Search

International audience; Session: Microbial roles in contaminant fate and bioremediationThe potential adverse effects of pesticides on stream ecosystems subjected to extreme hydrological events has been poorly explored. Therefore, there is an urgent need to increase our knowledge to propose suitable pesticide mitigation strategies in the context of global change scenarios affecting stream ecosystems. In this study, a microcosm experiment was performed to assess the dissipation kinetics of a pulse of pesticide mixture (glyphosate,aminomethylphosphonic acid (AMPA), tebuconazole, terbuthylazine and imidacloprid) by stream biofilms with a different hydrological history. The effects of two types of drought events caused by hydropeaking (short and more frequent droughts) and agricultural practices (long and less frequent droughts) on aquatic biofilm structure-function were evaluated and compared with an immersed control, before and after the pesticide cocktail pulse. Different biomass and functional parameters were analyzed in biofilms to determine the combined impact of droughts and pesticides, whereas the dissipation kinetics of each pesticide molecule of the cocktail wasevaluated in the water as well as in the biofilm. Algal biomass measured as chlorophylla concentration was significantly lower under drought conditions compared to the control and these differences were more remarkable after the application of the pesticides cocktail. Microbial respiration per unit of microbial (algal and bacterial) carbon was higher in biofilms subjected to droughts, which tended to accumulate lower extracellular polymeric substances (EPS) comparing to the immersed control. Droughts also modified the organic matter decomposition fingerprint of stream biofilms, independently from pesticides cocktail exposure. The diffusion of glyphosate and AMPA into biofilms was lower than that of tebuconazole, terbuthylazine and imidacloprid and this was explained by the differences in molecules’ hydrophobicity. However, long droughts tended to further decrease the diffusion of pesticides per unit of biofilm EPS in comparison the short droughts and control treatments.

International audience; Recent evidences hilighted the effect of urban effluent on the structure and functions of dowstream periphytic microbial communities associated to increased tolerance to further chemical stress. Nevertheless, the underlying (molecular) mechanisms of these responses remain poorly understood while usual descriptors provide a partial picture of the phenotype of the communities under chemical stress. In this context, the present study aims to gain knowledge about the molecular/biochemical responses of biofilms under urban stress through the implementation of untargeted metabolomics approach. To do so, following exposure in indoor channels connected to a urban effluent, the metabolomic responses of the biofilms were characterized in parallel of structural and functional responses. First, no significant effects were noted on the photosynthesis, respiration, primary/secondary production, as well as on biomass whereas exposure to the raw effluent led to tolerance acquisition to further chemical stress likely associated to significant shift in the microbial diversity. Then, the metabolomic profiles showed discrepancies between all the tested conditions demonstrating a clear effect of the effluent on the molecular phenotype of the biofilms. Moreover, the clear separation between raw and ultrafiltrated conditions highlighted the potential contribution of the microbes from the effluent in the response. Such pattern paralleled with microbial diversity one. The strong influence of the microbes was further confirmed through HCA showing that control and ultrafiltrated conditions were clustered together.Further trends analysis combined with pathways analysis revealed potential impact on porphyrin and chlorophyll metabolism, among others. Finally, statistical comparison between the metabolomic profiles of raw vs ultrafiltrated conditions highlighted that only 11% down- and 18% up-regulated signals contributed to the tolerance of the biofilm. Overall, this study demonstrates that the metabolomic response is more sensitive than usual descriptors through its ability to discriminate all the experimental conditions. Moreover, our results showed that microbes strongly contribute to these various molecular/biochemical responses, as well as to tolerance acquisition. Further investigations are ongoing to confirm the identity of the candidate metabolites and pathways and discriminate their link to urban microbes vs chemicals associated to the tolerance acquisition to chemical stress.

International audience; Facing increasing pollution of aquatic ecosystems, one major challenge is to better understand how organisms respond to chemical stress in order to gain knowledge on the resilience of these ecosystems and associated services. To this end, untargeted metabolomics is a method of choice which allows to provide a comprehensive picture of both exposure and effect. Also, its application to aquatic microbial communities, which play a key role in aquatic systems, could improve the ecological relevance of current ecotoxicological approaches through the discovery of ecosystemic biomarkers. Thus, we investigate the responses of aquatic biofilms to chemical stress by using high-resolution mass spectrometry based metabolomics. Our approach consists on the combined measurement of molecular, physiological and structural descriptors following exposure in controlled conditions and/or colonization/exposure on the field. Further chemometric methods help to identify metabolites and associated signalling pathways and link them to structural and functional changes. The specificity of these molecular makers regarding chemical stress and confounding factors can be then investigated through exposure under controlled conditions. Our research finally aims to gain knowledge on the molecular mechanisms involved in the alteration of key ecosystem functions and processes. Since urban activities represent a threat to the environment, we recently investigated the impact of wastewater effluents on microbial communities through colonization/exposure in channels connected to the effluents. Structural (microbial composition), physiological (photosynthesis, respiration) and metabolomic responses have been acquired and are under interpretation. Our approach and the results of this study will be presented.

International audience

International audience

International audience

Micropollutants (MPs) represent an important environmental and health issue. Identifying their sources and then reducing their discharges is the strategy used in France to fight against water pollution. This is also the approach implemented in the Regard project (Reduction and management of micropollutants in the Bordeaux metropolis). The first phase of the project aimed to carry out a territorial, global and integrated diagnosis combining both chemical and biological analyses of the natural environment and the sewerage network from the discharge points (wastewater treatment plant, separate stormwater overflow, by-pass) to the emission sources (domestic, industrial, hospital and stormwater). In addition, a social characterization of the sources was carried out in order to understand the practices, products and uses at the origin of MP discharges and to identify decisive actions to reduce these discharges. The strengths of this diagnostic assessment are the complementary of various approaches (engineering and social sciences, chemical and biological analyses, wastewater and stormwater studies) and the large number of sampling sites. The second phase of the project was to ensure the implementation of reduction actions to test and evaluate their impacts from the environmental (effectiveness in reducing the quantity, diversity and effect of MPs), social (appropriation and satisfaction of solutions) and economic (to guide public action) points of view. The best results have been achieved with (i) the “water families challenge” action on the domestic source, (ii) mechanical rat control actions, defoaming of tennis courts and grassing of cemeteries for the community source, and (iii) the treatment action of separate stormwater in real conditions on a pilot scale. This feedback on what has been achieved to date should help communities interested to implement such an approach in order to avoid the same “mistakes” and, on the contrary, to directly implement actions that give significant reduction results.; Les micropolluants (MP) représentent un enjeu environnemental et sanitaire important. Identifier leurs sources pour ensuite réduire leurs rejets est la stratégie privilégiée au niveau français pour lutter contre cette pollution. C’est aussi la démarche qui a été mise en oeuvre dans le projet Regard (réduction et gestion des micropolluants sur la métropole bordelaise). La première phase du projet correspondait ainsi à la réalisation d’un diagnostic territorial, global et intégré couplant à la fois des analyses chimiques et biologiques du milieu naturel et du réseau d’assainissement depuis les points de rejets (station de traitement des eaux usées, exutoires pluviaux, by-pass) jusqu’aux sources d’émission (domestique, industrielle, hospitalière et pluviale). En complément, une caractérisation sociale dessources a été faite afin de comprendre les pratiques, les produits et les usages à l’origine des rejets de MP et d’identifier des leviers d’action pour réduire ces rejets. Les points forts de ce diagnostic sont la complémentarité des approches (sciences de l’ingénieur et sciences sociales, analyses chimiques et biologiques, étude des eaux usées et pluviales) et le nombre important de sites d’étude. La seconde phase du projet correspondait à la mise en oeuvre d’actions de réduction pour les tester et les évaluer du point de vue environnemental (efficacité pour réduire la quantité, la diversité et l’effet des MP), social (appropriation et satisfaction vis-à-vis des solutions) et économique (aide à l’orientation de l’action publique). Les actions ayant eu les meilleurs résultats sont (i) l’action « Familles EAU Défi » sur la source domestique, (ii) les actions de dératisation mécanique, de démoussage des terrains de tennis et d’enherbement des cimetières pour la source collectivité et (iii) l’action de traitement des eaux pluviales strictes en conditions réelles à l’échelle d’un pilote. Le présent retour d’expérience sur ce qui a été fait doit aider les collectivités qui souhaiteraient effectuer une telle démarche à ne pas commettre les mêmes « erreurs » et, au contraire, à mettre en oeuvre directement les actions qui donnent des résultats satisfaisants.

International audience

International audience

Environmental risk assessment associated with aquatic and terrestrial contamination is mostly based on predicted or measured environmental concentrations of a limited list of chemicals in a restricted number of environmental compartments. High resolution mass spectrometry (HRMS) can provide a more comprehensive picture of exposure to harmful chemicals, particularly through the retrospective analysis of digitally stored HRMS data. Using this methodology, our study characterized the contamination of various environmental compartments including 154 surface water, 46 urban effluent, 67 sediment, 15 soil, 34 groundwater, 24 biofilm, 41 gammarid and 49 fish samples at 95 sites widely distributed over the Swiss Plateau. As a proof-of-concept, we focused our investigation on antifungal azoles, a class of chemicals of emerging concern due to their endocrine disrupting effects on aquatic organisms and humans. Our results demonstrated the occurrence of antifungal azoles and some of their (bio)transformation products in all the analyzed compartments (0.1–100 ng/L or ng/g d.w.). Comparison of actual and predicted concentrations showed the partial suitability of level 1 fugacity modelling in predicting the exposure to azoles. Risk quotient calculations additionally revealed risk of exposure especially if some of the investigated rivers and streams are used for drinking water production. The case study clearly shows that the retrospective analysis of HRMS/MS data can improve the current knowledge on exposure and the related risks to chemicals of emerging concern and can be effectively employed in the future for such purposes., Environment International, 139, ISSN:0160-4120, ISSN:1873-6750

Le récepteur humain Pregnane X Receptor (hPXR, NR1I2) est impliqué dans la biosynthèse, la distribution et le métabolisme de stéroïdes, d’acides biliaires et de xénobiotiques, parmi lesquels de nombreux pesticides. Quand des xénobiotiques se lient au hPXR, ce complexe forme un hétérodimère avec le récepteur X des rétinoïdes (RXR) et induit l’expression de gènes de détoxication comme les gènes codant pour les enzymes de phase I (CYP), de phase II (conjugaison) ou des protéines de transport (transporteurs de la famille ABC), résultant en une clairance des stéroïdes, des drogues et xénobiotiques par le foie et l’intestin (Timsit et al, 2007). L’activation de hPXR a été associée à des interactions médicamenteuses, la perturbation endocrinienne, la chimiorésistance, la croissance et l’agressivité de cancers du côlon et du foie (Planque et al, 2016 ; Wang et al, 2011 ; Kodama et al, 2015 ; Pondugula et al, 2016). Des études récentes ont révélé l’activité hPXR d’échantillons environnementaux de milieux aquatiques comme les effluents urbains et industriels, les eaux de surface et les sédiments de rivière (Creusot et al, 2011 ; Dagnino et al, 2012). Comparativement au PXR humain, il apparaît que le PXR de poisson zèbre (zfPXR) est activé par un plus faible nombre de pesticides (Ekins et al, 2008 ; Krasowski et al, 2011 ; Milnes et al, 2008) et que certains d’entre eux ont une meilleure affinité pour zfPXR que pour hPXR, suggérant d’importantes différences entre les deux récepteurs. De plus, les rôles du PXR dans le métabolisme du poisson ne sont pas encore pleinement élucidés. Dans ce contexte, nous avons cherché à caractériser les interactions entre les pesticides et le PXR chez l'homme et le poisson zèbre et à étudier le rôle de zfPXR dans le métabolisme du poisson. À cette fin, nous avons testé l'activité PXR d'échantillons environnementaux et de pesticides à l'aide de lignées cellulaires luminescentes exprimant soit hPXR soit zfPXR. Les pesticides les plus puissants ont ensuite été testés pour leur capacité à réguler l'expression des gènes cibles de PXR et à agir sur la prolifération cellulaire et la résistance aux médicaments dans ces cellules. Ces travaux seront combinés à des approches de modélisation (zfPXR) et de cristallographie (hPXR) pour étudier les interactions structurelles des pesticides avec hPXR et zfPXR, et à une approche in vivo utilisant le poisson zèbre pour étudier les effets des pesticides sur les gènes régulés par PXR. Dans l'ensemble, nos données permettent de mieux comprendre le rôle et les mécanismes d'action du PXR chez l'homme et le poisson zèbre et mettent en évidence des différences inter-espèces pour une meilleure évaluation des risques d'exposition aux pesticides pour la santé humaine et l'environnement.

In the context of contamination of aquatic ecosystems by endocrine disrupting chemicals (EDCs), this work aims to provide new insights on cross-species differences in the modulation of nuclear receptors (NRs) and the aryl hydrocarbon receptor (AhR) by individual chemicals and environmental mixtures, in order to help further cross-species extrapolation in the frame of the environmental risks of EDCs. To this end, a panel of individual ligands and environmental mixtures from an urban waste water treatment plant (WWTP), were screened on a set of recently developed in vitro reporter cell lines based on both human (h) and zebrafish (zf) NRs and AhR. Our results revealed that for some of the receptors marked cross-species differences occurred (PXR, PPARγ, PR) while for other receptors the differences were lower (ER, AR, GR, MR) or almost absent (AhR, ERRγ). For instance, promegestone acts as a full agonist of the hPR but as partial agonist of the zfPR whereas the dihydroxy-4-pregnen-3-one -reference ligand of the zfPR- antagonizes the hPR. In the same way, none of the reference ligands of the hPXR (T0913117) modulates the zfPXR whereas the clotrimazole -reference ligand of zfPXR- modulates also the hPXR but with lower potency. Then the hAR was more sensitive to the agonist mifepristone and the antagonist OH-flutamide than the zfAR whereas the dexamethasone was a more potent agonist of the zfGR than the hGR. Also significant differences in selectivity were noted among h and zf ER subtypes. Finally, the in vitro profiling of an urban WWTP confirmed these cross-species differences in terms of level, type (agonist vs antagonist), distribution along the WWTP. For instance, h and zf estrogenic activity was differentially detected in the sludge and the suspended material. In the same way, strong zf anti-androgenic activity was detected in the effluent while no human one can be detected. Also, strong zf mineralocorticoidic activity was detected in both influent and effluent whereas only h anti-mineralocorticoid activity was detected. Alogether, our results showed that h and zf NRs are, for some of them, differentially modulated by individual chemicals and environmental mixtures. Also, interaction of EDCs towards NRs cannot always be extrapolated between these species highlighting the need to further document NRs modulation between human and fish and associated responses, to improve human health and environmental risk assessment of EDCs.

National audience

National audience

Wastewater treatment plant (WWTP) effluents are major sources of endocrine-disrupting chemicals (EDCs) and other chemicals of toxicological concern for the aquatic environment. In the present study, we used an integrated strategy combining passive sampling (Chemcatcher®), developmental toxicity, and mechanism-based in vitro and in vivo bioassays to monitor the impacts of a WWTP on a river. In vitro screening revealed the WWTP effluent as a source of estrogen, glucocorticoid, and aryl hydrocarbon (AhR) receptor-mediated activities impacting the downstream river site where significant activities were also measured, albeit to a lesser extent than in the effluent. Effect-directed analysis of the effluent successfully identified the presence of potent estrogens (estrone, 17α-ethinylestradiol, and 17β-estradiol) and glucocorticoids (clobetasol propionate and fluticasone propionate) as the major contributors to the observed in vitro activities, even though other unidentified active chemicals were likely present. The impact of the WWTP was also assessed using zebrafish embryo assays, highlighting its ability to induce estrogenic response through up-regulation of the aromatase promoter-dependent reporter gene in the transgenic (cyp19a1b–green fluorescent protein [GFP]) zebrafish assay and to generate teratogenic effects at nonlethal concentrations in the zebrafish embryo toxicity test. The present study argues for the use of such an integrated approach, combining passive sampling, bioassays, and effect-directed analysis, to comprehensively identify endocrine active compounds and associated hazards of WTTP effluents.