Exposure of freshwater gastropods to chlorpyrifos and cadmium: Predictable and unpredictable responses

Aquatic environments near agricultural areas are increasingly contaminated by mixtures of pesticides and heavy metals, posing a threat to non-target aquatic invertebrates. Because mixtures may induce unexpected biological responses, their study is essential for realistic environmental risk assessment. This study investigated the acute and chronic effects of the pesticide chlorpyrifos (CPF), cadmium (Cd), and their mixture on the freshwater gastropods Biomphalaria glabrata and Planorbarius corneus under controlled laboratory conditions. A panel of biochemical and cellular biomarkers was used to evaluate neurotoxicity, oxidative stress, metabolic alterations and immunotoxicity. Acute CPF exposure, alone or combined with Cd, inhibited B-esterase activity and increased TBARS, indicating neurotoxic and pro-oxidant effects. Glutathione-S-transferase (GST) and catalase (CAT) remained unaltered. Hemolymph glucose levels increased with all treatments, with a synergistic hyperglycemic response to CPF + Cd observed only in B. glabrata. Cd exposure induced hypercholesterolemia in B. glabrata and hypertriglyceridemia in both species. Hemocyte revealed enhanced reactivity in response to contaminants alone, an effect absent in the mixture. Notably, under chronic CPF + Cd exposure, a pronounced hemocyte clumping occurred, an immunotoxic response, not previously reported. These findings highlight B-esterase inhibition and lipid peroxidation as sensitive biomarkers of acute CPF exposure; while hemolymph metabolic parameters may serve as complementary indicators of acute Cd exposure. GST and TBARS levels emerged as informative indicators of chronic Cd toxicity. Notably, the contaminant mixture unpredictably suppressed or amplified only certain biological responses, particularly on immune cells. This study underscore the value of integrative biomarker approaches to better predict ecological risks associated with contaminant mixtures.