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Carbamazepine (CBZ), an antiepileptic drug, is one of the most commonly detected pharmaceutical drugs in aquatic ecosystems, and is used as a marker of urban pollution. Since CBZ is designed to exert a biological effect, when it reaches aquatic environment high probability exist for toxic effects on non-target organisms. The present study evaluated the acute toxicity of environmentally relevant concentrations of CBZ (0.00, 0.03, 0.30, 3.00, 9.00 µg/L) in the edible clams Venerupis decussata (a native species) and Venerupis philippinarum (an invasive species) collected from the Ria de Aveiro. The effects on both species were assessed through the use of a battery of biomarkers mainly related with health status and oxidative stress. Furthermore, in this work an alternative and promising tool, the direct competitive immunoassay ELISA, for the direct CBZ quantification in clam's tissues, was applied.
The results of the present work showed that CBZ in clam's tissues increased with the exposure concentration and V. decussata gave slightly higher values than V. philippinarum. Although the clams accumulated lower levels of CBZ than the concentration of exposure, these concentrations were enough to impair the health status and induce oxidative stress. However, a different response to CBZ was observed in the two species. While in V. philippinarum the lipid peroxidation levels increased at the highest CBZ concentration (9.00 µg/L), in V. decussata a significant decrease was seen. Moreover, glutathionse S-transferase activity was stimulated in V. decussata and decreased in V. philippinarum. Nevertheless, an induction of glutathione reductase, superoxide dismutase and cytochrome P450 3A4 activities was found in both species as a result of the exposure. The results indicate that, probably, V. philippinarum have a less efficient antioxidant system than V. decussata, and are therefore less capable to neutralize oxidative stress and consequently more sensitive to CBZ. The risk quotient determined for the Ria de Aveiro was higher than 1 indicating that a ecotoxicological risk is suspected. Furthermore, bioaccumulation of CBZ in clams should be taken into consideration since this chemical might be transferred along the food chain and affect non-target organisms.
Several works evaluated the toxicity of pharmaceutical drugs and climate related changes in invertebrates but few explored the combined effects of both stressors, namely considering their mode of action (MoA). Carbamazepine (CBZ) and cetirizine (CTZ) are pharmaceutical drugs detected in the environment and the toxicity derived from the combined effects of these drugs with ocean acidification (OA) is poorly explored. Thus, the present study investigated the biochemical parameters related to an oxidative stress response and the transcription of genes related to the MoA of CBZ (1.0 mg/L) and CTZ (0.6 mg/L) in the clam Ruditapes philippinarum chronically exposed (28 days) to control (7.8) and low (7.5) pH conditions. The results obtained showed that despite the clams accumulated both drugs, at low pH the clams exposed to CTZ decreased drug concentration and BCF values (CTZ uptake: 2.0 ± 0.5 ng/g fresh weight; BCF: 3.8 ± 0.9) in comparison with clams exposed to control pH (CTZ uptake: 2.9 ± 0.3 ng/g fresh weight; BCF: 5.5 ± 0.6). No oxidative stress was induced by the exposure to CBZ or CTZ at each pH level, but the transcription of several genes related with the MoA (neurotransmission, immunity and biomineralization) was altered by low pH, drug exposure and the combination of both stressors. At both pH conditions, CBZ increased the transcription of GABA receptor gene (neurotransmission) and CTZ led to a decrease of Perlucin gene (biomineralization) transcription. The transcription of MyD88 gene (immunity) decreased at low pH (7.5) combined with drug exposure (CBZ or CTZ). Thus, it was highlighted that the interaction of drug exposure and low pH conditions can change bivalves’ sensitivity to drugs or alter drugs toxicity.
In the aquatic environment, organisms are exposed to complex mixtures of contaminants which may alter the toxicity profile of each compound, compared to its toxicity alone. Pharmaceutical drugs (e.g. carbamazepine (CBZ) and cetirizine (CTZ)) and metals (e.g. cadmium (Cd)) are among those contaminants that co-occur in the environment. However, most studies concerning their toxicity towards aquatic species are based on single exposure experiments. Thus, the present study aimed to evaluate single and combined effects of Cd and CBZ or CTZ (single conditions: Cd, CTZ, CBZ; combined conditions: CTZ+Cd, CBZ+Cd) on biomarkers related to oxidative stress and energy metabolism in the edible clam Ruditapes philippinarum, by exposing the organisms for 28 days to environmentally relevant concentrations of these contaminants. The biomarkers studied were: i) the electron transport system activity, protein and glycogen contents (indicators of organisms’ metabolic status and energy reserves); ii) lipid peroxidation and the ratio between reduced and oxidized glutathione (indicators of oxidative stress); iii) superoxide dismutase and catalase activities (enzymes indicators of antioxidant defence) and iv) activity of glutathione S-transferases (family of enzymes indicators of biotransformation capacity). Results obtained showed that the uptake of Cd and CBZ was not affected by the combined presence of the contaminants.
However, for CTZ, the uptake was higher in the presence than in the absence of Cd. Concerning toxicity data, in general, the combined exposures (CTZ+Cd, CBZ+Cd) had lower biological effects than the contaminants alone. Nevertheless, our data showed that despite the low concentrations tested, they were enough to exert biological effects that differed between single and combined treatments, evidencing the need to conduct more co-exposure studies to increase the environmental relevance of the gathered data.