The synergistic effects of chronic gas condensate pollution and warming on the survival, performance and reproduction of the mussel Brachidontes pharaonis

The individual and combined effects of chronic gas condensate pollution and warming of 3.1 °C on the survival, physiological and pathohistological responses of the mussel Brachidontes pharaonis and the mono/poly cyclic aromatic hydrocarbons (M/PAHs) bioconcentration patterns in the mussels' tissues. Clearance rate was measured by incubation of mussels (n=3 from each treatment) in seperate chambers, with the microalgae Tetraselmis tetrathele. Three chambers without mussels were used as control. At the end of the incubation, algal cells were counted under a microscope using a hemocytometer. Clearance rate was standardized to wet weight and shell length and calculated. Respiration rate was measured in seperate chamber of each mussel at the beginning and after one hour using a handheld sensor. Control jars with no mussels were used to account for microbial respiration. Respiration rate (µmol O2 h-1 g WW-1) was calculated per weight and per length. Two-way ANOVA and Tukey post-hoc were used to test the effect of gas condensate concentrations and temperature and their combination on respiration rates and clearance rates using with R v. 4.2.2 using the package “stats” (R Core Team, 2021). Tissue PAHs: triplicate tissue samples were pooled and extracted using a Speed Extractor E-916 (Büchi). Gas chromatography-mass spectrometry (GC/MS, Agilent 6890/5973) was used to identify and quantify the concentrations of each MAH and PAH. Water PAHs: the water was extracted using a solid-phase extraction (SPE) and the samples were analyzed using an Agilent GC/MS (6890/5973) against external selected MAHs standard and 16 PAHs standard Temperature logs: Temperatures in the ambient and warming treatments were monitored and logged hourly during the whole experimental period using temperature loggers (EnvLogger, Electric Blue, Portugal).