Benoît J.D. Ferrari

Towards a renewed research agenda in ecotoxicology

New concerns about biodiversity, ecosystem services and human health triggered several new regulations increasing the need for sound ecotoxicological risk assessment. The PEER network aims to share its view on the research issues that this challenges. PEER scientists call for an improved biologically relevant exposure assessment. They promote comprehensive effect assessment at several biological levels. Biological traits should be used for Environmental risk assessment (ERA) as promising tools to better understand relationships between structure and functioning of ecosystems. The use of modern high throughput methods could also enhance the amount of data for a better risk assessment. Improved models coping with multiple stressors or biological levels are necessary to answer for a more scientifically based risk assessment. Those methods must be embedded within life cycle analysis or economical models for efficient regulations. Joint research programmes involving humanities with ecological sciences should be developed for a sound risk management.

Chromium(VI) is more toxic than chromium(III) to freshwater algae: A paradigm to revise?

The behavior and toxicity of Cr(III) and Cr(VI) to the green algae Pseudokirchneriella subcapitata and Chlorella kessleri were studied in a standard culture medium (ISO medium) and, for P. subcapitata only, in ultrafiltered natural water enriched with all ISO components (modified ISO medium). In all solutions amended with Cr(III), initial chromium concentrations decreased by 60-90% over 72h (the duration of algal tests) indicating that protocols for testing poorly soluble substances are required to properly evaluate Cr(III) toxicity. After accounting for its behavior in test solutions, chromium(III) was 5-10 times more toxic than Cr(VI) in both media. For P. subcapitata, the average 72h EC50 of Cr(III) in ISO medium was 17.4+/-4.7 microg/L (n=9); lower than corresponding hardness-corrected Continuous Concentration Criteria of the US EPA and well within the range of Cr concentrations found in waters impacted by tannery discharges. These results follow from intrinsic chemical properties of Cr(III) in circumneutral solutions, so that the actual toxicity of Cr(III) to aquatic organisms may be generally underestimated.

Coupling geochemical and biological approaches to assess the availability of cadmium in freshwater sediment.

Sediments are considered as a sink for metals, and the assessment of metal bioavailability for benthic organisms represents a great challenge. Diffusive Gradient in Thin films (DGT), developed to measure labile metals in aquatic media, have more recently been applied to sediment. Nevertheless, few studies have determined the relation between measurements from DGT and bioaccumulation in different benthic organisms. The aim of our work was to determine if labile metal measured by DGT in sediment is representative of bioavailable metal for benthic organisms. We focused our work on Cd and chose to use the diversity of ecological traits from different organisms to better understand the measurement given by DGT. We exposed simultaneously DGT and 3 macroinvertebrates species (the chironomid, Chironomus riparius; the amphipod, Gammarus fossarum; the mudsnail, Potamopyrgus antipodarum) to a natural sediment Cd-spiked at environmental relevant concentrations. The nature of sediment-bound Cd was also determined by means of sequential extractions in order to better interpret DGT measurements. Cadmium concentrations were determined in DGT and in the 3 organisms after one week of exposure. Results provided by DGT indicated that Cd was poorly released from particulate phase to pore water, suggesting that Cd measured by DGT was representative of the pore water labile fraction. Sequential extractions showed that the percentage of Cd bound to carbonate fraction increased simultaneously with Cd-spiking level; hence, this Cd fraction was poorly reactive to supply DGT demand. Cadmium accumulation rates were similar between DGT measurements and P. antipodarum, suggesting that labile Cd in pore waters was representative of bioavailable Cd for this species. Cadmium accumulation rates in C. riparius were higher than in DGT, demonstrating that C. riparius can mobilize Cd bound to carbonate phase. G. fossarum showed the lowest Cd accumulation rates, suggesting that they were mainly exposed to Cd from overlaying waters.

Bioaccumulation of perfluoroalkyl compounds in midge (Chironomus riparius) larvae exposed to sediment.

Midge larvae (Chironomus riparius) were exposed to sediments from a deposition sampled at a site along the Rhône River (France) downstream of an industrial site releasing various perfluorinated chemicals. This sediment is characterized by high concentrations of perfluoroundecanoic acid (PFUnA) and perfluorotridecanoic acid (PFTrDA) and a low perfluorooctane sulfonate (PFOS) concentration. Concentrations of 23 perfluoroalkyl compounds, including C4-C14 carboxylate acids, C4-C10 sulfonates, and seven precursors, were analyzed in overlying and pore water, sediment, and larvae. Midge larvae accumulated carboxylate acids (C11-C14), PFOS, and two precursors (perfluorooctane sulfonamide: FOSA and 6:2 fluorotelomer sulfonic acid, 6:2 FTSA). These substances accumulated mainly during the fourth instar larvae exponential growth phase. Accumulation of 6:2 FTSA, PFUnA, and PFOS occured via trophic and tegumentary routes. Other compounds mainly accumulated from food. Kinetics followed a partition model, from which uptake and elimination constants were derived.

Mercury tissue residue approach in Chironomus riparius: Involvement of toxicokinetics and comparison of subcellular fractionation methods

Along with the growing body of evidence that total internal concentration is not a good indicator of toxicity, the Critical Body Residue (CBR) approach recently evolved into the Tissue Residue Approach (TRA) which considers the biologically active portion of metal that is available to contribute to the toxicity at sites of toxic action. For that purpose, we examined total mercury (Hg) bioaccumulation and subcellular fractionation kinetics in fourth stage larvae of the midge Chironomus riparius during a four-day laboratory exposure to Hg-spiked sediments and water. The debris (including exoskeleton, gut contents and cellular debris), granule and organelle fractions accounted only for about 10% of the Hg taken up, whereas Hg concentrations in the entire cytosolic fraction rapidly increased to approach steady-state. Within this fraction, Hg compartmentalization to metallothionein-like proteins (MTLP) and heat-sensitive proteins (HSP), consisting mostly of enzymes, was assessed in a comparative manner by two methodologies based on heat-treatment and centrifugation (HT&C method) or size exclusion chromatography separation (SECS method). The low Hg recoveries obtained with the HT&C method prevented accurate analysis of the cytosolic Hg fractionation by this approach. According to the SECS methodology, the Hg-bound MTLP fraction increased linearly over the exposure duration and sequestered a third of the Hg flux entering the cytosol. In contrast, the HSP fraction progressively saturated leading to Hg excretion and physiological impairments. This work highlights several methodological and biological aspects to improve our understanding of Hg toxicological bioavailability in aquatic invertebrates.

Potential exposure routes and accumulation kinetics for poly- and perfluorinated alkyl compounds for a freshwater amphipod: Gammarus spp. (Crustacea)

Gammarids were exposed to sediments from a deposition site located on the Rhône River (France) downstream of a fluoropolymer manufacturing plant. Gammarids accumulated to various extents four long-chain perfluoroalkyl carboxylic acids (PFCAs) from C9 to C13, one sulfonate, perfluorooctane sulfonate (PFOS) and three of its precursors (the perflurooctane sulfonamide (FOSA), the N-methyl perfluorooctane sulfonamidoacetic acid (MeFOSAA), the N-ethyl perfluorooctane sulfonamidoacetic acid (EtFOSAA) and the 6:2 fluorotelomer sulfonic acid (6:2 FTSA). Whatever the compound, the steady state was not achieved after a 3-week exposure; elimination was almost complete after a 3-week depuration period for perfluorononanoic acid (PFNA), PFOS, the three precursors and the 6:2FTSA. However, this was not the case for long-chain PFCAs, whose elimination rates decreased with increasing chain length. PFAS accumulation in gammarids occurred via the trophic and respiratory pathways, in proportions varying with the carbon chain length and the terminal moiety.

Bioaccumulation kinetics and effects of sediment-bound contaminants on chironomids in deep waters: new insights using a low-disturbance in situ system

Effective assessment of sediment quality and associated risks requires further integration between laboratory-based studies and field observations. Tools for in situ exposure of laboratory-reared organisms are particularly suitable for this purpose. However, available tools suffer from technical drawbacks that limit their use in deep waters and their general ability to provide results linking laboratory and field observations. To overcome these limitations, we developed a new device (Sediment–Water Interface Study System – S-WISS1) for in situ exposure of fourth instar larvae of Chironomus riparius at depths up to 18 m. S-WISS1 allows minimum alteration of the sediment natural structure (upon both deployment and recovery) and, in a way analogous to laboratory tests, repeated delivery of food doses to the in situ exposed organisms. Results obtained in situ with S-WISS1 were compared with standard laboratory tests and simple caging systems (large-mouthed bottles partly filled with sand) used as field controlks. A first series of experiments (48 h, no food addition) showed that element bioavailability to chironomids was higher in laboratory settings than on-site or in situ. A second series of experiments (96 h, food addition every 24 h) confirmed that sediments rather than overlying waters acted as the principal source of contaminants and that exposure to field sediments reduced chironomids’ growth compared with on-site controls. S-WISS1 represents a new family of in situ tools capable of combining the environmental realism associated with field monitoring with the controlled experimental conditions typical of laboratory testing.

Per- and poly-fluoroalkyl compounds in freshwater fish from the Rhône River: Influence of fish size, diet, prey contamination and biotransformation

Pools of aquatic plants and benthic invertebrates were collected along with 47 individuals from three cyprinid fish species (Barbus barbus, Gobio gobio, Rutilus rutilus) at a site in the Rhône River (France). Carbon and nitrogen isotopic ratios (δ13C and δ15N) and a wide range of per- and poly-fluorinated chemicals (PFASs) were analysed in all samples. The sum of PFAS concentrations (ΣPFAS) increased from aquatic plants to fish dorsal muscles; molecular profiles were dominated by C9-C13 perfluorocarboxylic acids (PFCAs), while perfluorooctane sulfonate (PFOS) and perfluorooctane sulfonamide (FOSA) were detected in all samples at lower concentrations. ΣPFAS and especially ΣPFCAs were higher in barbels (B. barbus) than in other species, while roaches (R. rutilus) were less contaminated by PFOS than barbels and gudgeons (G. gobio). Gudgeons accumulated significantly higher FOSA concentrations. Young (small) barbels displayed significantly higher PFOS, perfluorononanoic acid (PFNA) and perfluorodecanoic acid (PFDA) concentrations than did large specimens; conversely, perfluorotetradecanoic acid (PFTeDA) concentrations were significantly higher in large barbels. Multiple linear regressions were performed on the whole set of fish samples with size, mass and isotopic ratios as explicative variables, and several single compounds as explained variables. Regardless of the compound, the regressions did not explain much of the contamination variability. However, adding species as a qualitative variable, i.e. performing analyses of covariance (ANCOVAs) improved the fit greatly, while adding sex did not. Diet (i.e. δ13C and δ15N) was the main factor explaining interspecific differences. Biotransformation was assessed by comparing concentration ratios of PFOS or FOSA to their precursors in the food-web compartments. These ratios increased from invertebrates to fish, and differed among fish species, suggesting that biotransformation occurred but was species-specific. Biomagnification factor calculations showed that C11-C13 PFCAs, PFOS and FOSA were apparently biomagnified in barbels and gudgeons.

Cytochrome c oxidase barcodes for aquatic oligochaete identification: development of a Swiss reference database

Introduction Aquatic oligochaetes represent valuable indicators of the quality of sediments of watercourses and lakes, but their difficult identification based on morphological criteria compromises their more common use for eco-diagnostic analyses. This issue could be overcome by using DNA barcodes for species identification. A 10% threshold of cytochrome c oxidase (COI) divergence was proposed for differentiating between oligochaete species based on molecular and morphological data. A Swiss database of COI sequences of aquatic oligochaetes was initiated in 2012. The aim of this study is to complement the Swiss oligochaete database of COI sequences and to confirm the relevance of this threshold for species delimitation. Methods We sequenced the COI sequence of 216 specimens collected in different regions of Switzerland and ITS2 region of some lineages whose delimitation with COI data was doubtful. Results We distinguished 53 lineages, among which 34 were new for Switzerland and 17 sequenced for the first time. All the lineages were separated by more than 10% of COI variation, with the exception of some species within Nais and Uncinais. In these two genera, the threshold was lowered to 8% to be congruent with the morphological analysis. The total number of lineages reported so far for Switzerland is 75, including 59 morphospecies or unidentified species and 16 cryptic species. Discussion Our study shows that the threshold of 10% of COI divergence is generally appropriate to distinguish aquatic oligochaete lineages, but that it must be adjusted for some species. The database reported here will be complemented in the future in parallel to the development of genetic oligochaete indices.

On the bioavailability of trace metals in surface sediments: a combined geochemical and biological approach

The bioavailability of metals was estimated in three river sediments (Sensée, Scarpe, and Deûle Rivers) impacted by different levels of Cu, Cd, Pb, and Zn (Northern France). For that, a combination of geochemistry and biological responses (bacteria and chironomids) was used. The results obtained illustrate the complexity of the notion of “bioavailability.” Indeed, geochemical indexes suggested a low toxicity, even in surface sediments with high concentrations of total metals and a predicted severe effect levels for the organisms. This was also suggested by the abundance of total bacteria as determined by DAPI counts, with high bacterial cell numbers even in contaminated areas. However, a fraction of metals may be bioavailable as it was shown for chironomid larvae which were able to accumulate an important quantity of metals in surface sediments within just a few days.We concluded that (1) the best approach to estimate bioavailability in the selected sediments is a combination of geochemical and biological approaches and that (2) the sediments in the Deûle and Scarpe Rivers are highly contaminated and may impact bacterial populations but also benthic invertebrates.