A two years field experiment to assess the impact of two fungicides on earthworm communities and their recovery.

Recent EFSA (European Food Safety Authority) reports highlighted that the ecological risk assessment of pesticides needed to go further by taking more into account the impacts of chemicals on biodiversity under field conditions. We assessed the effects of two commercial formulations of fungicides separately and in mixture, i.e., Cuprafor Micro® (containing 500 g kg-1 copper oxychloride) at 4 (C1, corresponding to 3.1 mg kg-1 dry soil of copper) and 40 kg ha-1 (C10), and Swing® Gold (50 g L-1 epoxiconazole EPX and 133 g L-1 dimoxystrobin DMX) at one (D1, 5.81 10-2 and 1.55 10-1 mg kg-1 dry soil of EPX and DMX, respectively) and ten times (D10) the recommended field rate, on earthworms at 1, 6, 12, 18 and 24 months after the application following the international ISO standard no. 11268-3 to determine the effects on earthworms in field situations. The D10 treatment significantly reduced the species diversity (Shannon diversity index, 54% of the control), anecic abundance (29% of the control), and total biomass (49% of the control) over the first 18 months of experiment. The Shannon diversity index also decreased in the mixture treatment (both fungicides at the recommended dose) at 1 and 6 months after the first application (68% of the control at both sampling dates), and in C10 (78% of the control) at 18 months compared with the control. Lumbricus terrestris, Aporrectodea caliginosa, Aporrectodea giardi, Aporrectodea longa, and Allolobophora chlorotica were (in decreasing order) the most sensitive species to the tested fungicides. This study not only addressed field ecotoxicological effects of fungicides at the community level and ecological recovery, but it also pinpointed some methodological weaknesses (e.g., regarding fungicide concentrations in soil and statistics) of the guideline to determine the effects on earthworms in field situations.

Effects of two common fungicides on the reproduction of Aporrectodea caliginosa in natural soil.

The use of pesticides in agroecosystems can have negative effects on earthworms, which play key roles in soil functioning such as organic matter decomposition. The aim of this study was to assess the effects of two fungicides (Cuprafor micro®, composed of copper oxychloride, and Swing Gold®, composed of epoxiconazole (EPX) and dimoxystrobin (DMX)) on earthworm reproduction by exposing adults and cocoons. First, adult Aporrectodea caliginosa individuals were exposed for 28 days to 3.33, 10 and 30 times the recommended dose (RD) of Cuprafor micro® corresponding to 25.8, 77.5 and 232.5 mg kg-1 dry soil of copper, respectively, and 0.33, 1 and 3 times the RD of Swing Gold® (corresponding to 5.2 × 10-2 mg DMX kg-1 + 1.94 × 10-2 mg EPX kg-1, 1.55 × 10-1 mg DMX kg-1 + 5.81 × 10-2 mg EPX kg-1 and 4.62 × 10-1 mg DMX kg-1 + 1.74 × 10-1 mg EPX kg-1 respectively), in addition to a control soil with no fungicide treatment. Cocoon variables (production, weight, hatching success, hatching time) were monitored. Second, "naïve" cocoons produced by uncontaminated earthworms were exposed to soils contaminated by the same concentrations of the two fungicides, and we assessed hatching success and hatching time. In the first experiment, cocoon production was halved at the highest copper concentration (232.5 mg Cu kg-1 of dry soil) as compared to the control. Cocoons took 5 more days to hatch, and the hatching success decreased by 35% as compared to the control. In the Swing Gold® treatments, cocoon production was reduced by 63% at 3 times the RD, and the hatching success significantly decreased by 16% at the RD. In the second experiment, only the hatching success of cocoons was impacted by Swing Gold® at 3 times the RD (30% less hatching). It is concluded that the cocoon stock in the soil is crucial for the renewal of populations in the field. The most sensitive endpoint was the hatching success of the cocoons produced by exposed adults. This endpoint and the effects observed on the "naïve" cocoons could be taken into account in pesticide risk assessment.

Short-term effects of two fungicides on enchytraeid and earthworm communities under field conditions.

Because of the wide use of pesticides in agriculture, there is still a need of higher-tier field studies to assess ecotoxicological effects of commercial formulations on a wider variety of non-target soil organisms such as soil annelids. We here tested the effects of different concentrations of two fungicide formulations, i.e., Cuprafor Micro® (composed of 500 g kg-1 copper oxychloride) and Swing Gold® (composed of 50 g l-1 epoxiconazole and 133 g l-1 dimoxystrobin) on two families of terrestrial oligochaetes (Lumbricidae and Enchytraeidae) after 1 month of exposure. We also assessed the feeding activity of soil organisms using the bait lamina method. Along with the feeding activity, the enchytraeid density, diversity and communities were not different in the control and the contaminated plots. By contrast, epigeic earthworms were absent and earthworm diversity and densities of anecic species decreased significantly in the plots contaminated at ten times the recommended dose of the Swing Gold® formulation. The copper fungicide (at 0.75 and 7.5 kg Cu ha-1) and the treatment with the pesticide mixture (Cuprafor Micro® at 0.75 kg Cu ha-1 and Swing Gold® at the recommended dose) did not affect Oligochaeta communities compared with the control, except the Shannon index for earthworms in the mixture of both fungicides. Responses of the two annelid families to the tested pesticides were different with higher effects observed on the diversity and the community structure of earthworms compared with enchytraeids. This study allowed detecting early changes on oligochaete populations after pesticide application.

Differences in sensitivity between earthworms and enchytraeids exposed to two commercial fungicides.

The use of pesticides in crop fields may have negative effects on soil Oligochaeta Annelida, i.e., earthworms and enchytraeids, and thus affect soil quality. The aim of this study was to assess the effects of two commercial fungicide formulations on the earthworm Aporrectodea caliginosa and the enchytraeid Enchytraeus albidus in a natural soil. The fungicides were Cuprafor micro® (copper oxychloride), commonly used in organic farming, and Swing Gold® (epoxiconazole and dimoxystrobin), a synthetic fungicide widely used in conventional farming to protect cereal crops. Laboratory experiments were used to assess the survival, biomass loss and avoidance behaviour. No lethal effect was observed following exposure to the copper fungicide for 14 days, even at 5000mgkg-1 of copper, i.e. 650 times the recommended dose (RD). However, a significant decrease in biomass was observed from 50mgkg-1 of copper (6.5 times the RD) for A. caliginosa and at 5000mgkg-1 of copper (650 times the RD) for E. albidus. These sublethal effects suggest that a longer period of exposure would probably have led to lethal effects. The EC50 avoidance for the copper fungicide was estimated to be 51.2mgkg-1 of copper (6.7 times the RD) for A. caliginosa, and 393mgkg-1 of copper (51 times the RD) for E. albidus. For the Swing Gold® fungicide, the estimated LC50 was 7.0 10-3mLkg-1 (6.3 times the RD) for A. caliginosa and 12.7 10-3mLkg-1 (11.0 times the RD) for E. albidus. No effect on biomass or avoidance was observed at sublethal concentrations of this synthetic fungicide. It was concluded that enchytraeids were less sensitive than earthworms to the two commercial fungicides in terms of mortality, biomass loss and avoidance behaviour. Therefore we discuss the different strategies possibly used by the two Oligochaeta species to cope with the presence of the pesticides were discussed, along with the potential consequences on the soil functions.

Transgenerational Adaptation to Pollution Changes Energy Allocation in Populations of Nematodes.

Assessing the evolutionary responses of long-term exposed populations requires multigeneration ecotoxicity tests. However, the analysis of the data from these tests is not straightforward. Mechanistic models allow the in-depth analysis of the variation of physiological traits over many generations, by quantifying the trend of the physiological and toxicological parameters of the model. In the present study, a bioenergetic mechanistic model has been used to assess the evolution of two populations of the nematode Caenorhabditis elegans in control conditions or exposed to uranium. This evolutionary pressure resulted in a brood size reduction of 60%. We showed an adaptation of individuals of both populations to experimental conditions (increase of maximal length, decrease of growth rate, decrease of brood size, and decrease of the elimination rate). In addition, differential evolution was also highlighted between the two populations once the maternal effects had been diminished after several generations. Thus, individuals that were greater in maximal length, but with apparently a greater sensitivity to uranium were selected in the uranium population. In this study, we showed that this bioenergetics mechanistic modeling approach provided a precise, certain, and powerful analysis of the life strategy of C. elegans populations exposed to heavy metals resulting in an evolutionary pressure across successive generations.

A physiologically based toxicokinetic model for the zebrafish Danio rerio.

Zebrafish (Danio rerio) is a widely used model for toxicological studies, in particular those related to investigations on endocrine disruption. The development and regulatory use of in vivo and in vitro tests based on this species can be enhanced by toxicokinetic modeling. For this reason, we propose a physiologically based toxicokinetic (PBTK) model for zebrafish describing the uptake and disposition of organic chemicals. The model is based on literature data on zebrafish, other cyprinidae and other fish families, new experimental physiological information (volumes, lipids and water contents) obtained from zebrafish, and chemical-specific parameters predicted by generic models. The relevance of available models predicting the latter parameters was evaluated with respect to gill uptake and partition coefficients in zebrafish. This evaluation benefited from the fact that the influence of confounding factors such as body weight and temperature on ventilation rate was included in our model. The predictions for six chemicals (65 data points) yielded by our PBTK model were compared to available toxicokinetics data for zebrafish and 88% of them were within a factor of 5 of the corresponding experimental values. Sensitivity analysis highlighted that the 1-octanol/water partition coefficient, the metabolism rate, and all the parameters that enable the prediction of assimilation efficiency and partitioning of chemicals need to be precisely determined in order to allow an effective toxicokinetic modeling.

Consequences of a multi-generation exposure to uranium on Caenorhabditis elegans life parameters and sensitivity.

The assessment of toxic effects at biologically and ecologically relevant scales is an important challenge in ecosystem protection. Indeed, stressors may impact populations at much longer term than the usual timescale of toxicity tests. It is therefore important to study the evolutionary response of a population under chronic stress. We performed a 16-generation study to assess the evolution of two populations of the ubiquitous nematode Caenorhabditis elegans in control conditions or exposed to 1.1 mM of uranium. Several generations were selected to assess growth, reproduction, survival, and dose-responses relationships, through exposure to a range of concentrations (from 0 to 1.2 mM U) with all endpoints measured daily. Our experiment showed an adaptation of individuals to experimental conditions (increase of maximal length and decrease of fecundity) for both populations. We also observed an increase of adverse effects (reduction of growth and fertility) as a function of uranium concentration. We pointed out the emergence of population differentiation for reproduction traits. In contrast, no differentiation was observed on growth traits. Our results confirm the importance of assessing environmental risk related to pollutant through multi-generational studies.

Biology-based modeling to analyze uranium toxicity data on Daphnia magna in a multigeneration study.

Recent studies have investigated chronic toxicity of waterborne depleted uranium on the life cycle and physiology of Daphnia magna. In particular, a reduction in food assimilation was observed. Our aims here were to examine whether this reduction could fully account for observed effects on both growth and reproduction, for three successive generations, and to investigate through microscope analyses whether this reduction resulted from direct damage to the intestinal epithelium. We analyzed data obtained by exposing Daphnia magna to uranium over three successive generations. We used energy-based models, which are both able to fit simultaneously growth and reproduction and are biologically relevant. Two possible modes of action were compared - decrease in food assimilation rate and increase in maintenance costs. In our models, effects were related either to internal concentration or to exposure concentration. The model that fitted the data best represented a decrease in food assimilation related to exposure concentration. Furthermore, observations of consequent histological damage to the intestinal epithelium, together with uranium precipitates in the epithelial cells, supported the assumption that uranium has direct effects on the digestive tract. We were able to model the data in all generations and showed that sensitivity increased from one generation to the next, in particular through a significant increase of the intensity of effect, once the threshold for appearance of effects was exceeded.

A linear model to predict chronic effects of chemicals on Daphnia magna.

Chronic toxicity data for Daphnia magna are information requirements in the context of regulations on chemical safety. This paper proposes a linear model for the prediction of chemically-induced effects on the reproductive output of D. magna. This model is based on data retrieved from the Japanese Ministry of Environment database and it predicts chronic effects as a function of acute toxicity data. The proposed model proved to be able to predict chronic toxicities for chemicals not used in the training set. Our results suggest that experiments involving chronic exposure to chemicals could be reduced thanks to the proposed model.

Substance-tailored testing strategies in toxicology: an in silico methodology based on QSAR modeling of toxicological thresholds and Monte Carlo simulations of toxicological testing.

The design of toxicological testing strategies aimed at identifying the toxic effects of chemicals without (or with a minimal) recourse to animal experimentation is an important issue for toxicological regulations and for industrial decision-making. This article describes an original approach which enables the design of substance-tailored testing strategies with a specified performance in terms of false-positive and false-negative rates. The outcome of toxicological testing is simulated in a different way than previously published articles on the topic. Indeed, toxicological outcomes are simulated not only as a function of the performance of toxicological tests but also as a function of the physico-chemical properties of chemicals. The required inputs for our approach are QSAR predictions for the LOAELs of the toxicological effect of interest and statistical distributions describing the relationship existing between in vivo LOAEL values and results from in vitro tests. Our methodology is able to correctly predict the performance of testing strategies designed to analyze the teratogenic effects of two chemicals: di(2-ethylhexyl)phthalate and Indomethacin. The proposed decision-support methodology can be adapted to any toxicological context as long as a statistical comparison between in vitro and in vivo results is possible and QSAR models for the toxicological effect of interest can be developed.

Comparison of models to analyze mortality data and derive concentration-time response relationship of inhaled chemicals.

The derivation of thresholds for lethal effects for inhaled chemicals is a key issue in accidental risk management because they largely determine the outcome of land use planning, among which localization of habitations in the vicinity of a factory. This derivation is generally performed on the basis of rodent lethality data analyzed by statistical models able to extrapolate effects for different times and concentrations of exposure. A model commonly used in France is the standard probit model. In this model, effects is related to exposure concentration and duration according to the Haber's law and considers that individual thresholds, corresponding to the maximum tolerated effects before dying, are log-normally distributed among the population. This approach has been criticized for its lack of biological parameters and its inability to treat data characterized by only one time of exposure. In order to improve the current state of modeling, we proposed three alternative models. Two of them (DEBtox and Haber-TKTD models) incorporate the kinetics of the chemicals. The third one (Loguniform model) is a linearization of the standard probit model. We evaluated their performance by analyzing real data and simulated data generated with each model. For data characterized by several times of exposure, the standard probit model outperformed all other models in terms of goodness of fits and estimation of parameters. For data characterized by only one time of exposure, only DEBtox model was able to fit the data and estimate parameters, provided we dispose of several observation times, typically just after exposure and a long period afterwards.

Soil dissipation and bioavailability to earthworms of two fungicides under laboratory and field conditions.

The representativeness of laboratory studies of the fate of pesticides in soil in field conditions is questionable. This study aimed at comparing the dissipation and bioavailability to earthworms of two fungicides, dimoxystrobin (DMX) and epoxiconazole (EPX), over 12 months under laboratory and field conditions. In both approaches, the fungicides were applied to the same loamy soil as a formulated mixture at several concentrations. We determined total DMX and EPX concentrations in the soil using exhaustive extraction, their environmental availability using mild extraction and their bioavailability through internal concentrations in exposed earthworms. The initial fungicide application appeared as much better controlled in terms of dose and homogeneity in the laboratory than in the field. One year after application, a similar dissipation rate was observed between the laboratory and field experiments (ca 80% and 60% for DMX and EPX, respectively). Similarly, the ratio of available/total concentrations in soil displayed the same trend whatever the duration and the conditions (field or lab), EPX being more available than DMX. Finally, the environmental bioavailability of the two fungicides to earthworms was heterogeneous in the field, but, in the laboratory, the bioaccumulation was evidenced to be dose-dependent only for DMX. Our findings suggest that the actual fate of the two considered fungicides in the environment is consistent with the one determined in the laboratory, although the conditions differed (e.g., presence of vegetation, endogeic earthworm species). This study allowed better understanding of the fate of the two considered active substances in the soil and underlined the need for more research dedicated to the link between environmental and toxicological bioavailability.

An energy-based model to analyze growth data of earthworms exposed to two fungicides.

The pesticide risk assessment for earthworms is currently performed using standardized tests, the model species Eisenia fetida, and the analyses of the data obtained are performed with ad hoc statistical tools. We assessed the impact of two fungicides on the entire growth pattern of the earthworm species Aporrectodea caliginosa, which is highly representative of agricultural fields. Individuals of three different ages (from hatching to 56 days old) were exposed to Cuprafor micro® (copper oxychloride) and Swing® Gold (dimoxystrobin and epoxiconazole). Data were analyzed with an energy-based toxicodynamic model coupled with a toxicokinetic model. The copper fungicide caused a drastic growth inhibition once the no effect concentration (NEC), estimated at 65 mg kg-1 of copper, was exceeded. The Swing® Gold negatively affected the growth with NEC values estimated at 0.387 mg kg-1 and 0.128 mg kg-1 for the dimoxystrobin and the epoxiconazole in this fungicide formulation, respectively. The time-profile of the effects on A. caliginosa individuals was fully accounted for by the model, whatever their age of exposure. Furthermore, toxicity data analyses, supported by measurements of fungicide concentrations in earthworm at the end of the experiment, allowed bettering understanding of the mechanisms of action of the fungicides towards earthworm growth.

Development of a physiologically based kinetic model for 99m-technetium-labelled carbon nanoparticles inhaled by humans.

Particulate air pollution is associated with respiratory and cardiovascular morbidity and mortality. Recent studies investigated whether and to which extent inhaled ultrafine particles are able to translocate into the bloodstream in humans. However, their conclusions were conflicting. We developed a physiologically based kinetic model for (99m)technetium-labelled carbon nanoparticles (Technegas). The model was designed to analyse imaging data. It includes different translocation rates and kinetics for free technetium, and small and large technetium-labelled particles. It was calibrated with data from an experiment designed to assess the fate of nanoparticles in humans after inhalation of Technegas. The data provided time courses of radioactivity in the liver, stomach, urine, and blood. Parameter estimation was performed in a Bayesian context with Markov chain Monte Carlo (MCMC) techniques. Our analysis points to a likely translocation of particle-bound technetium from lung to blood, at a rate about twofold lower than the transfer rate of free technetium. Notably, restricting the model so that only free technetium would have been able to reach blood circulation resulted in much poorer fits to the experimental data. The percentage of small particles able to translocate was estimated at 12.7% of total particles. The percentage of unbound technetium was estimated at 6.7% of total technetium. To our knowledge, our model is the first PBPK model able to use imaging data to describe the absorption and distribution of nanoparticles. We believe that our modeling approach using Bayesian and MCMC techniques provides a reasonable description on which to base further model refinement.

Earthworms Mitigate Pesticide Effects on Soil Microbial Activities.

Earthworms act synergistically with microorganisms in soils. They are ecosystem engineers involved in soil organic matter degradation and nutrient cycling, leading to the modulation of resource availability for all soil organisms. Using a soil microcosm approach, we aimed to assess the influence of the earthworm Aporrectodea caliginosa on the response of soil microbial activities against two fungicides, i.e., Cuprafor Micro® (copper oxychloride, a metal) and Swing® Gold (epoxiconazole and dimoxystrobin, synthetic organic compounds). The potential nitrification activity (PNA) and soil enzyme activities (glucosidase, phosphatase, arylamidase, and urease) involved in biogeochemical cycling were measured at the end of the incubation period, together with earthworm biomass. Two common indices of the soil biochemistry were used to aggregate the response of the soil microbial functioning: the geometric mean (Gmean) and the Soil Quality Index (SQI). At the end of the experiment, the earthworm biomass was not impacted by the fungicide treatments. Overall, in the earthworm-free soil microcosms, the two fungicides significantly increased several soil enzyme and nitrification activities, leading to a higher GMean index as compared to the non-treated control soils. The microbial activity responses depended on the type of activity (nitrification was the most sensitive one), on the fungicide (Swing® Gold or Cuprafor Micro®), and on the doses. The SQI indices revealed higher effects of both fungicides on the soil microbial activity in the absence of earthworms. The presence of earthworms enhanced all soil microbial activities in both the control and fungicide-contaminated soils. Moreover, the magnitude of the fungicide impact, integrated through the SQI index, was mitigated by the presence of earthworms, conferring a higher stability of microbial functional diversity. Our results highlight the importance of biotic interactions in the response of indicators of soil functioning (i.e., microbial activity) to pesticides.

Statistical cautions when estimating DEBtox parameters.

DEBtox (Dynamic Energy Budget in toxicology) models have been designed to analyse various results from classic tests in ecotoxicology. They consist of a set of mechanistic models describing how organisms manage their energy, when they are exposed to a contaminant. Until now, such a biology-based modeling approach has not been used within the regulatory context. However, these methods have been promoted and discussed in recent guidance documents on the statistical analysis of ecotoxicity data. Indeed, they help us to understand the underlying mechanisms. In this paper, we focused on the 21 day Daphnia magna reproduction test. We first aimed to clarify and detail the model building process leading to DEBtox models. Equations were rederived step by step, and for some of them we obtained results different from the published ones. Then, we statistically evaluated the estimation process quality when using a least squares approach. Using both experimental and simulated data, our analyses highlighted several statistical issues related to the fitting of DEBtox models on OECD-type reproduction data. In this case, particular attention had to be paid to parameter estimates and the interpretation of their confidence intervals.

Assessing the risk of metal mixtures in contaminated sediments on Chironomus riparius based on cytosolic accumulation.

Sediments usually contain mixtures of trace metals introduced via natural geochemical processes and anthropogenic activities. Kinetics and effects of these metals are strongly dependent both on the composition of the mixture and on the physico-chemical characteristics of the sediment. Relating effects to metal concentration may consequently be advised. However, total accumulation may be a poor predictor of metal toxicity for Chironomus riparius exposed to contaminated field sediments. As an alternative, we proposed to relate effects on Chironomus growth with cytosolic metal accumulation, measured in larvae after a short exposure period. Dose-response relationships were derived for zinc, copper, and cadmium through single-metal exposure data analysed with toxicokinetics and toxicodynamics models. They permitted, on the basis of cytosolic accumulation measures, to predict successfully the effects of mixtures of cadmium, zinc, and copper on the growth of larvae exposed to spiked sediments, as well as to field sediments in which zinc and copper were assumed to be predominant.

How to assess the feeding activity in ecotoxicological laboratory tests using enchytraeids?

The risk assessment of pesticides on soil fauna is an issue to protect agroecosystem sustainability. Enchytraeids are recognized as relevant soil bioindicators of chemical stress in agroecosystems. In laboratory, the reproduction test was found to be sensitive to reveal chemical impacts on enchytraeids. However, it does not allow to assess the impacts on ecological functions in which enchytraeids are involved. The objectives of this study were (i) to explore the feasibility of the bait-lamina test with enchytraeids under laboratory conditions and (ii) to compare its sensitivity with the Enchytraeid Reproduction Test. We exposed individuals of Enchytraeus albidus to two commercial formulations of fungicides widely used in Europe. The EC50 reproduction for the Swing® Gold (50 g L-1 epoxiconazole and 133 g L-1 dimoxystrobin) and the Cuprafor micro® (50% copper oxychloride) were respectively estimated at 1.66 ± 0.93 times the recommended dose and > 496 mg kg-1 of copper. However, no impact was found on the feeding activity of enchytraeids. The bait-lamina test thus appeared less sensitive than the Enchytraeid Reproduction Test to the tested fungicides. Despite that, this test which is achievable under laboratory conditions and allows assessing indirect effects of pesticides is quick, cheap, and easy to perform. It would deserve to be used to explore longer-exposure effects through the repeated addition of bait-lamina sticks.

Aporrectodea caliginosa, a relevant earthworm species for a posteriori pesticide risk assessment: current knowledge and recommendations for culture and experimental design.

Ecotoxicological tests with earthworms are widely used and are mandatory for the risk assessment of pesticides prior to registration and commercial use. The current model species for standardized tests is Eisenia fetida or Eisenia andrei. However, these species are absent from agricultural soils and often less sensitive to pesticides than other earthworm species found in mineral soils. To move towards a better assessment of pesticide effects on non-target organisms, there is a need to perform a posteriori tests using relevant species. The endogeic species Aporrectodea caliginosa (Savigny, 1826) is representative of cultivated fields in temperate regions and is suggested as a relevant model test species. After providing information on its taxonomy, biology, and ecology, we reviewed current knowledge concerning its sensitivity towards pesticides. Moreover, we highlighted research gaps and promising perspectives. Finally, advice and recommendations are given for the establishment of laboratory cultures and experiments using this soil-dwelling earthworm species.

Rearing and estimation of life-cycle parameters of the tubicifid worm Branchiura sowerbyi: application to ecotoxicity testing.

This paper provides original collection, acclimatizing, rearing and toxicity test methods for the freshwater worm Branchiura sowerbyi, an alternative species to Tubifex tubifex for ecotoxicity evaluation of sediments. Influence of the substrate, type of food, and feeding level on individual performances was assessed in short-term tests, in order to set up optimal culture and test conditions. Low-size particles and high organic matter content favoured the growth and reproduction of B. sowerbyi. The relative contribution of sediments and fish food to the individual food intake was assessed using a foraging efficiency model based upon the dynamic energy budget theory. Individual performances were optimal when the substrate plus fish food provided the energy equivalent to 5 mg Tetramin per worm per day, which is the ad libitum food level for adults at 21 degrees C. The life-cycle of the worm was fully characterized using a life-cycle test conducted under the previously defined optimal conditions. Hatching rates were low (32%), whereas newborn and juveniles exhibited high survival (>80%) and growth (2.4 mg/day in juveniles) rates. Age at puberty was low (60 days) when compared to the maximal life span (1100 days) as predicted using a Weibull model. Adults reproduced every other month with a constant fecundity (0.16 cocoon/worm/day). The mean values of the life-cycle parameters and their variability and reproducibility among laboratory studies were discussed in order to identify relevant endpoints to be used in ecotoxicity tests. Survival, juvenile growth, and fecundity may constitute suitable test endpoints, whereas hatching rate and adult growth should not be used as endpoints in B. sowerbyi.