The influence of temperature rise on the metabolic response of Ruditapes philippinarum clams to 17-α-ethinylestradiol


Untargeted Nuclear Magnetic Resonance metabolomics was employed to study the effects of warming conditions (17–21 °C) and exposure to 17-α-ethinylestradiol (EE2) on the polar metabolome of Ruditapes philippinarum clams, to identify metabolic markers for monitoring/prediction of deviant environmental conditions. Warming alone triggered changes in alanine/aspartate/glutamate, aromatic amino acids, taurine/hypotaurine and homarine/trigonelline pathways, as well as in energy metabolism, suggesting osmoregulatory adaptations and glycolytic/tricarboxylic acid (TCA) cycle activation, possibly accompanied to some extent by gluconeogenesis to preserve glycogen reserves. At 17 °C, the lowest EE2 concentration (5 ng/L) specifically engaged branched-chain and aromatic amino acids to activate the glycolysis/TCA cycle. Notably, a partial metabolic recovery was observed at 25 ng/L, whereas higher EE2 concentrations (125 and 625 ng/L) again induced significant metabolic disturbances. These included enhanced glycogen biosynthesis and increased lipid reserves, sustained by low-level glutathione-based antioxidative mechanisms that seemed active. At 21 °C, response to EE2 was notably weak at low/intermediate concentrations, becoming particularly significant at the highest EE2 concentration (625 ng/L), suggesting higher protection capacity of Ruditapes philippinarum clams under warming conditions. At 625 ng/L, disturbances in alanine/aspartate/glutamate and taurine/hypotaurine metabolisms were observed, with no evidence of enhanced carbohydrate/protein catabolism. This low energy function profile was accompanied by marked antioxidative mechanisms and choline compounds modulation for cell membrane protection/repair. These results help monitor clams´ response to temperature rise and EE2 exposure, paving the way for future effective guidance and prediction of environmental damaging effects.


Joao A. Rodrigues, Mónica Silva, Rita Araújo, Leonor Madureira, Amadeu M.V.M. Soares, Rosa Freitas, Ana M. Gil

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This work was developed within the CICECO-Aveiro Institute of Materials project (UIDB/50011/2020, UIDP/50011/2020 & LA/P/0006/2020) financed by national funds through the FCT/MCTES (PIDDAC). We are also grateful to the Portuguese National NMR Network (PTNMR), supported by FCT funds as the NMR spectrometer used is part of PTNMR and partially supported by Infrastructure Project No. 022161 (co-financed by FEDER through COMPETE 2020, POCI and PORL, and the FCT through PIDDAC). This work was also financially supported by the project BISPECIAl: BIvalveS under Polluted Environment and ClImate chAnge (POCI-01-0145-FEDER-028425) funded by FEDER, through COMPETE2020 - Programa Operacional Competitividade e Internacionalização (POCI), and by national funds (OE), through FCT/MCTES. Mónica G. Silva benefited from Research Grant (MSc) (BI/CESAM/0043_2019/POCI-01-0145-FEDER-028425) under the project BISPECIAl: BIvalveS under Polluted Environment and ClImate change PTDC/CTA-AMB/28425/2017 (POCI-01-0145-FEDER-028425).

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