Role of Biofilms in Contaminant Bioaccumulation and Trophic Transfer in Aquatic Ecosystems: Current State of Knowledge and Future Challenges.

In freshwater environments, microbial assemblages attached to submerged substrates play an essential role in ecosystem processes such as primary production, supported by periphyton, or organic matter decomposition, supported by microbial communities attached to leaf litter or sediments. These microbial assemblages, also called biofilms, are not only involved in nutrients fluxes but also in contaminants dynamics. Biofilms can accumulate metals and organic contaminants transported by the water flow and/or adsorbed onto substrates. Furthermore, due to their high metabolic activity and their role in aquatic food webs, microbial biofilms are also likely to influence contaminant fate in aquatic ecosystems. In this review, we provide (1) a critical overview of the analytical methods currently in use for detecting and quantifying metals and organic micropollutants in microbial biofilms attached to benthic substrata (rocks, sediments, leaf litter); (2) a review of the distribution of those contaminants within aquatic biofilms and the role of these benthic microbial communities in contaminant fate; (3) a set of future challenges concerning the role of biofilms in contaminant accumulation and trophic transfers in the aquatic food web. This literature review highlighted that most knowledge on the interaction between biofilm and contaminants is focused on contaminants dynamics in periphyton while technical limitations are still preventing a thorough estimation of contaminants accumulation in biofilms attached to leaf litter or sediments. In addition, microbial biofilms represent an important food resource in freshwater ecosystems, yet their role in dietary contaminant exposure has been neglected for a long time, and the importance of biofilms in trophic transfer of contaminants is still understudied.

Contribution of aqueous and dietary uptakes to lead (Pb) bioaccumulation in Gammarus pulex: From multipathway modeling to in situ validation.

Although dynamic approaches are nowadays used increasingly to describe metal bioaccumulation in aquatic organisms, the validation of such laboratory-derived modeling is rarely assessed under environmental conditions. Furthermore, information on bioaccumulation kinetics of Pb and the significance of its uptake by dietary route is scarce in freshwater species. This study aims at modeling aqueous and dietary uptakes of Pb in the litter-degrader Gammarus pulex and assessing the predictive quality of multipathway modeling from in situ bioaccumulation data. In microcosms, G. pulex were exposed to environmentally realistic concentrations of Pb (from 0.1 to 10µg/L) in the presence of Pb-contaminated poplar leaves, which were enclosed or not in a net to distinguish aqueous and dietary uptakes. Results show that water and food both constitute contamination sources for gammarids. Establishing biodynamic parameters involved in Pb aqueous and dietary uptake and elimination rates enabled to construct a multipathway model to describe Pb bioaccumulation in gammarids. This laboratory-derived model successfully predicted bioaccumulation measured in native populations of G. pulex collected in situ when local litter was used as dietary exposure source. This study demonstrates not only the suitable applicability of biodynamic parameters for predicting Pb bioaccumulation but also the necessity of taking dietary uptake into account for a better interpretation of the gammarids' contamination in natural conditions.

Screening potential toxicity of currently used herbicides in the freshwater amphipod Gammarus fossarum based on multi-level biomarker responses to field-realistic exposures.

Herbicides are widely used to control weeds and maximize crop growth. Because of agricultural runoff, these chemicals are potentially hazardous to aquatic wildlife. However, their ecotoxicity and resulting disturbance in individual performance remain scarcely documented in freshwater crustaceans. This study aimed to screen the potential toxicity of currently used herbicides in the ecosystem engineer Gammarus fossarum using multi-level biomarkers. In microcosms, gammarids were exposed for 72 h to 12 herbicides individually (quinmerac, mesotrione, bentazone, isoproturon, chlortoluron, metazachlor, chloridazone, diflufenican, flufenacet, aclonifen, prosulfocarb and metolachlor) at a field-realistic concentration (i.e. 10 µg/L). The sublethal effects were assessed by monitoring several biochemical, physiological and behavioural traits. In exposed gammarids, alterations in behavioural activities were observed, i.e. increased locomotion and respiration as a general trend. Moreover, biochemical biomarkers suggested herbicide-dependent disruptions in moulting, antioxidant responses and cell integrity. Integrating multi-metric variations through statistical analyses allowed us to identify herbicide clusters likely to trigger common sets of biological responses. Depressed antioxidant defence at the cell level and impaired respiration at the individual level were the predominant toxic effects of herbicides, related to their hydrophobic feature. Furthermore, establishing relationships between sublethal alterations in gammarids and acute lethality or chronic toxicity values defined for regulatory purposes supports the relevance of these alterations as early warnings of toxicity. Our findings demonstrate that currently used herbicides have unexpected toxicological effects in a non-target wild animal, with possible long-term alterations in population dynamics and associated ecological functions, which constitute promising diagnostic tools for risk assessment in agricultural areas.

Modelling copper bioaccumulation in Gammarus pulex and alterations of digestive metabolism.

Bioaccumulation enables to integrate the ability of aquatic organisms to regulate metals and effects of water chemistry on metal bioavailability. Linking this process to biological responses offers thus promising lines of enquiry for protecting aquatic ecosystems. This study aims at characterizing the mechanisms involved in waterborne Cu bioaccumulation and assessing metal impact on digestive metabolism in an ecosystem engineer widely distributed in Europe, Gammarus pulex. The organism was exposed to several Cu concentrations (from 0.5 to 100 µg/L) in aquatic microcosms to establish kinetic parameters for the construction and comparison of two bioaccumulation models, i.e. the biodynamic and saturation models. Cu uptake was recorded in waters exhibiting various concentrations of Na, Mg and Ca at environmental levels to assess the influence of cationic composition on bioaccumulation. Then, the effect of increasing Cu in exposure media on the digestive metabolism of G. pulex was investigated by measuring enzymatic activities (ß-glucosidase, N-acetyl-ß-glucosaminidase, ß-galactosidase). We showed that the saturation model is more suitable than the biodynamic model to describe Cu bioaccumulation in gammarids due to a maximal capacity of animals to accumulate the metal. Cationic composition of water affected insignificantly Cu uptake. All activities of tested enzymes decreased with increasing Cu in exposure media but with different degrees. High correlations were established between the inhibition of enzymatic activities and amounts of Cu bioaccumulated by gammarids. These biological responses could thus provide early-warming of Cu impact on aquatic biota.

Individual performances and biochemical pathways as altered by field-realistic exposures of current-use fungicides and their mixtures in a non-target species, Gammarus fossarum.

Persistent fungicides, which are widely applied to agricultural soils to protect crops, are frequently detected in freshwaters because of hydraulic transfer, possibly resulting in unintentional adverse effects on wildlife. However, the ecotoxicity of fungicides in aquatic species remains scarcely assessed at environmentally relevant concentrations, and there is scant information available on their combined effects. This study aims at investigating multi-level changes elicited by two currently used fungicides, boscalid and tebuconazole, in the amphipod Gammarus fossarum. In microcosms, gammarids were exposed for 72 h to fungicides tested individually or in binary mixture at 0.01, 0.1 and 1.0 µg/L to monitor individual performances (locomotion, respiration and amplexus formation) and biochemical parameters (involved in energy metabolism, growth, moulting and cell stress). This range of exposure concentrations was field-realistic and largely lower than local environmental quality standards for the protection of aquatic wildlife. Overall, results showed that single and combined exposures altered the mobility and respiratory activity of individuals. At the cellular level, boscalid inhibited energy-based biomarkers whereas tebuconazole led to cytotoxicity associated with reduced antioxidant defences. In binary mixtures, the biochemical responses were mainly driven by the presence of boscalid. Multi-variable analyses, integrating individual alterations and cellular impairments alike, confirmed the relevance of the multi-level approach in forecasting the toxicity of fungicides and their mixtures towards other aquatic species. This study demonstrates dose-related sublethal effects of fungicides on multiple functional traits in an ecosystem engineer under realistic exposure scenarios, highlighting the harmful signs of these toxicants. Such sublethal alterations could thus constitute reliable tools for the early diagnosis of the organisms' health and the ecological status of agriculturally impacted hydrosystems.

High denitrification potential but low nitrous oxide emission in a constructed wetland treating nitrate-polluted agricultural run-off.

Constructed wetlands (CW) can efficiently remove nitrogen from polluted agricultural run-off, however, a potential caveat is nitrous oxide (N2O), a harmful greenhouse gas and stratospheric ozone depleter. During five sampling campaigns, we measured N2O fluxes from a 0.53 ha off-stream CW treating nitrate-rich water from the intensively fertilized watershed in Rampillon, France, using automated chambers with a quantum cascade laser system, and manual chambers. Sediment samples were analysed for potential N2 flux using the HeO2 incubation method. Both inlet nitrate (NO3-) concentrations and N2O emission varied significantly between the seasons. In the Autumn and Winter inlet concentrations were about 11 mg NO3--N L-1, and < 6.5 mg NO3--N L-1 in the Spring and Summer. N2O emission was highest in the Autumn (mean ± standard error: 9.7 ± 0.2 µg N m-2 h-1) and lowest in the Summer (wet period: 0.2 ± 0.3 µg N m-2 h-1). The CW was a very weak source of N2O emitting 0.32 kg N2O-N ha-1 yr-1 and removing around 938 kg NO3--N ha-1 yr-1, the ratio of N2O-N emitted to NO3--N removed was 0.033%. The automated and manual chambers gave similar results. From the potential N2O formation in the sediment, only 9% was emitted to the atmosphere, the average N2 N 2O ratio was high: 89:1 for N2-Npotential: N2O-Npotential and 1353:1 for N2-Npotential: N2O-Nemitted. These results indicate complete denitrification. The focused principal component analysis showed strong positive correlation between the gaseous N2O fluxes and the following environmental factors: NO3--N concentrations in inlet water, streamflow, and nitrate reduction rate. Water temperature, TOC and DOC in the water and hydraulic residence time showed negative correlations with N2O emissions. Shallow off-stream CWs such as Rampillon may have good nitrate removal capacity with low N2O emissions.

Insights into the development of fungal biomarkers for metal ecotoxicity assessment: case of Trametes versicolor exposed to copper.

The relationship between the physiological state of fungi and the response of their functional system to metals is not known, limiting the use of fungal enzymes as tools for assessing metal ecotoxicity in terrestrial ecosystems. The present study attempts to establish how the development phases modulate the secretion of enzymes in the filamentous fungus Trametes versicolor after exposure to Cu. For that purpose, extracellular hydrolases (acid and alkaline phosphatases, aryl-sulfatase, beta-glucosidase, beta-galactosidase, and N-acetyl-beta-glucosaminidase) and oxidoreductases (laccase, manganese and lignin peroxidases) were monitored in liquid cultures for 2 weeks. Copper was added during either the growth or the stationary phases at 20 or 200 ppm. Results of the present study showed that Cu at the highest concentration modifies the secretion of enzymes, regardless of the development phase to which the fungus was exposed. However, the sensitivity of enzyme responses to Cu depended on the phase development and the type of secreted enzyme. In a general way, the production of hydrolases was decreased by Cu, whereas that of oxidoreductases was highly increased. Furthermore, lignin peroxidase was not detected in control cultures and was specifically produced in the presence of Cu. In conclusion, fungal oxidoreductases may be enzymatic biomarkers of copper exposure for ecotoxicity assessment.

Single and combined effects of insecticides on multi-level biomarkers in the non-target amphipod Gammarus fossarum exposed to environmentally realistic levels.

Aquatic media are ultimate recipients of various contaminants including pesticides pervasively applied in agrosystems. Characterizing the ecotoxicity of pesticides and their mixtures to aquatic wildlife at field-realistic levels is thus crucial for environmental risk assessment. This study aims at assessing the effects of two current-use insecticides, imidacloprid and chlorpyrifos, on Gammarus fossarum using multi-level biomarkers. In microcosms, gammarids were exposed for 72 h to insecticides tested individually or in mixture at 0.01, 0.1 and 1 µg/L of each chemical. Multi-metric responses were assessed at the individual level (behavioural traits: locomotion, respiration and amplexus formation) and the cellular level (enzymes involved in growth, moulting, digestion and cell stress). The results showed insecticide-elicited behavioural and biochemical responses from the lowest concentration of 0.01 µg/L. Overall, single exposures stimulated behavioural traits and inhibited enzymatic activities, highlighting subtle impacts at different organizational levels but these were not dose related. For binary mixtures, antagonistic effects (i.e. less-than-additive) on biomarkers were mainly observed when compared with single exposures. Multi-variable analyses indicated the complementarity of behavioural and biochemical biomarkers in identifying sublethal biological alterations and dose-dependent multiple action sites of insecticides. Besides, the mortality observed only for the mixture at 1 µg/L demonstrated a high lethal potential of insecticides in a simple binary combination. To conclude, this study demonstrates disturbances in individual performances and cellular impairments occurring at environmentally realistic exposure levels in a non-target wild species. Since the sublethal effects, such as those identified with this multi-biomarker approach, could lead to long-term alterations in population dynamics of agricultural areas, they constitute promising early endpoints for risk assessment of insecticides.

Application of a multidisciplinary and integrative weight-of-evidence approach to a 1-year monitoring survey of the Seine River.

Quality assessment of environments under high anthropogenic pressures such as the Seine Basin, subjected to complex and chronic inputs, can only be based on combined chemical and biological analyses. The present study integrates and summarizes a multidisciplinary dataset acquired throughout a 1-year monitoring survey conducted at three workshop sites along the Seine River (PIREN-Seine program), upstream and downstream of the Paris conurbation, during four seasonal campaigns using a weight-of-evidence approach. Sediment and water column chemical analyses, bioaccumulation levels and biomarker responses in caged gammarids, and laboratory (eco)toxicity bioassays were integrated into four lines of evidence (LOEs). Results from each LOE clearly reflected an anthropogenic gradient, with contamination levels and biological effects increasing from upstream to downstream of Paris, in good agreement with the variations in the structure and composition of bacterial communities from the water column. Based on annual average data, the global hazard was summarized as "moderate" at the upstream station and as "major" at the two downstream ones. Seasonal variability was also highlighted; the winter campaign was least impacted. The model was notably improved using previously established reference and threshold values from national-scale studies. It undoubtedly represents a powerful practical tool to facilitate the decision-making processes of environment managers within the framework of an environmental risk assessment strategy.

Behavioural and biochemical responses to metals tested alone or in mixture (Cd-Cu-Ni-Pb-Zn) in Gammarus fossarum: From a multi-biomarker approach to modelling metal mixture toxicity.

Metals are usually present as mixtures at low concentrations in aquatic ecosystems. However, the toxicity and sub-lethal effects of metal mixtures on organisms are still poorly addressed in environmental risk assessment. Here we investigated the biochemical and behavioural responses of Gammarus fossarum to Cu, Cd, Ni, Pb and Zn tested individually or in mixture (M2X) at concentrations twice the levels of environmental quality standards (EQSs) from the European Water Framework Directive. The same metal mixture was also tested with concentrations equivalent to EQSs (M1X), thus in a regulatory context, as EQSs are proposed to protect aquatic biota. For each exposure condition, mortality, locomotion, respiration and enzymatic activities involved in digestive metabolism and moult were monitored over a 120h exposure period. Multi-metric variations were summarized by the integrated biomarker response index (IBR). Mono-metallic exposures shed light on biological alterations occurring at environmental exposure levels in gammarids and depending on the considered metal and gender. As regards mixtures, biomarkers were altered for both M2X and M1X. However, no additive or synergistic effect of metals was observed comparing to mono-metallic exposures. Indeed, bioaccumulation data highlighted competitive interactions between metals in M2X, decreasing subsequently their internalisation and toxicity. IBR values indicated that the health of gammarids was more impacted by M1X than M2X, because of reduced competitions and enhanced uptakes of metals for the mixture at lower, EQS-like concentrations. Models using bioconcentration data obtained from mono-metallic exposures generated successful predictions of global toxicity both for M1X and M2X. We conclude that sub-lethal effects of mixtures identified by the multi-biomarker approach can lead to disturbances in population dynamics of gammarids. Although IBR-based models offer promising lines of enquiry to predict metal mixture toxicity, further studies are needed to confirm their predictive quality on larger ranges of metallic combinations before their use in field conditions.

Oxidoreductases provide a more generic response to metallic stressors (Cu and Cd) than hydrolases in soil fungi: new ecotoxicological insights.

The present study investigates the effect of metals on the secretion of enzymes from 12 fungal strains maintained in liquid cultures. Hydrolases (acid phosphatase, ß-glucosidase, ß-galactosidase, and N-acetyl-ß-glucosaminidase) and ligninolytic oxidoreductases (laccase, Mn, and lignin peroxidases) activities, as well as biomass production, were measured in culture fluids from fungi exposed to Cu or Cd. Our results showed that all fungi secreted most of the selected hydrolases and that about 50% of them produced a partial oxidative system in the absence of metals. Then, exposure of fungi to metals led to the decrease in biomass production. At the enzymatic level, Cu and Cd modified the secretion profiles of soil fungi. The response of hydrolases to metals was contrasted and complex and depended on metal, enzyme, and fungal strain considered. By contrast, the metals always stimulated the activity of ligninolytic oxidoreductases in fungal strains. In some of them, oxidoreductases were specifically produced following metal exposure. Fungal oxidoreductases provide a more generic response than hydrolases, constituting thus a physiological basis for their use as biomarkers of metal exposure in soils.

Seasonal variability and inter-species comparison of metal bioaccumulation in caged gammarids under urban diffuse contamination gradient: implications for biomonitoring investigations.

Although caging of Gammarus species offers promising lines of inquiry to monitor metal bioavailability in freshwaters, the interspecies responsiveness to metal exposures is still unclear. In addition, abiotic factors inherent to transplantation can hamper the interpretation of field bioaccumulation data. To assess the relevance of using gammarids as biomonitors, we investigated the seasonal influence on metal bioaccumulation in two common species, Gammarus pulex and Gammarus fossarum. During four seasons, caged gammarids were deployed on three sites along the Seine River exhibiting a diffuse gradient of multi-metal contamination: a site upstream and two sites downstream from the Paris megacity. For each seasonal deployment, metal concentrations in animals were determined after 7d-exposure in situ (Ag, Cd, Co, Cu, Mn, Ni, Pb and Zn). Results show that the seasonal patterns of metal contaminations are similar between both Gammarus species, and closely related to the river axis' contamination gradient. Statistical analyses indicate that bioaccumulation of essential metals in both species is influenced by season, especially by water temperature. This highlights the necessity to consider this climatic factor inherent to the deployment period for a reliable interpretation of bioaccumulation data in the field. The comparison of accumulation factors suggests that these two species coming from different geochemical origins display similar abilities to internalize metals. This generic responsiveness of caged gammarids supports their use as sentinel organisms to quantify low spatiotemporal variations in metal bioavailabilities.

Essential metal contents in indigenous gammarids related to exposure levels at the river basin scale: metal-dependent models of bioaccumulation and geochemical correlations.

Biomonitoring, assumed to be an integrative measurement of the chemical exposure of aquatic organisms, is not straightforward for essential metals because they can be actively regulated by animals. Although increasing bioaccumulation with exposure levels is a crucial endpoint for the development of biomonitors, it is rarely verified in real environments, where the metal concentrations are rather low and vary little. This study was designed at the scale of a river basin to assess the ability of Gammarus pulex indigenous populations to accumulate Cu, Zn and Mn in realistic exposure conditions. During two annual campaigns, water and gammarids were collected at various sites contrasted in terms of physicochemistry and contamination. The results show significant relationships between metal concentrations in animals and in freshwaters established by conceptual models of bioaccumulation, but with patterns specific to each metal (base level, internal regulation and maximal accumulation). In particular, a saturation process of Cu accumulation occurs at environmental exposure levels, unlike Mn and Zn. Statistical analyses performed from field data show that Cu and Zn bioaccumulations may be influenced by a complex combination of geochemical variables, unlike Mn. We conclude that G. pulex is a useful candidate to monitor metal bioavailability in freshwaters due to its responsiveness to low exposures of surrounding environments. Nevertheless, a reliable quantification of bioavailability of essential metals requires characterizing some geochemical effects on metal bioaccumulation.

Bioaccumulation of polybrominated diphenyl ethers (PBDEs) in Gammarus pulex: relative importance of different exposure routes and multipathway modeling.

Characterizing the exposure routes of an organism and its ability to regulate accumulated contaminants is a crucial step toward developing a biomonitor. To date, very little data are available on the bioaccumulation kinetics of PBDEs in freshwater biota. This study aims at investigating the potential use of a litter-degrader widely distributed in European freshwaters, Gammarus pulex, as an indicator of exposure to PBDEs. In aquatic microcosms, gammarids were exposed to a mixture of brominated congeners (BDE-28, 47, 66, 85, 99, 100, 153, 154 and 183) to assess their ability to bioconcentrate PBDEs. Results show that all tested congeners are highly internalized by G. pulex and uptake rates of PBDEs are closely related to their partition coefficients (Kow). The determination of the elimination rate of BDE-47, the congener most readily accumulated by gammarids, indicated that metabolism and excretion of this congener are low in G. pulex, which argues in favor of its use as a quantitative biomonitor. Finally, bioaccumulation experiments were performed using contaminated leaves to determine the relative importance of dietary uptake in the contamination of gammarids. Even though water is the preeminent exposure route, a significant uptake of BDE-47 through food was observed (27%). We propose a biodynamic model that takes into account both exposure routes to describe BDE-47 bioaccumulation. This study supports the use of this ubiquitous amphipod as an early warning monitor of the bioavailable contamination of freshwaters by PBDEs.

Favouring the bioavailability of Zn and Cu to enhance the production of lignin-modifying enzymes in Trametes versicolor cultures.

Metal effect on the enzyme secretion in fungi is usually related to total concentrations but not to bioavailable metal species. In this work, we aimed at enhancing the secretion of lignin-modifying oxidoreductases in Trametes versicolor by favouring the bioavailability of essential metals. For this purpose, the fungus was exposed to Cu or Zn in liquid culture media exhibiting different complexation levels. Metal speciation was determined experimentally or theoretically to quantify free metal species, supposed to be the most bioavailable, and species complexed to ligands. Although Zn(2+) contents were high in media, Zn had no effect on the oxidoreductase production. Conversely, Cu highly induced the manganese peroxidase and laccase productions until 40 and 310 times when compared to unexposed controls. This inductive potential was highly correlated to Cu(2+) contents in media. Furthermore, in poorly complexing media, the response threshold of oxidoreductases to Cu greatly decreased and an unexpected production of lignin peroxidase occurred.

Secretion profiles of fungi as potential tools for metal ecotoxicity assessment: a study of enzymatic system in Trametes versicolor.

The relationship between the expression of extracellular enzymatic system and a metal stress is scarce in fungi, hence limiting the possible use of secretion profiles as tools for metal ecotoxicity assessment. In the present study, we investigated the effect of Zn, Cu, Pb and Cd, tested alone or in equimolar cocktail, on the secretion profiles at enzymatic and protein levels in Trametesversicolor. For that purpose, extracellular hydrolases (acid phosphatase, ß-glucosidase, ß-galactosidase and N-acetyl-ß-glucosaminidase) and ligninolytic oxidases (laccase, Mn-peroxidase) were monitored in liquid cultures. Fungal secretome was analyzed by electrophoresis and laccase secretion was characterized by western-blot and mass spectrometry analyses. Our results showed that all hydrolase activities were inhibited by the metals tested alone or in cocktail, whereas oxidase activities were specifically stimulated by Cu, Cd and metal cocktail. At protein level, metal exposure modified the electrophoretic profiles of fungal secretome and affected the diversity of secreted proteins. Two laccase isoenzymes, LacA and LacB, identified by mass spectrometry were differentially glycosylated according to the metal exposure. The amount of secreted LacA and LacB was strongly correlated with the stimulation of laccase activity by Cu, Cd and metal cocktail. These modifications of extracellular enzymatic system suggest that fungal oxidases could be used as biomarkers of metal exposure.

Waterborne nickel bioaccumulation in Gammarus pulex: comparison of mechanistic models and influence of water cationic composition.

The biodynamic and saturation models offer promising lines of enquiry to predict the bioaccumulation of metals by aquatic organisms. However, in order to construct these models, the accumulation strategies have to be defined for each metal/organism couple in controlled conditions. This study aims at modelling the waterborne bioaccumulation of Ni and the influence of the water's geochemical properties on this process in a crustacean that is widely distributed in Europe, Gammarus pulex. In the laboratory, G. pulex was exposed to several Ni concentrations (from 0.001 to 100 mg L(-1)) in aquatic microcosms. Our results show that G. pulex is very tolerant to Ni (LC50(48 h)=477 mg L(-1) Ni). Time course experiments enabled the construction of a biodynamic model by determining the uptake (k(u)) and elimination (k(e)) rate constants. When the exposure concentration exceeded 1 mg L(-1) Ni, the metal uptake reached a maximum due to a limited number of binding sites for Ni. Therefore, the organism's maximal capacity to accumulate the metal (B(max)) and the half-saturation constant (K) were determined to establish the saturation model. We showed that the two models are comparable for the lowest exposure concentrations (<1 mg L(-1) Ni), with k(u)/k(e)=B(max)/K. Then, the bioaccumulation of Ni was recorded in waters exhibiting various concentrations of three major ions (Na(+), Mg(2+) and Ca(2+)). Only Ca had an inhibitory effect on the Ni uptake. This study reports for the first time the bioaccumulation of Ni in G. pulex. Because of its high tolerance to Ni and its high capacity to accumulate this metal, this crustacean could be used as an indicator of Ni bioavailability in freshwaters.