Christelle Clérandeau

Evaluation of enzymatic biomarkers and lipoperoxidation level in Hediste diversicolor exposed to copper and benzo[a]pyrene.

This study aims to evaluate the effects of exposure to copper, benzo[a]pyrene, and to their mixture on enzymatic and lipid peroxidation biomarkers in Hediste diversicolor. Worms were submitted to 1 microM of both single compounds and to their mixture during a period of test of 12, 24, 36, and 48 h. The biomarkers selected in this work were the activities of cytochrome P450-dependent NADPH cytochrome c reductase (NADPH red) as phase I enzyme, glutathione-S-transferase (GST) as phase II enzyme, and the acetylcholinesterase (AChE) activity as neurotoxicity marker. Oxidative stress was evaluated using catalase activity (CAT) and malondialdehyde accumulation (MDA). The NADPH red activity was not significantly affected by copper exposure; it shows a drastic increase in both B[a]P and mixture-exposed organisms. GST activities were significant in B[a]P-exposed worms only after 36 h, and in animals exposed to the mixture after 12 and 48 h. The ACHE activity was inhibited only in B[a]P-exposed worms.

Mixture toxicity assessment of cadmium and benzo[a]pyrene in the sea worm Hediste diversicolor.

In the present study, Hediste diversicolor biotransformation and anti-oxidant responses to acute exposure to cadmium (Cd) and to the polycyclic aromatic hydrocarbons benzo[a]pyrene (B[a]P) were investigated. Worms were submitted to 0.2, 0.4 and 1 microM of each contaminant and to their mixture during a period of test of 48h. Following biological responses were measured: (1) NADPH cytochrome c reductase (NADPH cyt c) activity, as phase I biotransformation parameter; (2) gluthathione-S-transferase (GST) activity as a phase II conjugation enzyme, (3) catalase activity as anti-oxidant response and (4) malondialdehyde accumulation (MDA) as lipid peroxydation marker. The cholinergic system was evaluated using the acetylcholinesterase activity (AChE). Exposure to the mixture resulted in low dose level additive effects on the investigated biomarkers. However, worms exposed to 1 microM of the single compounds and to their mixture exhibited the highest MDA accumulation and the lowest enzymatic biomarkers activities suggesting severe toxicological effects. These data should be carefully considered in view of the biological effects of mixture pollutants and particularly in marine sediment ecosystems.

Assessment of pollution in the Bizerte lagoon (Tunisia) by the combined use of chemical and biochemical markers in mussels, Mytilus galloprovincialis.

In order to assess the environmental quality of the Bizerte lagoon (Tunisia), biomarker and contaminant levels were measured in Mediterranean mussels (Mytilus galloprovincialis) from five selected sites. Persistent organic pollutants (POPs) were quantified in whole body and enzyme activities such as acetylcholinesterase (AChE), catalase (CAT) and glutathione S-transferase (GST) in gills. Despite the relatively low levels of organic contaminants, the selected biomarkers responded differently according to the pollution level at the different sites. GST and AChE activities were correlated with the amount of DDTs in mussel tissues. These two enzymatic activities were also correlated to temperature and pH. No significant difference was observed for CAT activity. Principal component analysis showed a clear separation of sampling sites in three different assemblages which is consistent with POP body burden in mussels. Our results confirmed the usefulness of combining biomarker and chemical analyses in mussels to assess chemical pollution in the Bizerte lagoon.

Combined effects of temperature and copper and S-metolachlor on embryo-larval development of the Pacific oyster, Crassostrea gigas

This study evaluates the combined effects of two contaminants (copper and a herbicide S-metolachlor) and temperature on the early life stages of the Pacific oyster Crasssotrea gigas, which is native to Arcachon Bay (southwest France). The responses of D-larvae, obtained from wild and cultivated oysters, were investigated for one year during the oyster breeding period at different sampling sites and compared with the response of D-larvae from a commercial hatchery. Embryotoxicity was measured by considering the percentage of abnormal D-shaped larvae. Normal development of embryos occurred only from 22°C to 26°C. There were synergistic effects of copper at low and high temperatures. Native oysters appear particularly sensitive to an increase in water temperature, suggesting a future increase in the percentage of larval abnormalities as a result of global climate change. Hatchery oysters represent a good alternative model for studying the effects of both pollutants and climate change stressors.

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.

Assessing the toxicity of sediments using the medaka embryo–larval assay and 2 other bioassays

Sediments are sinks for aquatic pollutants, and analyzing toxicity in such complex matrices is still challenging. To evaluate the toxicity of bioavailable pollutants accumulated in sediments from the Bizerte lagoon (Tunisia), a novel assay, the medaka embryo-larval assay by sediment contact, was applied. Japanese medaka (Oryzias latipes) embryos were incubated in direct contact with sediment samples up to hatching. Lethal and sublethal adverse effects were recorded in embryos and larvae up to 20 d postfertilization. Results from medaka embryo-larval assay were compared with cytotoxicity (Microtox®), genotoxicity (SOS chromotest), and pollutant content of sediments. The results highlight differences in the contamination profile and toxicity pattern between the different studied sediments. A significant correlation was shown between medaka embryo-larval assay by sediment contact and SOS chromotest responses and concentrations of most organic pollutants studied. No correlation was shown between pollutant levels and Microtox. According to the number of sediment samples detected as toxic, medaka embryo-larval assay by sediment contact was more sensitive than Microtox, which in turn was more sensitive than the SOS chromotest; and medaka embryo-larval assay by sediment contact allowed sediment toxicity assessment of moderately polluted sediments without pollutant extraction and using an ecologically realistic exposure scenario. Although medaka embryo-larval assay by sediment contact should be tested on a larger sample set, the results show that it is sensitive and convenient enough to monitor the toxicity of natural sediments. Environ Toxicol Chem 2016;35:2270-2280.

Usefulness of RTL-W1 and OLCAB-e3 fish cell lines and multiple endpoint measurements for toxicity evaluation of unknown or complex mixture of chemicals

Fish are currently used for the assessment of chemical toxicity. The REACh regulation and the European directive on the protection of animals used for scientific purposes both recommend the use of methods other than animal testing. In view of this, fish cell lines are increasingly used to provide fast and reliable toxic and ecotoxic data on new chemicals. The sensitivity of the Rainbow trout liver cell line RTL-W1 and Japanese medaka embryos cell line OLCAB-e3 were used with different toxicity endpoints, namely cytotoxicity, EROD activity, ROS production and DNA damage for various classes of pollutants displaying different modes of action but also with complex environmental mixtures. Toxicity tests were coupled with chemical analysis to quantify the chemical concentrations in cell cultures. Differences in sensitivity were found between fish cell lines. MTT reduction assay revealed that OLCAB-e3 cells were more sensitive than RTL-W1 cells. On the contrary, RTL-W1 gave higher response levels for the Fpg-modified comet assay and ROS assay. The OLCAB-e3 cell line did not express EROD activity unlike RTL-W1. This study highlights the capacity of the two different fish cell lines to measure the toxicity of individual toxicants but also environmental mixtures. Then, results obtained here illustrate the interest of using different cell lines and toxicity endpoints to assess the toxicity of complex or unknown mixture of chemicals.

An innovative and integrative assay for toxicity testing using individual fish embryos. Application to oxazepam

This paper describes the development of an integrative embryo-toxicity assay in Japanese medaka allowing analysis of several toxicological endpoints together in a same individual. In this assay, embryos are topically exposed, and survival, hatching success, malformations, biometry, behaviour, and target gene expression are subsequently analysed in each individual. This assay was applied to oxazepam, an anxiolytic pharmaceutical compound currently found in wastewater treatment plant effluent. Even if oxazepam accumulation in embryos was very low, it caused spinal and cardiac malformations, delayed growth, erratic swimming and deregulation of genes involved in apoptosis, DNA repair and mitochondrial metabolism. Relationship between gene deregulation, abnormal behaviour, and developmental anomalies was demonstrated. This assay is sensitive enough to detect adverse effects at low chemical concentrations and at multiple endpoints in a unique fish embryo. This integrative embryo-toxicity assay is a powerful tool to characterize the spectrum of effects of new chemicals and also to link effects induced at different molecular, tissue and physiological levels.

Toxicity Assessment of Pollutants Sorbed on Microplastics Using Various Bioassays on Two Fish Cell Lines

Microplastics are tiny plastic fragments resulted from runoff and weathering breakdown of larger plastics debris. They represent an emerging concern for marine ecosystems. Microplastics impacts on aquatic life are little studied. These small particles could be ingested directly by organisms and cause chronic physical and toxicological effects. Moreover, microplastics are the support for a lot of chemicals present in aquatic environment, especially hydrophobic substances. The aims of this study is to investigate toxicity of environmental microplastics samples collected during the Race for Water Odyssey in fish cell lines. A difference of sensitivity was observed between fish cell lines. Preliminary results obtained here support that RTLW-1 could be an interesting tool to evaluate the potential chemical toxicity of microplastics and sorbed pollutants. Toxicity of extracts from MP collected worldwide suggests a biological impact?

Using an Integrated Approach to Assess the Sediment Quality of an Mediterranean Lagoon, the Bizerte Lagoon (Tunisia)

The present study investigates the quality of surface sediments from the Bizerte lagoon (North Tunisia) using an integrated approach including chemical contaminant analysis, bioassays and sediment quality guidelines (SQGs). Sediment samples were collected at 9 sites and analyzed for eight heavy metals (Hg, Cd, Cr, Cu, Pb, Zn, Ni, Fe and Mn). PAHs, PCBs, OCPs were measured previously in the same sediment samples. Our results indicated that the highest concentrations of metals were found near urban areas due to the municipial and industrial wastewater discharges. Sediment pollution assessment was carried out using geoaccumulation index and enrichment factor, which indicate a widespread pollution by Cd, Pb, Ni and Zn in the studied sediments. For bioassays, aqueous and organic extracts were used to assess toxicity and genotoxicity in sediments by using Microtox® and SOS Chromotest, respectively. Toxicity levels were compared to metallic and organic pollutants contents. Our results highlight differences in the pattern of responses between the different assays and show no correlation with all the studied contaminants, emphasizing the influence of other contaminants not analyzed in the present study. Based on SQGs, the results of toxicity assessment indicated that adverse effects caused by Ni and Zn would be expected frequently. Nickel was found to have the highest predicted acute toxicity, followed by Zn, Pb, Cd, Cu and Cr. There was no significant relationship between sediment toxicity calculated from heavy metal concentrations (SQG approach) and those measured with bioassays. These findings support the use of integrated approachs for evaluating the environmental risks of sediments.