Antioxidant enzyme responses of Mytilus galloprovincialis to olive mill wastewater and Vibrio harveyi co-exposure under static and flow-through conditions.

Coastal marine organisms are increasingly exposed to co-occurring chemical effluents and opportunistic pathogens; however, the mechanisms governing their combined toxicological effects remain insufficiently resolved. This study evaluated the combined effects of olive mill wastewater (OMW) and Vibrio harveyi on Mytilus galloprovincialis, with a focus on antioxidant enzyme responses, microbial dynamics, and environmental conditions under static and flow-through exposure regimes. Mussels were exposed to sublethal OMW concentrations in the presence and absence of V. harveyi, and tissue-specific activities of the antioxidant enzymes, namely superoxide dismutase (SOD), catalase (CAT), and glutathione S-transferase (GST), were assessed alongside bacterial abundance, dissolved oxygen concentration (DO), and pH. Under static conditions, V. harveyi persisted throughout the exposure period (4.08-4.48 log CFU/g at 48 h), coinciding with reduced DO (3.4 mg/L) and sustained elevation of antioxidant enzyme activities. In contrast, flow-through conditions promoted rapid bacterial elimination (below detection limits within 3-12 h) and stabilised environmental parameters (DO >5 mg/L), resulting in reduced antioxidant enzyme responses. OMW alone induced concentration-dependent changes in antioxidant enzyme activities, with gill SOD activity reaching 135.28 ± 8.10 U/mg protein at 5% OMW. Notably, under co-exposure conditions, 1% OMW was associated with lower antioxidant enzyme activities compared with V. harveyi exposure alone. Multivariate analyses revealed clear associations between antioxidant enzyme activities, microbial persistence, and dissolved oxygen concentration dynamics across exposure regimes. These findings indicate that short-term antioxidant enzyme response patterns differed between static and flow-through exposure regimes and highlight the importance of environmental context when interpreting biochemical responses in marine bivalves.