Use of an integrated biomarker-based strategy to evaluate physiological stress responses induced by environmental concentrations of caffeine in the Mediterranean mussel Mytilus galloprovincialis

The occurrence of caffeine (CF), a biologically active drug, has widely been documented in coastal waters, and whether its environmental concentrations do represent a threat for marine organisms is unclear. The present study aimed at assessing sub-lethal effects induced by a 7-day exposure to environmentally relevant concentrations of CF (5, 50 and 500 ng/L) in the Mediterranean mussel, Mytilus galloprovincialis. CF in water and mussel tissues, and a battery of biomarkers, including lysosomal parameters of general stress, oxidative stress responses and endpoints of neurological and genetic damages, were evaluated and tested for significance vs controls (p < 0.05). CF exposure triggered a significant decrease of lysosomal membrane stability in both haemocytes and digestive gland (at 50 and 500 ng/L CF) and a significant increase of lysosomal content of neutral lipids (at 500 ng/L CF), indicating the onset of a stress syndrome. No effects were noted on lipid peroxidation parameters, such as malondialdehyde and lipofuscin content. The activity of the antioxidant enzymes glutathione S-transferase (GST) and catalase was unmodified in gills, while a significant increase of GST activity was observed in digestive gland (at 5 and 500 ng/L CF), suggesting the occurrence of GST-mediated phase II detoxifying processes. CF did not induce geno/neurotoxicity, as shown by the lack of effects on primary DNA damages and acetylcholinesterase activity. In line with its high hydrophilicity, CF did not bioaccumulate in mussel tissues. Data were integrated using the Mussel Expert System, which assigned a low stress level to mussels exposed to 500 ng/L CF, whereas no alterations of animal health status were highlighted at lower dosages. This study revealed a low profile of toxicity for environmental concentrations of CF, and confirmed the suitability of an integrated biomarker-based approach to provide a comprehensive picture of the degree of stress induced by emerging contaminants in marine invertebrates.