Oxidative stress and immune signatures of carbamazepine in freshwater mussels.

Abstract

The persistence of pharmaceutical compounds in aquatic environments and their limited removal by conventional treatment processes necessitate evaluating organism-level effects of long-term exposure using biomarkers. To determine how carbamazepine (CBZ), commonly detected in environmental waters, modulates immune and oxidative stress responses in the freshwater mussel Unio delicatus, individuals (n = 160) were exposed to sublethal CBZ concentrations for 21 and 45 days under semi-static conditions. Exposure concentrations were set at 199 µg/L (D1), 995 µg/L (D2), and 1.99 mg/L (D3), corresponding to 1%, 5%, and 10% of the LC₅₀ (19.9 mg/L). The experimental environment was conducted under controlled physicochemical conditions, and the water was periodically replaced to maintain target concentrations. Total haemocyte counts (THCs) were determined from haemolymph tissue; malondialdehyde (MDA) and reduced glutathione (GSH) levels were measured in gill and digestive gland tissues. In addition, GSH and MDA data were integrated, and the total stress load was assessed using the Integrated Biomarker Response v2 (IBRv2) index. CBZ exposure led to significant increases in THCs, particularly at low doses, and this response became more pronounced after 45 days of exposure. Oxidative stress markers showed a tissue- and time-dependent relationship: MDA tended to increase in the gills during the early period, while it generally decreased with long-term exposure; an increase was observed in the digestive gland at high doses during the early period. GSH levels decreased with short-term exposure, but increased after 45 days of exposure, especially in the digestive gland. IBRv2 revealed that the total stress load increased at high doses, reaching 4.19 and 4.80 at 21 and 45 days, respectively, in the D3 group. In conclusion, CBZ regulates the immune system and redox balance in U. delicatus in a tissue, dose, and time-dependent manner. However, the decrease in MDA and the increase in GSH observed in some tissues after prolonged exposure should be interpreted cautiously as possible compensatory redox responses rather than definitive evidence of adaptation, since the elevated IBRv2 values indicate that the overall stress burden persisted.