Co-exposure to nano-TiO2 and carbamazepine induces multidimensional toxicity in mussels, from physiological responses to gut microbiome dysbiosis

Pharmaceuticals and personal care products represent a diverse class of globally pervasive emerging contaminants, raising significant concerns due to their environmental persistence and potential health risks. However, ecotoxicological risks posed by engineered nanomaterial nano-titanium dioxide (nano-TiO 2 ) and the recalcitrant pharmaceutical carbamazepine (CBZ), both widely detected micropollutants in coastal environments, are still poorly understood. This study employed the ecologically and economically important mussel Mytilus coruscus as a bioindicator to assess the physiological and biochemical response to individual and combined exposure to nano-TiO 2 (100 μg/L) and CBZ (10 μg/L) for 30 days. Impacts were evaluated across multiple tissues (gill, foot, digestive gland, mantle), and both contaminants accumulated in the digestive gland. Individual and combined exposures triggered antioxidant defense responses and neurotoxicity, concurrently disrupting key physiological functions including feeding, digestion, and nutrient reserves, while metagenomic analysis demonstrated that exposure disrupted the intestinal microbiome. Specifically, exposure induced intestinal microbiota dysbiosis, characterized by reduced alpha diversity, altered community structure, and an increased abundance of potentially pathogenic bacteria. Furthermore, exposure stimulated KEGG pathways within the gut microbiome associated with environmental adaptation, immune function, and energy metabolism. These findings reveal the potential role of the immune–neuroendocrine system, digestive/energy metabolic pathways, and gut microbiota in bivalve stress responses to environmental contaminants, providing deeper insights into the mechanisms underlying multi-pollutant toxicity.