Effect-directed analysis (EDA) aims at identifying the compound(s) responsible for toxicity in a complex environmental sample where several dozens of contaminants can be present. In this study, we used an environmental mixture extracted from the Polar Organic Chemical Integrative Sampler (POCIS) previously immersed downstream a landfill (River Ponteils, South West France), to perform an EDA approach using a microalgal bioassay based on the photosynthetic capacities of diatom (Nitzschia palea) cultures. Adverse effects on photosynthetic capacities were recorded when algae were exposed to the entire POCIS extract (> 85% inhibition at the highest concentration tested). This result was coherent with the detection of diuron and isoproturon, which were the 2 most concentrated herbicides in the extract. However, the EDA process did not allow pointing out the specific compound(s) responsible for the observed toxicity but rather suggested that multiple compounds were involved in the overall toxicity and caused mixture effects.

In this study, we examined the dose-dependent effects of an environmentally relevant pesticide cocktail (metalachlor, linuron, isoproturon, tebucanazole, aclonifen, atrazine, pendimethalin, and azinphos-methyl) and temperature change (22 vs. 32 °C for 4-week exposure) on Na⁺/K⁺-ATPase, 3-nitrotyrosine protein (NTP), dinitrophenyl protein (DNP), catalase (CAT), and superoxide dismutase (SOD) expressions in gills of goldfish (Carassius auratus). Histopathological analysis showed widespread damage to gill in elevated temperature (32 °C) and pesticide co-exposure groups, including fusion of secondary lamellae, club-shaped primary lamellae, rupture of epithelial layer, loss of normal architecture, and hemorrhaging. Immunohistochemical and qRT-PCR analyses showed significant decreases in Na⁺/K⁺-ATPase protein and mRNA expressions in gills exposed to higher temperature and pesticides; however, combined exposure to heat and pesticides significantly increases NTP, DNP, CAT, and SOD expressions. In situ TUNEL assay revealed elevated levels of apoptotic cells in response to combined exposure. Collectively, our results suggest the combined effects of heat and pesticide stress cause cellular damage, upregulate oxidative/nitrative stress biomarkers, and increase apoptotic cells, downregulate Na⁺/K⁺-ATPase expression in gills. This provides new evidence for oxidant/antioxidant-dependent mechanisms for downregulation of Na⁺/K⁺-ATPase expression in gills during combined exposure.

Pesticides are still widely used by agriculture, leading to the exposure of surface water. This may be the case for fish ponds located in farmland landscapes. To address this issue, the present study investigated the contamination by pesticides of fish ponds located in the mixed agriculture-pond landscape of the Dombes area, France. Ten ponds were selected in water catchments with a gradient of 3–57 ha of cropland with maize and winter cereals as the dominant crops. A total of 197 water samples were collected in the ponds during the fish production season over 3 years. Recently used pesticides were the most frequent residues occurring. Occurrences greater than 0.1 µgL⁻¹ particularly concerned chlorotoluron and S-metolachlor. Maximum observed concentrations were slightly above 3 µgL⁻¹ for S-metolachlor, acetochlor, and dimethenamide, all herbicides allowed for maize cultivation. Isoproturon and chlorotoluron, herbicides allowed in cereal crops, reached up to 1.2 and 1.0 µgL⁻¹, respectively. We found a significant positive effect of crop area in catchments on the pond contamination frequency by pesticides and more significantly on the contamination frequency by broad-spectrum herbicides (glyphosate and AMPA residues). The cumulative antecedent rainfall was best correlated to the frequency of highest contaminations (> 0.5 µgL⁻¹). In such a hydrological context, the crop area within catchment was identified as a good indicator of fish pond exposure to pesticide residues. Finally, we proposed to adapt some mitigation measures to reduce fish pond contamination.