Temperature rise and microplastics interact with the toxicity of the antibiotic cefalexin to juveniles of the common goby (Pomatoschistus microps): Post-exposure predatory behaviour, acetylcholinesterase activity and lipid peroxidation.

The goal of this study was to investigate the toxicity of cefalexin to Pomatoschistus microps juveniles in relation to the presence of microplastics in the water and temperature rise. After acclimatization, groups of wild juveniles were exposed for 96h to artificial salt water (control), microplastics alone (0.184mg/l), cefalexin alone (1.3-10mg/l) and in mixture with microplastics (cefalexin: 1.3-10mg/l; microplastics: 0.184mg/l) at 20 and 25°C. Effect criteria were mortality, post-exposure predatory performance (PEPP), acetylcholinesterase activity (AChE) and lipid peroxidation levels (LPO). At 20°C, concentrations of cefalexin alone≥5mg/l significantly reduced PEPP (up to 56%; 96h-EC50=8.4mg/l), indicating toxicity of the antibiotic to juveniles after short-term exposure to water concentrations in the low ppm range. At 20°C, fish exposed to microplastics alone did not have significant differences in any of the parameters tested relative to the control group but tended to have an inhibition of the PEPP (23%) and AChE (21%); at 25°C, microplastics alone caused mortality (33%) and PEPP inhibition (28%). Thus, microplastics are toxic to P. microps juveniles. At 20°C, under simultaneous exposure to cefalexin and microplastics, the PEPP was significantly reduced (at cefalexin concentrations≥1.25mg/l). Moreover, at 25°C, the toxicity curves of cefalexin (PEPP based), alone and in mixture with microplastics, were significantly different (p<0.05; 96h-EC50 of 3.8 and 5.2mg/l, respectively), and the integrated data analysis indicated significant interactions between the two substances for all biomarkers. Thus, the presence of microplastics in the water influenced the toxicity of cefalexin. The rise of water temperature (from 20°C to 25°C), increased the microplastics-induced mortality (from 8 to 33%), and the inhibitory effects of cefalexin on the PEPP (up to 70%). Significant differences (p<0.05) between the toxicity curves of cefalexin alone at distinct temperatures were found, with a lower 96h-EC50 at 25°C (3.8mg/l) than at 20°C (8.4mg/l). Moreover, at 25°C, increases of AChE activity (14%) and LPO (72%) in fish exposed to the mixture treatment containing the highest cefalexin concentration were found, and the integrated analysis of data indicated significant interactions between cefalexin and temperature for PEPP, and among all stressors for LPO. Thus, the temperature rise increased the toxicity of microplastics and of cefalexin, alone and in mixture with microplastics, to P. microps juveniles. These findings raise concern on the long-term exposure of wild populations to complex mixtures of pollutants, likely decreasing their fitness, and highlight the need of more research on the combined effects of widely used pharmaceuticals, microplastics and temperature increase on wild species to improve environmental and human risk assessments of chemicals and their safe use under a global warming scenario.

Effects of multi-stressors on juveniles of the marine fish Pomatoschistus microps: Gold nanoparticles, microplastics and temperature.

Knowledge on multi-stressors effects required for environmental and human risk assessments is still limited. This study investigated the combined effects of gold nanoparticles (Au-NP), microplastics (MP) and temperature increase on Pomatoschistus microps, an important prey for several higher level predators, including some species edible to humans. Four null hypotheses were tested: H01: P. microps juveniles do not take up Au-NP through the water; H02: Au-NP (ppb range) are not toxic to juveniles; H03: the presence of MP do not influence the effects of Au-NP on juveniles; H04: temperature increase (20-25°C) does not change the effects of the tested chemicals on juveniles. Wild juveniles were acclimated to laboratory conditions. Then, they were exposed to Au-NP (≈5nm diameter) and MP (polyethylene spheres, 1-5µm diameter), alone and in mixture, at 20°C and 25°C, in semi-static conditions. After 96h of exposure to Au-NP, fish had gold in their body (0.129-0.546µg/g w.w.) leading to H01 refusal. Exposure to Au-NP alone caused a predatory performance decrease (≈-39%, p<0.05) leading to H02 refusal. MP did not change the Au-NP toxicity leading to H03 acceptance. Temperature rise significantly increased the concentration of gold in fish exposed to Au-NP (≈2.3 fold), and interacted with chemical effects (e.g. glutathione S-transferases activity) leading to H04 refusal. Thus, the results of this study highlight the importance of further investigating the effects of multi-stressors on marine fish, particularly the effects of temperature on the uptake, biotransformation, elimination and effects of nanoparticles and microplastics, either alone or in mixture. This knowledge is most important to improve the basis for environmental and human risk assessments of these environmental contaminants of high concern.

Does the presence of microplastics influence the acute toxicity of chromium(VI) to early juveniles of the common goby (Pomatoschistus microps)? A study with juveniles from two wild estuarine populations.

Toxicological interactions between microplastics (MP) and other environmental contaminants are of grave concern. Here, the potential influence of MP in the short-term toxicity of chromium to early juveniles of Pomatoschistus microps was investigated. Three null hypotheses were tested: (1) exposure to Cr(VI) concentrations in the low ppm range does not induce toxic effects on juveniles; (2) the presence of microplastics in the water does not influence the acute toxicity of Cr(VI) to juveniles; (3) the environmental conditions of the natural habitat where fish developed do not influence their sensitivity to Cr(VI)-induced acute stress. Fish were collected in the estuaries of Minho (M-est) and Lima (L-est) Rivers (NW Iberian Peninsula) that have several abiotic differences, including in the water and sediment concentrations of various environmental contaminants. After acclimatization to laboratory conditions, two 96h acute bioassays were carried out with juveniles from both estuaries to: (i) investigate the effects of Cr(VI) alone; (ii) investigate the effects of Cr(VI) in the presence of MP (polyethylene spheres 1-5µm ∅). Cr(VI) alone induced mortality (96h-LC50s: 14.4-30.5mg/l) and significantly decreased fish predatory performance (≤74%). Thus, in the range of concentrations tested (5.6-28.4mg/l) Cr(VI) was found to be toxic to P. microps early juveniles, therefore, we rejected hypothesis 1. Under simultaneous exposure to Cr(VI) and MP, a significant decrease of the predatory performance (≤67%) and a significant inhibition of AChE activity (≤31%) were found. AChE inhibition was not observed in the test with Cr(VI) alone and MP alone caused an AChE inhibition ≤21%. Mixture treatments containing Cr(VI) concentration ≥3.9mg/l significantly increased LPO levels in L-est fish, an effect that was not observed under Cr(VI) or MP single exposures. Thus, toxicological interactions between Cr(VI) and MP occurred, therefore, we rejected hypothesis 2. In the presence of MP, the negative effect caused by high concentrations of Cr(VI) on the predatory performance was significantly reduced in L-est fish but not in M-est fish, and Cr(VI) concentrations higher than 3.9mg/l caused oxidative damage in L-est fish but not in M-est fish. The acclimatization and test conditions were similar for fish from the two estuaries and these ecosystems have environmental differences. Thus, long-term exposure to distinct environmental conditions in the natural habitat during previous developmental phases influenced the sensitivity and responses of juveniles to Cr(VI), therefore, we rejected hypothesis 3. Overall, the results of this study indicate toxicological interactions between MP and Cr(VI) highlighting the importance of further investigating the combined effects of MP and other common contaminants.