimes (cold, 17-18 °C; warm, 21-22 °C), and 2 interspecific competition levels (no interspecific Brachionus competition = no Brachionus calyciflorus added; interspecific Brachionus competition = B. calyciflorus added). Interspecific Brachionus competition and temperature by itself had a limited effect on the Daphnia abundance but significantly interacted with the highest Zn concentration. Without Brachionus competition the D. longispina juvenile and adult abundances under warm conditions were up to 5.5 and 21 times lower, respectively, in the high Zn treatment in comparison with the Zn control, whereas under cold conditions no significant Zn effect was observed. However, with Brachionus competition the highest Zn treatment was on average 2.2 times less toxic to the D. longispina juvenile abundance at higher temperatures. Under cold conditions the highest Zn treatment affected the juvenile abundance sooner and up to 9 times more negatively when simultaneously faced with Brachionus competition. It is possible that the competition for food reduced the amount of energy that could be used by D. longispina for reproduction, and the metabolic costs increased as a result of Zn stress. The present study clearly illustrated the influence of temperature and competition on the effects of a chemical stressor. Thus, not considering such factors in ecological risk assessment may underestimate or overestimate risks in aquatic ecosystems when extrapolating data from standard single-species tests to the field. Environ Toxicol Chem 2018;37:1668-1678. © 2018 SETAC.

International audience; Our aim was to investigate epigenetic changes in Daphnia magna after a 25-day chronic external γ irradiation (generation F0 exposed to 6.5 μGy·h-1 or 41.3 mGy·h-1) and their potential inheritance by subsequent recovering generations, namely, F2 (exposed as germline cells in F1 embryos) and F3 (the first truly unexposed generation). Effects on survival, growth, and reproduction were observed and DNA was extracted for whole-genome bisulfite sequencing in all generations. Results showed effects on reproduction in F0 but no effect in the subsequent generations F1, F2, and F3. In contrast, we observed significant methylation changes at specific CpG positions in every generation independent of dose rate, with a majority of hypomethylation. Some of these changes were shared between dose rates and between generations. Associated gene functions included gene families and genes that were previously shown to play roles during exposure to ionizing radiation. Common methylation changes detected between generations F2 and F3 clearly showed that epigenetic modifications can be transmitted to unexposed generations, most likely through the germline, with potential implications for environmental risk. © 2018 American Chemical Society.

Although metal mixture toxicity has been studied relatively intensely, there is no general consensus yet on how to incorporate metal mixture toxicity into aquatic risk assessment. We combined existing data on chronic metal mixture toxicity at the species level with species sensitivity distribution (SSD)-based in silico metal mixture risk predictions at the community level for mixtures of Ni, Zn, Cu, Cd, and Pb, to develop a tiered risk assessment scheme for metal mixtures in freshwater. Generally, independent action (IA) predicts chronic metal mixture toxicity at the species level most accurately, whereas concentration addition (CA) is the most conservative model. Mixture effects are noninteractive in 69% (IA) and 44% (CA) and antagonistic in 15% (IA) and 51% (CA) of the experiments, whereas synergisms are only observed in 15% (IA) and 5% (CA) of the experiments. At low effect sizes (∼ 10% mixture effect), CA overestimates metal mixture toxicity at the species level by 1.2-fold (i.e., the mixture interaction factor [MIF]; median). Species, metal presence, or number of metals does not significantly affect the MIF. To predict metal mixture risk at the community level, bioavailability-normalization procedures were combined with CA or IA using SSD techniques in 4 different methods, which were compared using environmental monitoring data of a European river basin (the Dommel, The Netherlands). We found that the simplest method, in which CA is directly applied to the SSD (CASSD ), is also the most conservative method. The CASSD has median margins of safety (MoS) of 1.1 and 1.2 respectively for binary mixtures compared with the theoretically more consistent methods of applying CA or IA to the dose-response curve of each species individually prior to estimating the fraction of affected species (CADRC or IADRC ). The MoS increases linearly with an increasing number of metals, up to 1.4 and 1.7 for quinary mixtures (median) compared with CADRC and IADRC , respectively. When our methods were applied to a geochemical baseline database (Forum of European Geological Surveys [FOREGS]), we found that CASSD yielded a considerable number of mixture risk predictions, even when metals were at background levels (8% of the water samples). In contrast, metal mixture risks predicted with the theoretically more consistent methods (e.g., IADRC ) were very limited under natural background metal concentrations (

Urban regions of the world are expanding rapidly, placing additional stress on water resources. Urban water bodies serve many purposes, from washing and sources of drinking water to transport and conduits for storm drainage and effluent discharge. These water bodies receive chemical emissions arising from either single or multiple point sources, diffuse sources which can be continuous, intermittent, or seasonal. Thus, aquatic organisms in these water bodies are exposed to temporally and compositionally variable mixtures. We have delineated source-specific signatures of these mixtures for diffuse urban runoff and urban point source exposure scenarios to support risk assessment and management of these mixtures. The first step in a tiered approach to assessing chemical exposure has been developed based on the event mean concentration concept, with chemical concentrations in runoff defined by volumes of water leaving each surface and the chemical exposure mixture profiles for different urban scenarios. Although generalizations can be made about the chemical composition of urban sources and event mean exposure predictions for initial prioritization, such modeling needs to be complemented with biological monitoring data. It is highly unlikely that the current paradigm of routine regulatory chemical monitoring alone will provide a realistic appraisal of urban aquatic chemical mixture exposures. Future consideration is also needed of the role of nonchemical stressors in such highly modified urban water bodies. Environ Toxicol Chem 2018;37:703-714. © 2017 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals, Inc. on behalf of SETAC.

Freshwater biota are usually exposed to mixtures of different metals in the environment, which raises concern because risk-assessment procedures for metals are still mainly based on single-metal toxicity. Because microalgae are primary producers and therefore at the base of the food web, it is of utmost importance to understand the effects of metal mixtures on these organisms. Most studies that have investigated the combined interactive effects of mixtures on microalgae performed tests in only one specific water. The objective of the present study was to test if combined effects of mixtures to Pseudokirchneriella subcapitata were the same or different across natural waters showing diverse water-chemistry characteristics. This was done by performing experiments with ternary Cu-Ni-Zn mixtures in 3 natural waters and with binary Cu-Ni mixtures in 5 natural waters. We showed that the ternary mixture acted noninteractively on algal growth, except in one water in which the mixture acted antagonistically. We suggest that a low-cationic competition situation in the latter water could be the reason for the antagonistic interaction between the metals. On the other hand, the binary mixture acted noninteractively on algal growth in all tested waters. We showed that both the concentration addition and independent action models can serve as accurate models for toxicity of ternary Cu-Ni-Zn and binary Cu-Ni mixtures to P. subcapitata in most cases and as protective models in all cases. In addition, we developed a metal mixture bioavailability model, by combining the independent action model and the single-metal bioavailability models, that can be used to predict Cu-Ni-Zn and Cu-Ni toxicity to P. subcapitata as a function of metal concentration and water characteristics. Environ Toxicol Chem 2018;37:587-598. © 2017 SETAC.

The effect of multiple stressors on marine ecosystems is poorly understood. To partially bridge this knowledge gap we investigated the relative contribution of environmental variables to density and diversity dynamics of the zooplankton community in the Belgian part of the North Sea. We applied multimodel inference on generalized additive models to quantify the relative contribution of chlorophyll a, temperature, nutrients, salinity and anthropogenic chemicals (i.e. polychlorinated biphenyls and polycyclic aromatic hydrocarbons) to the dynamics of calanoid copepod species in the Belgian part of the North Sea. Temperature was the only predictor consistently showing a high importance in all models predicting the abundances of the selected copepod species. The relative contribution of other predictors was species-dependent. Anthropogenic chemicals were important predictors for three out of six species indicating that chemical mixtures at low concentrations should not be left unattended when performing risk assessments in a natural environment.

Little is known about the effect of metal mixtures on marine organisms, especially after exposure to environmentally realistic concentrations. This information is, however, required to evaluate the need to include mixtures in future environmental risk assessment procedures. We assessed the effect of copper (Cu)-Nickel (Ni) binary mixtures on Mytilus edulis larval development using a full factorial design that included environmentally relevant metal concentrations and ratios. The reproducibility of the results was assessed by repeating this experiment 5 times. The observed mixture effects were compared with the effects predicted with the concentration addition model. Deviations from the concentration addition model were estimated using a Markov chain Monte-Carlo algorithm. This enabled the accurate estimation of the deviations and their uncertainty. The results demonstrated reproducibly that the type of interaction-synergism or antagonism-mainly depended on the Ni concentration. Antagonism was observed at high Ni concentrations, whereas synergism occurred at Ni concentrations as low as 4.9 μg Ni/L. This low (and realistic) Ni concentration was 1% of the median effective concentration (EC50) of Ni or 57% of the Ni predicted-no-effect concentration (PNEC) in the European Union environmental risk assessment. It is concluded that results from mixture studies should not be extrapolated to concentrations or ratios other than those investigated and that significant mixture interactions can occur at environmentally realistic concentrations. This should be accounted for in (marine) environmental risk assessment of metals. Environ Toxicol Chem 2017;36:3471-3479. © 2017 SETAC.

In the present study, we observed that the chronic Zn Daphnia magna biotic ligand model (BLM) could not accurately predict the chronic Zn toxicity to Ceriodaphnia dubia in a series of (modified) field waters, although the intrinsic sensitivities of the D. magna BLM was specifically calibrated on the sensitivity of C. dubia. Moreover, it was observed that the D. magna BLM underestimated the effect of pH on toxicity of the free Zn2+ ion. A preliminary species-specific chronic Zn C. dubia bioavailability model was developed based on the data. The preliminary Zn C. dubia bioavailability model was shown to predict Zn toxicity to C. dubia in a series of natural waters with pH ranging between 7 and 8.3 with reasonable accuracy. The difference in the effect of pH on Zn2+ toxicity between D. magna and C. dubia (two phylogenetically closely related species) suggested by our results deserves further attention, because of the importance of the cross-species application of bioavailability models in ecological risk assessment procedures and environmental quality standard derivations for metals. This article is protected by copyright. All rights reserved

In this work, the three-dimensional elemental, distribution profile within the freshwater crustacean Ceriodaphnia dubia was constructed at a spatial resolution down to S mu m via a data, fusion approach employing state-of-the-art laser ablation inductively coupled plasma-time-of-flight mass spectrometry (LAICP-TOFMS) and laboratory-based absorption microcomputed tomography (mu-CT). C. dubia was exposed to elevated Cu, Ni, and Zn concentrations, chemically fixed, dehydrated, stained, and embedded, prior to mu-CT analysis. Subsequently, the sample was cut into 5 pm thin sections that were subjected to LA-ICPTOFMS imaging. Multimodal image registration was performed to spatially align the 2D LA-ICP-TOFMS images relative to the Corresponding slices of the 3D mu-CT reconstruction. Mass channels corresponding to the isotopes of a single element were merged to improve the signal-to-noise ratios within the elemental images. In order to aid the visual interpretation of the data, LA-ICP-TOEMS data wete projected onto the mu-CT voxels representing tissue. Additionally, the image resolution and elemental sensitivity were compared to those obtained with synchrotron radiation based 3D confocal mu-X-ray fluorescence imaging upon a chemically fixed and air-dried C. dubia specimen.

Water quality standards for cobalt (Co) have not been developed for the European Union or United States. The objective of the present study was to produce freshwater Co toxicity data that could be used by both the European Union and the United States to develop appropriate regulatory standards (i.e., environmental quality standards or predicted-no-effect concentrations in Europe and ambient water quality criteria or state water quality standards in the United States). Eleven species, including algae, an aquatic plant, and several invertebrate and fish species, were used in the performance of acute and chronic Co toxicity tests. Acute median lethal or median effective concentration (LC50 or EC50) values ranged from 90.1 µg Co/L for duckweed (Lemna minor) to 157 000 µg Co/L for midges (Chironomus tentans). Chronic 10% effect concentration (EC10) values ranged from 4.9 µg Co/L for duckweed to 2170 µg Co/L for rainbow trout (Oncorhynchus mykiss). Chronic 20% effect concentration (EC20) values ranged from 11.1 µg Co/L for water flea (Ceriodaphnia dubia) to 2495 µg Co/L for O. mykiss. Results indicated that invertebrate and algae/plant species are more sensitive to chronic Co exposures than fish. Acute-to-chronic ratios (derived as acute LC50s divided by chronic EC20s) were lowest for juvenile O. mykiss (0.6) and highest for the snail Lymnaea stagnalis (2670). Following the European-based approach and using EC10 values, species sensitivity distributions (SSDs) were developed and a median hazardous concentration for 5% of the organisms of 1.80 µg Co/L was derived. Chronic EC20 values were used, also in an SSD approach, to derive a US Environmental Protection Agency-style final chronic value of 7.13 µg Co/L. Environ Toxicol Chem 2020;39:799-811. © 2020 SETAC.

The generalized bioavailability model (gBAM) has been proposed as an alternative to the biotic ligand model (BLM) for modeling bioavailability and chronic toxicity of copper (Cu). The gBAM combines a log-linear effect of pH on free Cu2+ ion toxicity with BLM-type parameters for describing the protective effects of major cations (calcium [Ca]2+ , magnesium [Mg]2+ , and sodium [Na]+ ). In the present study, a Windermere Humic Aqueous Model (WHAM) VII-based gBAM for fish was parametrized based on an existing chronic (30-d) dataset of juvenile rainbow trout (Oncorhynchus mykiss). The model, with defined parameters (pH slope parameter [SpH ] = 0.4449 and biotic ligand competition constants [log KCaBL = 4.0, log KMgBL = 3.4, and log KNaBL = 3.0]), was shown to accurately predict the effects of pH, dissolved organic carbon, Ca, and Mg on chronic Cu toxicity to juvenile rainbow trout at the effect levels relevant for environmental risk assessment (i.e., median prediction error of 1.3-fold for 10 and 20% lethal concentrations). The gBAM predicted the effect of pH more accurately than a previously published Cu BLM for juvenile rainbow trout, especially at pH > 8. We also evaluated the cross-species and cross-life stage applicability of the newly developed juvenile rainbow trout gBAM using existing chronic Cu toxicity data with early life stages of fathead minnow (Pimephales promelas) and rainbow trout. We did this because using a single bioavailability model for all fish species and life stages is practical from a regulatory point of view. Although the early life stage datasets exhibit considerable uncertainties, 91% of the considered toxicity values at the effect levels most relevant in European environmental regulations (10% effect on survival or growth) were predicted within a 2-fold error. Overall, the chronic Cu gBAM we developed is a valuable alternative for the existing chronic Cu BLM for rainbow trout and performs sufficiently well to be used in risk assessment according to currently accepted standards of bioavailability model performance (from the current European regulatory point of view). However, our analysis also suggests that bioavailability relations differ between different fish life stages and between endpoints (e.g., mortality vs growth), which is currently not accounted for in environmental risk assessments. Environ Toxicol Chem 2020;39:2424-2436. © 2020 SETAC.

Water quality assessments provide essential information for protecting aquatic habitats and stakeholders downstream of mining sites. Moreover, mining companies must comply with environmental quality standards and include public participation in water quality monitoring (WQM) practices. However, overarching challenges beyond corporate environmental responsibility are the scientific soundness, political relevance and harmonization of WQM practices. In this study, a mountainous watershed supporting large-scale gold mining in the headwaters, besides urban and agricultural landuses at lower altitudes, is assessed in the dry season. Conventional physicochemical and biological (Biological MonitoringWater Party-Colombia index) freshwater quality parameters were evaluated, including hydromorphological and land-use characteristics. According to the indicators used, water quality deterioration by mining was absent, in contrast to the e ects of urban economic activities, hydromorphological alterations and (less important) agricultural pollutants. We argue that mining impacts are hardly captured due to the limited ecological knowledge of high-mountain freshwaters, including uncharacterized mining-specific bioindicators, environmental baselines and groundwater processes, as well as ecotoxicological and microbial freshwater quality components. Lessons for overcoming scientific and operational challenges are drawn from joint e orts among governments, academia and green economy competitiveness. Facing a rapid development of extractive industries, interinstitutional and multidisciplinary collaborations are urgently needed to implement more integrated freshwater quality indicators of complex mining impacts. ispartof: Water vol:11 issue:9 pages:1-20 status: Published online

Over the past decade, significant advances have been made to unravel molecular mechanisms of stress response in different ecotoxicological model species. Within this study, we focus on population level transcriptomic responses of a natural population of Daphnia magna Straus, (1820), to heavy metals. We aim to characterize the population level transcriptomic responses, which include standing genetic variation, and improve our understanding on how populations respond to environmental stress at a molecular level. We studied population level responses to two heavy metals, copper and arsenic, and their binary mixture across time. Transcriptomic patterns identified significantly regulated gene families and genes at the population level including cuticle proteins and resilins. Furthermore, some of these differentially regulated gene families, such as cuticle proteins, were also significantly enriched for genetic variations including SNPs and MNPs. In general, genetic variation was observed in specific gene families, many of which are known to be involved in stress response. Overall, our results indicate that molecular stress responses can be identified within natural populations and that linking molecular mechanisms with genetic variation at the population level could contribute significantly to adverse outcome frameworks.

Although chemical risk assessment is still mainly conducted on a substance-by-substance basis, organisms in the environment are typically exposed to mixtures of substances. Risk assessment procedures should therefore be adapted to fit these situations. Four mixture risk assessment methodologies were compared for risk estimations of mixtures of copper (Cu), zinc (Zn), and nickel (Ni). The results showed that use of the log-normal species sensitivity distribution (SSD) instead of the best-fit distribution and sampling species sensitivities independently for each metal instead of using interspecies correlations in metal sensitivity had little impact on risk estimates. Across 4 different monitoring datasets, between 0% and 52% of the target water samples were estimated to be at risk, but only between 0% and 15% of the target water samples were at risk because of the mixture of metals and not any single metal individually. When a natural baseline database was examined, it was estimated that 10% of the target water samples were at risk because of single metals or their mixtures; when the most conservative method was used (concentration addition [CA] applied directly to the SSD, i.e., CASSD ). However, the issue of metal mixture risk at geochemical baseline concentrations became relatively small (2% of target water samples) when a theoretically more correct method was used (CA applied to individual dose response curves, i.e., CADRC ). Finally, across the 4 monitoring datasets, the following order of conservatism for the 4 methods was shown (from most to least conservative, with ranges of median margin of safety [MoS] relative to CASSD ): CASSD > CADRC (MoS = 1.17-1.25) > IADRC (independent action (IA) applied to individual dose-response curves; MoS = 1.38-1.60) > IASSD (MoS = 1.48-1.72). Therefore, it is suggested that these 4 methods can be used in a general tiered scheme for the risk assessment of metal mixtures in a regulatory context. In this scheme, the CASSD method could serve as a first (conservative) tier to identify situations with likely no potential risk at all, regardless of the method used (the sum toxic unit expressed relative to the 5% hazardous concentration [SumTUHC5 ] 0.05). The CADRC and IADRC methods could be used for site-specific assessment for situations that fall in between (SumTUHC5 > 1 and msPAFIA,SSD

To improve the ecological relevance of environmental risk assessment, an improved understanding is needed of 1) the influence of environmental conditions on the toxicity of pollutants, and 2) the effect of these factors in combination with possible interpopulation variability. The influences of salinity and dissolved organic carbon (DOC) on the accumulation and effect of copper (Cu) to settled mussels were investigated with mussels from a North Sea and a Baltic Sea population. We found that both populations were equally Cu-sensitive, even though the Baltic Sea population lives in suboptimal conditions. Baltic Sea mussels, however, accumulated more Cu. This suggests that these populations may have different ways of coping with excess Cu. The influence of salinity on Cu toxicity to settled mussels was limited for both populations. An increase in DOC did not decrease the Cu accumulation or effect in either population. This suggests that DOC-Cu complexes are bioavailable for settled mussels. These findings are in contrast with previous research which indicated that DOC decreased the toxicity and accumulation of Cu in the D-larvae life stage. As a consequence, the mussel larval stage is not the most Cu-sensitive life stage at high DOC concentrations. Furthermore, a DOC correction factor for Cu toxicity cannot be used for settled mussels. This should be accounted for in future marine Cu environmental risk assessment. Environ Toxicol Chem 2017;36:2074-2082. © 2017 SETAC.

Few studies have considered the effect of temperature on the chronic sensitivity of Daphnia magna to other stressors. The present study investigated the effect of temperature on chronic metal toxicity and whether this effect differed among 4 different D. magna clones. Life table experiments were performed with copper, zinc, and nickel at 15 °C, 20 °C, and 25 °C. General linear modeling indicated that chronic Cu, Zn, and Ni toxicity to D. magna were all significantly affected by temperature. When averaged across clones, our results suggest that chronic metal toxicity to D. magna was higher at 15 °C than at 20 °C, which is the temperature used in standard toxicity tests. At 15 °C, the 21-d median effect concentrations (EC50s) of Cu, Zn, and Ni were 1.4 times, 1.1 times, and 1.3 times lower than at 20 °C, respectively. At 25 °C, chronic Cu and Zn toxicity did not change in comparison with 20 °C, but chronic Ni toxicity was lower (21-d EC50 of nickel at 25 °C was 1.6 times higher than at 20 °C). The same trends were observed for Cu and Ni when the 21-d 10% and 20% effect concentrations were considered as the effect estimator, but not for Zn, which warns against extrapolating temperature effects on chemical toxicity across effect sizes. Overall, however, chronic metal toxicity was generally highest at the lowest temperature investigated (15 °C), which is in contrast with the usually observed higher acute metal toxicity at higher temperatures. Furthermore, the effect of temperature on chronic Ni toxicity depended significantly on the clone. This warns against extrapolating results about effect of temperature on chemical toxicity from single clone studies to the population level. Environ Toxicol Chem 2017;36:1909-1916. © 2016 SETAC.

Pollution and temperature increase are two of the most important stressors that aquatic organisms are facing. Exposure to elevated temperatures and metal contamination both induce heat shock proteins (HSPs), which may thus be involved in the induced cross-tolerance in various organisms. This study aimed to test the hypothesis that exposure to a non-lethal heat shock (NLHS) causes an increased tolerance to subsequent metal exposure. Using gnotobiotic cultures of the brine shrimp Artemia franciscana, the tolerance to Cd and Zn acute exposures was tested after a prior NLHS treatment (30min exposure to 37°C). The effects of NLHS and metal exposure were also assessed by measuring 70kDa-HSPs production, along with the analysis of epigenetic markers such as DNA methylation and histone H3 and histone H4 acetylation. Our results showed that heat-shocked Artemia had increased acute tolerance to Cd and Zn. However, different patterns of HSPs were observed between the two metal compounds and no epigenetic alterations were observed in response to heat shock or metal exposure. These results suggest that HSP production is a phenotypically plastic trait with a potential role in temperature-induced tolerance to metal exposure.

The regulatory decision-making process regarding chemical safety is most often informed by evidence based on ecotoxicity tests that consider growth, reproduction and survival as end-points, which can be quantitatively linked to short-term population outcomes. Changes in these end-points resulting from chemical exposure can cause alterations in micro-evolutionary forces (mutation, drift, selection and gene flow) that control the genetic composition of populations. With multi-generation exposures, anthropogenic contamination can lead to a population with an altered genetic composition, which may respond differently to future stressors. These evolutionary changes are rarely discussed in regulatory or risk assessment frameworks, but the growing body of literature that documents their existence suggests that these important population-level impacts should be considered. In this meta-analysis we have compared existing contamination levels of polychlorinated biphenyls (PCBs) and polycyclic aromatic hydrocarbons (PAHs) that have been documented to be associated with evolutionary changes in resident aquatic organisms to regulatory benchmarks for these contaminants. The original intent of this project was to perform a meta-analysis on evolutionary events associated with PCB and PAH contamination. However, this effort was hindered by a lack of consistency in congener selection for "total" PCB or PAH measurements. We expanded this manuscript to include a discussion of methods used to determine PCB and PAH total contamination in addition to comparing regulatory guidelines and contamination that has caused evolutionary effects. Micro-evolutionary responses often lead populations onto unique and unpredictable trajectories. Therefore, to better understand the risk of population-wide alterations occurring, we need to improve comparisons of chemical contamination between affected locations. In this manuscript we offer several possibilities to unify chemical comparisons for PCBs and PAHs that would improve comparability among evolutionary toxicology investigations, and with regulatory guidelines. In addition, we identify studies documenting evolutionary change in the presence of PCB and PAH contamination levels below applicable regulatory benchmarks.

The univariate effects of Ca, pH, and dissolved organic carbon (DOC) on chronic (48-h) Pb toxicity to the freshwater rotifer Brachionus calyciflorus were investigated. High pH (8.2) and higher concentrations of DOC were protective against filtered Pb toxicity, whereas Ca was not. However, expressed as the free Pb2+ ion, Pb2+ toxicity increased with increasing DOC concentration, indicating that Pb-fulvic acid (FA) complexes may be bioavailable and may contribute to toxicity. Two different bioavailability models were developed. The pH effect was modeled as a single-site competition effect by H+ (log KHBL = 7.14). In the first model, only this H+ effect was considered; in a second model, a log-linear fulvic acid bioavailability effect (SFA = 0.602) was also incorporated. Both models predicted chronic Pb toxicity for most waters used for model development within a 2-fold error. To validate the predictive capacities of the models, chronic Pb toxicity to B. calyciflorus was tested in 5 natural waters and a reference water. Both models consistently underestimated Pb toxicity in the natural waters, as a result of a shift in rotifer sensitivity to Pb between the development and the validation test series. However, optimizing the intrinsic sensitivity in the models specific for the validation test series resulted in reasonable predictions of Pb toxicity in the natural waters. The second, more complex model predicted chronic Pb toxicity most accurately. The protective effects of water chemistry on Pb toxicity to Brachionus are comparable to those observed for Ceriodaphnia. However, the developed Brachionus models were not able to accurately predict chronic Pb toxicity to the rotifer Philodina rapida. It remains unclear why the influence of water chemistry on chronic Pb toxicity appears to be different between 2 rotifer species. Environ Toxicol Chem 2016;35:2977-2986. © 2016 SETAC.

The authors investigated the effect of binary mixtures of zinc (Zn), copper (Cu), cadmium (Cd), and nickel (Ni) on the growth of a freshwater diatom, Navicula pelliculosa. A 7 × 7 full factorial experimental design (49 combinations in total) was used to test each binary metal mixture. A 3-d fluorescence microplate toxicity assay was used to test each combination. Mixture effects were predicted by concentration addition and independent action models based on a single-metal concentration-response relationship between the relative growth rate and the calculated free metal ion activity. Although the concentration addition model predicted the observed mixture toxicity significantly better than the independent action model for the Zn-Cu mixture, the independent action model predicted the observed mixture toxicity significantly better than the concentration addition model for the Cd-Zn, Cd-Ni, and Cd-Cu mixtures. For the Zn-Ni and Cu-Ni mixtures, it was unclear which of the 2 models was better. Statistical analysis concerning antagonistic/synergistic interactions showed that the concentration addition model is generally conservative (with the Zn-Ni mixture being the sole exception), indicating that the concentration addition model would be useful as a method for a conservative first-tier screening-level risk analysis of metal mixtures. Environ Toxicol Chem 2016;35:2765-2773. © 2016 SETAC.

A major problem with risk assessment of chemicals is the extrapolation of laboratory single-species toxicity tests, which oversimplify the actual field situation by ignoring species interactions, to natural communities. The authors tested if the bioavailability-normalized 5% hazardous concentration (HC5) estimated from chronic planktonic single-species toxicity data (HC5plankton ) for zinc (Zn) is protective for a plankton community and investigated the direct and indirect effects of Zn (at HC5 and HC50) on a freshwater community's structure and function. Microcosms were exposed to 3 different Zn concentrations (background, HC5plankton = 75 μg Zn/L and HC50plankton = 300 μg Zn/L) for 5 wk. The planktonic groups revealed a consistent no-observed-effect concentration for the community of 75 μg Zn/L, similar to or higher than the HC5plankton , thus suggesting its protectiveness in the present study. At 300 μg Zn/L a significant reduction in cladocerans resulted in increases of rotifer, ciliate, and phytoplankton abundance. In addition, the phytoplankton community shifted in dominance from grazing-resistant to edible species. Contrary to the species sensitivity distribution (SSD) prediction, which identified phytoplankton as the most sensitive group, only the total chlorophyll and the abundance of 2 phytoplankton species were adversely affected at 300 μg Zn/L. Thus, although the HC5 estimated from the bioavailability-normalized SSD was overall protective for the plankton community, the SSD was not able to correctly predict the species sensitivity ranking within their community context at the HC50. Environ Toxicol Chem 2016;35:2698-2712. © 2016 SETAC.

Metals occur as mixtures in the environment. Risk assessment procedures for metals currently lack a framework to incorporate chronic metal mixture toxicity. In the present study, the toxicity of binary and ternary mixture combinations of Ni, Zn, and Pb was investigated in 3 large-scale experiments using the standard chronic (7-d) Ceriodaphnia dubia reproductive toxicity test. These metals were selected because of anticipated differences in mode of action. The toxicity of the metals in most mixtures, expressed as either free metal ion activities or as dissolved metal concentrations, were antagonistic relative to the concentration addition model, whereas no significant (p < 0.05) interactive effects were observed relative to the independent action model. The only exception was the binary Pb-Zn mixture, for which mixture effects were noninteractive based on the dissolved concentrations, but antagonistic based on free ion activities all relative to the independent action model. Overall, the independent action model fitted the observed toxicity better than the concentration addition model, which is consistent with the different modes of action of these metals. The concentration addition model mostly overestimated toxicity. Finally, the present study warns against extrapolation of the type of interactive effects between species, even when they are closely related. Environ Toxicol Chem 2016;35:1796-1805. © 2015 SETAC.

The degree of biological variability within a population is an important factor for its ecological success. Yet, individual-based population models (IBMs) that utilize the Dynamic Energy Budget (DEB) theory as a mechanistic basis to simulate an individual's life history, largely rely on rule-of-thumb estimates of inter-individual variability of their parameter values. In this study, we explored how data from previous life-history experiments with the copepod Nitocra spinipes could be used to make realistic estimates of variability in DEB parameter values for this species. We specifically investigated to which one of 12 possible parameters variability should be added, and what shape and scale its probability distribution should have, to most accurately reproduce the experimental data's variation with model simulations. The best result was achieved by drawing the parameter ‘surface-area-specific maximum assimilation rate’ from a log-normal distribution with a standard deviation of 0.15 or by drawing the maturity thresholds for birth and puberty (assuming proportionality between the two) from a log-normal distribution with a standard deviation of 0.40. In both cases, the model not only accurately reproduced the variation observed in development time data (which had been used in the fitting process) but also gave adequate predictions of the variation observed in brood size data (which had not been used for fitting). Although the findings of this study apply to one species only, the methods presented here are a proof of principle for how the question of intra-specific variability in a DEB model, for any species, can be addressed in a simple, yet purposeful manner.

Marine zooplankton are increasingly being affected by recent environmental changes, such as climate change, and respond with profound spatial relocations and shifts in phenology and physiology. In order to predict whether populations are able to persist or adapt to such new conditions, it is essential to understand the molecular basis of such adaptations, which ultimately get translated into these physiological responses. To explore variation in population gene expression across time and space, we investigated transcriptome-level profiles of the calanoid copepod Temora longicornis, that were collected at four different locations in the Belgian Part of the North Sea (BPNS) on three different time points (April, June, October) in 2018. RNA-seq analysis of field collected adults identified large seasonal differences in gene expression, mainly between spring-summer and autumn samples. The largest log-fold changes occurred in a set of genes encoding for ribosomal and myosin (heavy chain) transcripts. Enrichment analysis revealed a strong seasonal pattern in vitellogenin, cuticle and glycolytic gene expression as well. We also found a positive correlation between vitellogenin expression and densities of T. longicornis. No clear spatial variation in expression patterns was found in the BPNS. This study underlines the potential of field gene expression studies for biomonitoring purposes and the significance of considering seasonal variation in future studies.

It is well known that temperature can affect the ecotoxicity of chemicals (including metals) to aquatic organisms. It was recently reported that nickel (Ni), a priority substance under the European Water Framework directive, showed decreasing chronic toxicity to Daphnia magna with increasing temperature, between 15 and 25 degrees C. We performed a toxicokinetic study to contribute to an increased mechanistic understanding of this effect. More specifically, we investigated the effect of temperature on Ni uptake and elimination in D. magna (in 4 clones) using an experimental design that included Ni exposures with different stable isotopic composition and using a one-compartment model for data analysis. Both Ni uptake and elimination were affected by temperature, and some clear interclonal differences were observed. On average (across all clones), however, a similar pattern of the effect of temperature was observed on both Ni uptake and elimination, that is, the uptake rate constant (k(u)) and elimination rate constant (k(e)) during 72 h of Ni exposure were lower at 25 than at 19 degrees C, by 2.6-fold and 1.6-fold, respectively, and they were similar at 19 and 15 degrees C. This pattern does not correspond to the effects of temperature on chronic Ni toxicity reported previously, suggesting that Ni compartmentalization and/or toxicodynamics may also be affected by temperature. The data gathered with our specific experimental design also allowed us to infer that 1) the k(u) was up-regulated over time, that is, the k(u) after 2 d of Ni exposure was significantly higher than the initial k(u), by 1.5- to 2.3-fold, and 2) the k(e) decreased significantly when the external Ni exposure was stopped, by 1.2- to 1.9-fold. These 2 findings are in contrast with 2 commonly used assumptions in toxicokinetic models, that is, that k(u) is constant during exposure and k(e) is independent of external exposure. We suggest that future toxicokinetic studies consider these factors in their experimental designs and data analyses. Overall, our study contributes to the growing body of evidence that temperature affects toxicokinetics of metals (and chemicals in general), but at the same time we emphasize that knowledge of toxicokinetics alone is not necessarily sufficient to explain or predict temperature effects on (chronic) toxicity. Environ Toxicol Chem 2019;38:784-793. (c) 2019 SETAC

Polyunsaturated fatty acids (PUFAs) have key biological roles in fish cells. We recently showed that the phospholipid composition of rainbow trout liver cells (RTL-W1 cell line) modulates their tolerance to an acute cadmium (Cd) challenge. Here, we investigated (i) the extent to which PUFAs and Cd impact fatty acid homeostasis and metabolism in these cells and (ii) possible mechanisms by which specific PUFAs may confer cytoprotection against Cd. First, RTL-W1 cells were cultivated for one week in growth media spiked with 50 μmol L −1 of either alpha-linolenic acid (ALA, 18:3n-3), eicosapentaenoic acid (EPA, 20:5n-3), linoleic acid (LA, 18:2n-6) or arachidonic acid (AA, 20:4n-6) in order to modulate their fatty acid profile. Then, the cells were challenged with Cd (0, 50 or 100 μmol L −1) for 24 h prior to assaying viability, fatty acid profile, intracellular Cd content, proteomic landscape and expression levels of genes involved in fatty acid metabolism, synthesis of PUFA-derived signalling molecules and stress response. We observed that the fatty acid supply and, to a lesser extent, the exposure to Cd influenced cellular fatty acid homeostasis and metabolism. The cellular fatty acid composition of fish liver cells modulated their tolerance to an acute Cd challenge. Enrichments in ALA, EPA, and, to a lesser extent, AA conferred cytoprotection while enrichment in LA had no impact on cell viability. The present study ruled out the possibility that cytoprotection reflects a decreased Cd burden. Our results rather suggest that the PUFA-derived cytoprotection against Cd occurs through a reduction of the oxidative stress induced by Cd and a differential induction of the eicosanoid cascade, with a possible role of peroxiredoxin and glutaredoxin (antioxidant enzymes) as well as cytosolic phospholipase A2 (enzyme initiating the eicosanoid cascade).

To estimate mixture effects caused by the high number of chemicals simultaneously present in the environment, methods for routine effect assessment of environmentally realistic contaminant mixtures are needed. We repeatedly exposed the marine diatom Phaeodactylum tricornutum to SpeediskTM passive sampler extracts and observed statistically significant growth stimulation up to 6 and 7% for samples from inside and outside the harbor of Zeebrugge, respectively. These effects were found at summed contaminant concentrations (159-166 ng L-1 ) that were within a 1.1- to 2.4-fold range of those observed in grab water samples taken during sampler deployment. These stimulatory effects were confirmed in 2 independent tests with extracts stored for

Recent studies have shown that temperature affects chronic nickel (Ni) toxicity to Daphnia magna at the individual (apical) level. However, the effect of temperature on Ni toxicity to D. magna at the population level is unknown. The present study investigated whether the effect of temperature on chronic Ni toxicity to D. magna assessed on apical endpoints can be extrapolated to the population level. The results of the population experiment showed no consistent Ni effects on total D. magna population abundance at 15, 20, and 25 degrees C, although the Ni concentrations tested were previously reported to significantly reduce reproduction in D. magna individuals. This result supports the idea that ecological risk assessment should not extrapolate as such from apical endpoints to the population level. A dynamic energy budget individual-based model (DEB-IBM) was calibrated using apical Ni toxicity data at 15, 20, and 25 degrees C. The goal was to investigate whether the calibrated DEB-IBM would be able to predict the unexpected absence of effects at the population level and to further investigate the effect of temperature on Ni toxicity to a D. magna population. At the population level, the calibrated DEB-IBM correctly predicted the unexpected absence of an effect of Ni on a D. magna population. Detailed analysis of simulation output suggests that the predicted lower Ni sensitivity at the population level occurs because Ni-induced mortality is compensated by reduced starvation (less intraspecific competition). Extrapolated median effective concentration (EC50) values for population density predicted that the effect of temperature on Ni toxicity to D. magna populations was smaller (1.9-fold higher at 25 degrees C than at 15 degrees C) than on Ni toxicity to D. magna apical reproduction (the EC50 is 6.5-fold higher at 25 degrees C than at 15 degrees C). These results show that the DEB-IBM can help to replace population experiments by in silico simulations and to optimize the experimental design of population studies. Environ Toxicol Chem 2019;38:1423-1433. (c) 2019 SETAC

The Belgian part of the North Sea (BPNS) is subjected to multiple environmental stressors. The impact of these stressors includes the modulation of fatty acid (FA) composition of the zooplankton. This study recorded temporal and spatial patterns of the FA profiles of two dominant calanoid copepods within the BPNS: Temora longicornis (Muller, 1785) and Acartia clausi (Giesbrecht, 1889). By means of distance-based linear modelling and by applying multi model inference to generalized additive models, environmental stressors were linked to patterns of the FA profiles of these species. The FA profiles of A. clausi and T. longicornis showed distinct intraspecific, spatial and temporal differences within the BPNS. Temperature and algal food quality (marked by the ratio of silicate concentration to dissolved inorganic nitrogen concentration, SiO4/DIN) were the most important drivers of seasonal fluctuations in the DHA/EPA ratio of both species. DHA/EPA ratio can be used as marker for stress in copepods in the BPNS in order to have a quick indication of food quality changes at the basis of the food web.