Effects of wastewater-spiked nanoparticles of silver and titanium dioxide on survival, growth, reproduction and biochemical markers of Daphnia magna.

Silver (Ag) and titanium dioxide (TiO2) nanoparticles (NPs) are released into aquatic environments through wastewater treatment plants (WWTPs). Even though these NPs are mostly retained in WWTPs, a small fraction can be found in released effluents and may exert toxic effects on aquatic biota. Currently, the available information about the sublethal effects of wastewater-borne NPs on aquatic organisms is inconclusive and the importance of exposure media remains poorly understood. Previously, we demonstrated that rainbow trout juveniles chronically exposed to wastewater-borne AgNPs or TiO2NPs caused no effects on growth, but antioxidative stress mechanisms were triggered in fish organs. Accordingly, this study aimed to: (i) assess the chronic (21-d) effects of wastewater-borne AgNPs (0.3-23.5 µg L-1 Ag) and TiO2NPs (2.7-3.9 µg L-1 Ti) on survival, growth and reproduction of Daphnia magna; (ii) determine the short-term (96-h) effects of wastewater-borne AgNPs (30.3 µg L-1 Ag) and TiO2NPs (6.3 µg L-1 Ti) at the subcellular level (biochemical markers of neurotoxicity, anaerobic metabolism and oxidative stress); and (iii) compare the effects obtained in (i) and (ii) with the corresponding ones induced by effluent-supplemented and water-dispersed NPs. Total Ag and Ti levels were analytically quantified in all treatments. It was demonstrated that both wastewater-borne NPs are considered non-toxic to daphnids at tested concentrations, considering the endpoints at the individual (survival, growth, reproduction) and subcellular (biochemical markers) levels. Contrarily, when pristine forms of NPs were supplemented to effluents or water, concentration-dependent effects were noticed, particularly on cumulative offspring of daphnids. The significant effects on anaerobic metabolism and detoxification pathways caused by the effluent indicate background toxicity. Bearing in mind the achievement of a suitable risk assessment of NPs in aquatic environments, this combined approach looking at both the individual and subcellular levels responses come up with relevant information about the ecotoxicological harmlessness of wastewater-borne NPs in complex environmental matrices like WWTP effluents.

Chronic effects of wastewater-borne silver and titanium dioxide nanoparticles on the rainbow trout (Oncorhynchus mykiss).

Even though nanoparticles (NPs) are mostly removed by wastewater treatment plants, wastewater-borne NPs may show an altered toxicity to aquatic organisms. The main objectives of this work were: i) to assess the chronic (28 days) effects of wastewater-borne NPs of silver (AgNPs, 1.4-36.2 µg L-1) and titanium dioxide (TiO2NPs, 3.1-50.2 µg L-1) at the individual (growth) and biochemical (biomarkers of neurotoxicity, oxidative stress and energy metabolism) levels in rainbow trout Oncorhynchus mykiss; and ii) to compare them with their effluent-supplemented and water-dispersed counterparts. The total Ag and Ti levels were determined in several fish organs. The growth of O. mykiss was not affected by the NPs in any treatment, except a 29% increase at 5.5 µg L-1 of total Ag supplemented to effluents. The Ag level in organs of O. mykiss was significantly higher after exposure to water-dispersed AgNPs than their wastewater-borne or effluent-supplemented counterparts. No significant Ti uptake could be observed. Effluent-supplemented TiO2NPs (50.1 µg L-1 Ti) potentially induced neurotoxic effects, indicated by a 24% increase in acetylcholinesterase activity comparatively to controls. Energy reserves were unaffected by TiO2 treatments, while nearly all AgNP-containing treatments caused a depletion of total lipids, proteins and carbohydrates in the muscle, suggesting an increased energy demand for detoxification processes to cope with AgNPs. Besides NPs, the effluent matrix and dispersing agent (for AgNPs) induced significant effects on energetic reserves and oxidative stress, indicating background toxicity of both treatments at the biochemical level. Our study is the first to assess chronic effects of wastewater-borne NPs on rainbow trout. While no effects were found at the individual level, several biochemical markers were changed by the NPs exposure. Our results highlight the importance of using complex matrices for a reliable risk assessment of NPs in the aquatic environment.

Impact of wastewater-borne nanoparticles of silver and titanium dioxide on the swimming behaviour and biochemical markers of Daphnia magna: An integrated approach.

Due to their widespread use, silver (Ag) and titanium dioxide (TiO2) nanoparticles (NPs) are commonly discharged into aquatic environments via wastewater treatment plants. The study was aimed to assess the effects of wastewater-borne AgNPs (NM-300 K; 15.5 ±â€¯2.4 nm; 25-125 µg L-1) and TiO2NPs (NM-105; 23.1 ±â€¯6.2 nm; 12.5-100 µg L-1), from a laboratory-scale wastewater treatment plant, on Daphnia magna, at individual and subcellular level. For effect comparison, animals were also exposed to ASTM-dispersed NPs at the same nominal concentrations. The behaviour of D. magna was evaluated through monitoring of swimming height and allocation time for preferred zones after 0 h and 96 h of exposure. Biochemical markers of neurotransmission, anaerobic metabolism, biotransformation, and oxidative stress were subsequently determined. No 96-h EC50 (immobilization ≤ 4 %) could be obtained with wastewater-borne NPs and ASTM-dispersed TiO2NPs, whereas the ASTM-dispersed AgNPs resulted in an immobilization 96-h EC50 of 113.8 µg L-1. However, both wastewater-borne and ASTM-dispersed TiO2NPs, at 12.5 µg L-1, caused immediate (0 h) alterations on the swimming height. Allocation time analyses showed that animals exposed to ASTM-dispersed AgNPs spent more time on the surface and bottom at 0 h, and in the middle and bottom at 96 h. This pattern was not observed with ASTM-dispersed TiO2NPs nor with wastewater-borne AgNPs and wastewater-borne TiO2NPs. At the biochemical level, the more pronounced effects were observed with wastewater-borne AgNPs (e.g. induction of lactate dehydrogenase and glutathione S-transferase activities, and inhibition of catalase activity). This integrative approach showed that: (i) the behavioural and biochemical response-patterns were distinct in D. magna exposed to environmentally relevant concentrations of wastewater-borne and ASTM-dispersed NPs; (ii) the most pronounced effects on allocation time were induced by ASTM-dispersed AgNPs; and (iii) at the subcellular level, wastewater-borne AgNPs were more toxic than wastewater-borne TiO2NPs. This study highlights the need for the assessment of the effects of wastewater-borne NPs under realistic exposure scenarios, since processes in wastewater treatment plants may influence their toxicity.

Basagran® induces developmental malformations and changes the bacterial community of zebrafish embryos.

This study aimed to assess the effects of Basagran® on zebrafish (Danio rerio) embryos. The embryos were exposed to Basagran® at concentrations ranging from 120.0 to 480.6 mg/L, and the effects on embryo development (up to 96 h) and bacterial communities of 96 h-larvae were assessed. The embryo development response was time-dependent and concentration-dependent (106.35 < EC50 < 421.58 mg/L). The sensitivity of embryo-related endpoints decreased as follows: blood clotting in the head and/or around the yolk sac > delay or anomaly in yolk sac absorption > change in swimming equilibrium > development of pericardial and/or yolk sac oedema > scoliosis. A PCR-DGGE analysis was used to evaluate changes in the structure, richness, evenness and diversity of bacterial communities after herbicide exposure. A herbicide-induced structural adjustment of bacterial community was observed. In this study, it was successfully demonstrated that Basagran® affected zebrafish embryos and associated bacterial communities, showing time-dependent and concentration-dependent embryos' developmental response and structural changes in bacterial community. Thus, this work provides for the first time a complementary approach, which is useful to derive robust toxicity thresholds considering the embryo-microbiota system as a whole. The aquatic hazard assessment will be strengthened by combining current ecotoxicological tests with molecular microbiology tools.

Toxicity of seven priority hazardous and noxious substances (HNSs) to marine organisms: Current status, knowledge gaps and recommendations for future research.

Shipping industry and seaborne trade have rapidly increased over the last fifty years, mainly due to the continuous increasing demand for chemicals and fuels. Consequently, despite current regulations, the occurrence of accidental spills poses an important risk. Hazardous and noxious substances (HNSs) have been raising major concern among environmental managers and scientific community for their heterogeneity, hazardous potential towards aquatic organisms and associated social-economic impacts. A literature review on ecotoxicological hazards to aquatic organisms was conducted for seven HNSs: acrylonitrile, n-butyl acrylate, cyclohexylbenzene, hexane, isononanol, trichloroethylene and xylene. Information on the mechanisms of action of the selected HNS was also reviewed. The main purpose was to identify: i) knowledge gaps in need of being addressed in future research; and ii) a set of possible biomarkers suitable for ecotoxicological assessment and monitoring in both estuarine and marine systems. Main gaps found concern the scarcity of information available on ecotoxicological effects of HNS towards marine species and their poorly understood mode of action in wildlife. Differences were found between the sensitivity of freshwater and seawater organisms, so endpoints produced in the former may not be straightforwardly employed in evaluations for the marine environment. The relationship between sub-individual effects and higher level detrimental alterations (e.g. behavioural, morphological, reproductive effects and mortality) are not fully understood. In this context, a set of biomarkers associated to neurotoxicity, detoxification and anti-oxidant defences is suggested as potential indicators of toxic exposure/effects of HNS in marine organisms. Overall, to support the development of contingency plans and the establishment of environmental safety thresholds, it will be necessary to undertake targeted research on HNS ecotoxicity in the marine environment. Research should address these issues under more realistic exposure scenarios reflecting the prevailing spatial and temporal variability in ecological and environmental conditions.

Exposure of the cyanobacterium Nostoc muscorum from Portuguese rice fields to Molinate (Ordram(®)): Effects on the antioxidant system and fatty acid profile.

Herbicide contamination of aquatic ecosystems is a serious global environmental concern. Several herbicides enhance the intracellular formation of reactive oxygen species, and can lead to the damage of macromolecules and to a decrease of oxidant defenses in a wide range of non-target microorganisms including cyanobacteria. The effects of molinate (a thiocarbamate herbicide used for controlling grassy weeds in rice fields) on the activities of antioxidant enzymes such as superoxide dismutase, catalase, ascorbate peroxidase, glutathione reductase, and glutathione S-transferase were evaluated in Nostoc muscorum, a freshwater cyanobacterium with a significant spread in Portuguese rice fields. These were determined in N. muscorum cultures acutely (72h) exposed to concentrations ranging from 0.75 to 2mM of molinate. This study also analyzed the effects of molinate on: (1) the nonenzymatic antioxidant contents (reduced and oxidized glutathione, carotenoids, and proline), (2) the oxidative cell damage measured in terms of lipid peroxidation (MDA level) and electrolyte leakage (intactness of plasma membrane), and (3) the total fatty acid profile. The results showed that the activities of all antioxidant enzymes decreased dramatically with the rising concentration of molinate after 72h. Time-dependent and concentration-dependent increase in MDA and enhanced cell membrane leakage were indicative of lipid peroxidation, formation of free radicals and oxidative damage. Compared to control, 72-h herbicide exposure increased lipid peroxidation by 5.4%, 19% and 28% with 0.75, 1.5 and 2mM of molinate, respectively. Similarly, herbicide stress induced an increase in electrolyte leakage (5.8%, 29.5% and 30.2% above control, with 0.75, 1.5 and 2mM of molinate, respectively). The increased production of proline at higher molinate concentrations (the values rose above control by 45%, 95% and 156% with 0.75, 1.5 and 2mM, respectively) indicated the involvement of this osmoprotectant in a free radical scavenging mechanism. Moreover, a radical decline in both glutathione pool, carotenoids and saturated fatty acids were also observed. The results of the present study lead us to conclude that: (1) both enzymatic and nonenzymatic antioxidative defense system of N. muscorum are dramatically affected by molinate, (2) the herbicide induces peroxidation, (3) it contributes to an increase of the unsaturation level of cell membrane fatty acids. These evidences should be taken in account when using N. muscorum as an environmental indicator species in studies of herbicide biotransformation and biomarker response as well as in environmental monitoring programmes.

Bentazon triggers the promotion of oxidative damage in the Portuguese ricefield cyanobacterium Anabaena cylindrica: response of the antioxidant system.

Rice fields are frequently exposed to environmental contamination by herbicides and cyanobacteria, as primary producers of these aquatic ecosystems, are adversely affected. Anabaena cylindrica is a cyanobacterium with a significantly widespread occurrence in Portuguese rice fields. This strain was studied throughout 72 h in laboratory conditions for its stress responses to sublethal concentrations (0.75-2 mM) of bentazon, a selective postemergence herbicide recommended for integrated weed management in rice, with special reference to oxidative stress, role of proline and intracellular antioxidant enzymes in herbicide-induced free radicals detoxification. Activities of antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), and glutathione S-transferase (GST) increased in a time- and herbicide dose-response manner and were higher than those in the control samples after 72 h. A time- and concentration-dependent increase of malondialdehyde (MDA) levels and the enhanced cell membrane leakage following bentazon exposure are indicative of lipid peroxidation, free radicals formation, and oxidative damage, while increased amounts of SOD, CAT, APX, GST, and proline indicated their involvement in free radical scavenging mechanisms. The appreciable decline in the reduced glutathione (GSH) pool after 72 h at higher bentazon concentrations could be explained by the reduction of the NADPH-dependent glutathione reductase (GR) activity. The obtained results suggested that the alterations of antioxidant systems in A. cylindrica might be useful biomarkers of bentazon exposure. As the toxic mechanism of bentazon is a complex phenomenon, this study also adds relevant findings to explain the oxidative stress pathways of bentazon promoting oxidative stress in cyanobacteria.

Comparative toxicity of bentazon and molinate on growth, photosynthetic pigments, photosynthesis, and respiration of the Portuguese ricefield cyanobacterium Nostoc muscorum.

Bentazon and molinate are selective herbicides recommended for integrated weed management in rice. Their toxicity on growth and some biochemical and physiological parameters of Nostoc muscorum, an abundant cyanobacterium in Portuguese rice fields, was evaluated under laboratory conditions during time- and concentration-dependent exposure for 72 h. Results showed that toxic concentrations (0.75-2 mM) of both herbicides have pleiotropic effects on the cyanobacterium. Molinate was more toxic than bentazon to growth, respiration, chlorophyll-a, carotenoids, and phycobiliproteins contents. Protein content was increased by both herbicides although the effect was particularly evident with higher concentrations of molinate (1.5-2 mM). The herbicides had contrasting effects on carbohydrates content: molinate increased this organic fraction whereas bentazon decreased it. Photosynthesis and respiration were inhibited by both herbicides.