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Marine organisms are constantly exposed to stress, such as changes in seawater temperature, which may elicit biochemical, molecular, and physiological changes. In the present study, an integrative approach to evaluate stress in the green-lipped mussel Perna canaliculus exposed to control, moderate, and severe heat stress was used. Flow cytometry (FCM) was used to measure hemolymph parameters, including hemocyte concentration, viability, and reactive oxygen species (ROS) production. Spectrophotometry and a handheld electrochemical meter were used to determine the total antioxidant capacity (TAC) of hemolymph and gill tissue, and metabolomics was used to contrast specific metabolic responses among treatments. For hemolymph parameters measured using FCM, the number of nonviable hemocytes increased with the severity of heat shock. The proportion of superoxide-positive hemocytes mirrored the increase in nonviability; by contrast, the general ROS production initially increased following moderate heat shock, but then decreased after severe heat shock. Trolox-equivalent antioxidant capacity in the gill tissue and fast-acting antioxidant levels in the haemolymph showed small but significant decreases following heat shock, whereas the reciprocal pattern was seen for slow-acting and TAC in the hemolymph. Metabolomic profiles discriminated between treatments, with energy metabolism pathways appearing the most affected. The approach used in this study provides a deeper mechanistic understanding of the stress responses of organisms exposed to heat stress; however, these analytical techniques may be combined or used independently to measure stress in marine organisms in the laboratory or in the field, which can improve health assessments of wild and farmed aquatic organisms. KEY WORDS: mussel, Perna canaliculus, hemocytes, ROS, antioxidant capacity, flow cytometry, electrochemistry, redox, metabolites, biomarker, aquaculture, climate change, INTRODUCTION Marine invertebrates are constantly being challenged by changes in environmental conditions and other stressors such as changes in seawater temperature, salinity, and pH, which will only become more prevalent [...]

The 2010 explosion of the Deepwater Horizon (DWH) oil rig generated the largest marine oil spill in US history with millions of barrels of crude oil released in the Gulf of Mexico (GoM). The eastern oyster, Crassostrea virginica, is an ecologically and economically important species in the northern GoM. Due to its biological characteristics (sessile, filter feeding), juvenile oysters may have been affected. This study investigated the effects of surface-collected DWH oil prepared as high-energy water-accommodated fraction (HEWAF) on the survival of 2-month-old oyster spat, and evaluated the potential impacts of HEWAF on particle clearance rate and spat tissue. Exposure of oysters to a range of oil/HEWAF (0-7-66-147-908-3450 μg tPAH50 (sum of 50 polycyclic aromatic hydrocarbons) L -1 ) resulted in non-dose-dependent mortalities and reduced clearance rates of algal food (Tisochrysis lutea). A morphometric study of the digestive tubules (DGTs) indicated a dose-dependent response to oil exposure on lumen dilation, on epithelium thinning of the DGT, and a significant change in DGT synchrony (LOEC = 66 μg tPAH50 L -1 ). This finding suggests that structural changes occurred in the digestive gland of exposed oysters most likely due to an oil-related stress. In addition, histological observations showed that tissues in contact with HEWAF (gills, palp, connective tissue, digestive gland) were adversely impacted at ≥ 7 μg tPAH50 L -1 , and exhibited pathological symptoms typical of an inflammatory response (e.g., hemocyte diapedesis and infiltration, syncytia, epithelium sloughing).