Microplastics (MP) are contaminants able to cause adverse effects on organisms. MPs are capable to interact with other environmental contaminants, including pesticides, altering their toxicity. The objective of the study was to research the sublethal effects (enzymatic activity) of pesticides alone and in combination with MPs. Cholinesterase enzymes are used as biomarkers to determine and evaluate the effects produced in organisms after exposure to pollutants. This study showed the acetylcholinesterase (AChE) enzymatic activity in the tissue of Solea senegalensis exposed to two environmental pollutants, the insecticide chlorpyrifos (CPF) and antibacterial triclosan (TCS) with and without microplastics (MPs). Solea senegalensis was chosen because it is a species in high demand because of its high economic value in southern Europe, as well as the use of this species in ecotoxicology and its increasing use as sentinel species, which justify using it to assess biological effects of pollutants. Toxicity tests were performed in organisms exposed to concentrations of between 5 and 80 mu g/L CPF and 0.1 and 0.4 mg/L TCS for 96 h. In addition, each test incorporated MPs that were added at different concentrations in order to evaluate their role as a possible enhancer of the effects caused by the previous pollutants. In the case of CPF, the head and muscle tissue cholinesterase activity was inhibited from a concentration of 5 mu g/L both without and with MPs, and the AChE enzymatic activity for the treatment with MPs was approximately half the activity for the treatment without MPs in the tissues studied. Besides, TCS inhibited the cholinesterase activity at a concentration of 0.3 mg/L in the muscle of S. senegalensis. In contrast, no significant differences were observed in the TCS + MP treatment compared to the controls. These results showed the importance of studies in assessing the anticholinesterase effects of pesticides combined with microplastics due to the abundance of these contaminants in the marine environment and the role of cholinesterase activity (biomarker) in the neurotransmission of key physiological processes.

A broad-spectrum fungicide, dichlofluanid is widely used in antifouling paints and agricultural pesticides. In this study, the acute and chronic effects of sublethal concentrations, namely, no observable effect concentration (NOEC) and 50% lethal concentration (LC₅₀) of dichlofluanid (1/10 NOEC, NOEC, 1/10 LC₅₀, and LC₅₀) were evaluated on the marine mysid, Neomysis awatschensis. Acute toxicity test (96 h) showed higher sensitivity to dichlofluanid in juvenile mysids (LC₅₀ 3.1 μg L⁻¹) than adults (LC₅₀ 24.5 μg L⁻¹), with lower survival rate and reduction in food consumption. Exposure with dichlofluanid considerably induced oxidative imbalance, as NOEC (0.006 μg L⁻¹ for juveniles and 0.074 μg L⁻¹ for adults) and 1/10 LC₅₀ values increased intracellular concentrations of malondialdehyde and glutathione, and the enzymatic activities of catalase and superoxide dismutase, whereas exposure to LC₅₀ value decreased the values of oxidative parameters. Enzymatic activity of acetylcholinesterase decreased considerably when exposed to LC₅₀ value. In the case of chronic effects, exposure to NOEC for 4 weeks markedly decreased the juvenile survival rate, while adults showed tolerance. Multigenerational monitoring in response to NOEC showed a significant growth retardation with an increase in intermolt duration as well as a decrease in the number of newborn mysids from females of the third generation. Consistent exposure to environmentally relevant sublethal concentrations of dichlofluanid would be detrimental to mysid individuals and the survival of the mysid population.

The effect of low concentration Cd stress on bivalves is unclear. In this study, Asian clams (Corbicula fluminea) were continuously exposed to 0, 0.05, 0.10, and 0.20 mg/L Cd for 14 d (exposure phase) and to artificial freshwater for 7 d (depuration phase). A total of 16 variables were measured to explore the toxic effects on C. fluminea. All physiological characteristics were significantly inhibited in the treatments (p < 0.05), and the negative effects of Cd did not return to normal levels in the short term. Tissue damage was found in the feet and gills of C. fluminea in all the treatments. On the 7th day (D7), enzyme activity in all the treatments was significantly higher (p < 0.05) than in the control group. Acetylcholinesterase, superoxide dismutase, and catalase activities were enhanced on D14 in all the treatments. However, only glutathione S-transferase activity was significantly higher in all the treatments (p < 0.05) than in the control group on D21. The instability of the enzymes indicated that the adaptability of C. fluminea became stronger throughout the experiment. In each group, the maximum bioaccumulation of Cd followed the order: 0.20 mg/L > 0.05 mg/L > 0.10 mg/L, which might be caused by the filtration capacity of C. fluminea in the 0.05-mg/L group, which was higher than that of the 0.10-mg/L group. Thus, low Cd concentrations effect the physiological characteristics, tissue health, and antioxidant system of C. fluminea and may require a long recovery time to be restored to normal levels.

Although many polymers are known by their toxicity, we know nothing about the impact of polyethylene glycol (PEG) on anurofauna. Its presence in different products and disposal in aquatic environments turn assessments about its impact on amphibians an urgent matter. Accordingly, we tested the hypothesis that short-time exposure (72 h) of tadpoles belonging to the species Physalaemus cuvieri (Anura, Leptodactylidae) to PEG induces oxidative stress and neurotoxicity on them. We observed that polymer uptake in P. cuvieri occurred after exposure to 5 and 10 mg/L of PEG without inducing changes in their nitrite levels neither at the levels of substances reactive to thiobarbituric acid. However, hydrogen peroxide and reactive oxygen species production was higher in animals exposed to PEG, whose catalase and superoxide dismutase levels were not enough to counterbalance the production of these reactive species. Therefore, this finding suggests physiological changes altering REDOX homeostasis into oxidative stress. In addition, the increased activity of acetylcholinesterase and butyrylcholinesterase, and reduction in superficial neuromasts, confirmed PEG’s neurotoxic potential. To the best of our knowledge, this is the first report on PEG’s biological impact on a particular amphibian species. The study has broadened the understanding about ecotoxicological risks associated with water pollution by these polymers, as well as motivated further investigations on its impacts on amphibians’ health and on the dynamics of their natural populations.

Moxidectin is an antiparasitic drug belonging to the class of the macrocyclic lactones, subgroup mylbemicins. It is used worldwide in veterinary practice, but little is known about its potential environmental risks. Thus, we used the zebrafish embryo as a model system to study the potential effects of moxidectin on aquatic non-target organisms. The analyses were performed in two experimental sets: (1) acute toxicity and apical endpoints were characterized, with biomarker assays providing information on the activity levels of catalase (CAT), glutathione S-transferase (GST), lactate dehydrogenase (LDH), and acetylcholinesterase (AChE); and (2) internal concentration and spatial distribution of moxidectin were determined using ultraperformance liquid chromatography quadrupole-time-of-flight mass spectrometry (UPLC-QToF-MS) and matrix-assisted laser desorption/ionization-MS imaging (MALDI-MSi). The acute toxicity to zebrafish embryos (96 hpf) appeared mainly as a decrease in hatching rates (EC₅₀ = 20.75 μg/L). It also altered the enzymatic activity of biomarker enzymes related to xenobiotic processing, anaerobic metabolism, and oxidative stress (GST, LDH, and CAT, respectively) and strongly accumulated in the embryos, as internal concentrations were 4 orders of magnitude higher than those detected in exposure solutions. MALDI-MSi revealed accumulations of the drug mainly in the head and eyes of the embryos (72 and 96 hpf). Thus, our results show that exposure to moxidectin decreases hatching success by 96 h and alters biochemical parameters in the early life stages of zebrafish while accumulating in the head and eye regions of the animals, demonstrating the need to prioritize this compound for environmental studies.

The flame retardants hexabromobenzene (HBB) and pentabromobenzene (PBB) have been extensively used and become ubiquitous pollutants in the aquatic environment and biota, but their potential toxic effects on wildlife remained unknown. In this study, by using zebrafish (Danio rerio) as a model, the bioconcentration and developmental neurotoxicity were investigated. Zebrafish embryos were exposed to HBB and PBB (0, 30, 100 and 300 μg/L) from 2 until 144 h post-fertilization (hpf). Chemical analysis showed bioconcentrations of both chemicals, while HBB is readily metabolized to PBB in zebrafish larvae. Embryonic exposure to both chemicals did not cause developmental toxicity, but induced locomotor behavioral anomalies in larvae. Molecular docking results indicated that both chemicals could bind to zebrafish acetylcholinesterase (AChE). Furthermore, HBB and PBB significantly inhibited AChE activities, accompanied by increased contents of acetylcholine and decreased choline in larvae. Downregulation of the genes associated with central nervous system (CNS) development (e.g., mbp, α1-tubulin, gfap, shha) as well as the corresponding proteins (e.g., Mbp, α1-Tubulin) was observed, but gap-43 was upregulated at both gene and protein levels. Together, our results indicate that both HBB and PBB exhibit developmental neurotoxicity by affecting various parameters related to CNS development and indications for future toxicological research and risk assessment of the novel brominated flame retardants.

As agriculture expands to provide food and wellbeing to the world’s growing population, there is a simultaneous increasing concern about the use of agrochemicals, which can harm non-target organisms, mainly in the aquatic environment. The fungicide Mancozeb (MZ) has been used on a large-scale and is a potent inducer of oxidative stress. Therefore, there is an urgent need for the development of more sensitive biomarkers designed to earlier biomonitoring of this compound. Here we tested the hypothesis that behavioral changes induced by sublethal MZ concentrations would occur first as compared to biochemical oxidative stress markers. Embryos at 4 h post-fertilization (hpf) were exposed to Mancozeb at 5, 10 and 20 μg/L. Controls were kept in embryo water only. Behavioral and biochemical parameters were evaluated at 24, 28, 72, and 168 hpf after MZ exposure. The results showed that MZ significantly altered spontaneous movement, escape responses, swimming capacity, and exploratory behavior at all exposure times. However, changes in ROS steady-stead levels and the activity of antioxidant enzymes were observable only at 72 and 168 hpf. In conclusion, behavioral changes occurred earlier than biochemical alterations in zebrafish embryos exposed to MZ, highlighting the potential of behavioral biomarkers as sensitive tools for biomonitoring programs.

To assess the time-dependent stress evidence in dynamic allocation of physiological metabolism of Nilaparvata lugens nymphs in response to elevated CO₂, we measured the time-dependent allocation of nutrient compositions and physiological metabolism in the bodies of N. lugens at 1h, 4h and 12h under elevated CO₂. Elevated CO₂ significantly increased the contents of nutrient compositions (protein, glucose and total amino acids) and catalase (CAT) enzyme activity in the body of N. lugens at 12h relative to 1h and 4h (P < 0.05). Significantly higher genes expression levels of acetylcholinesterase (AChE), heat shock protein (HSP70) and vitellogenin gene (vg) were observed in the body of N. lugens compared with those in ambient CO₂ at 4h (P < 0.05). These results showed that there was an instantaneous reaction of N. lugens nymphs to elevated CO₂, which indicated N. lugens may enhance stress defense response to future increasing CO₂ levels.