Interaction between cadmium and carbendazim in soil ecosystems: Impacts on earthworm health and biomarker responses

Cadmium (Cd) and carbendazim (CBZ) are widespread soil contaminants that frequently co-occur in agricultural environments, yet their combined toxicity to terrestrial invertebrates remains insufficiently understood. This study investigated the individual and combined effects of Cd (5-90 mg/g) and CBZ (7-126 mg/g) on Eisenia fetida using multiple biomarkers, including mortality, oxidative stress indicators (CAT, SOD, TAC, and MDA), total protein content, weight loss, sperm parameters (TUNEL-positive cells, sperm count, and sperm deformation), and subcellular Cd partitioning. Sperm DNA fragmentation increased in a dose-dependent manner under both single and combined exposures, reaching a maximum increase of 719% under mixture exposure, while sperm count was significantly reduced, particularly in the combined treatment. Cadmium was primarily localized in the cytosolic fraction, followed by debris and granules, under both single and combined exposures. Carbendazim exposure induced minor but measurable (6-7%) changes in Cd intracellular distribution. Under single exposure, cytosolic Cd and total body burden were significant predictors of most biomarker responses. In contrast, under combined exposure, these correlations were weaker and limited. Cadmium and mixture exposures significantly altered CAT, SOD, and TAC activities, with sperm-related parameters emerging as the most sensitive toxicity endpoints. The biomarker response index indicated severe adverse effects on earthworm health, while the Effect Addition Index demonstrated antagonistic interactions under combined exposure. Overall, reproductive and oxidative stress biomarkers were the most responsive endpoints, highlighting the importance of integrative biomarker approaches for assessing mixture toxicity in soil ecosystems.