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The greater white-toothed shrew Crocidura russula has been used as a sentinel species for estimating environmental risks to human populations. Previous studies in mining areas have focused on the liver of shrews as the primary target of physiological and metabolic changes due to heavy metal pollution. However, populations persist even when detoxification by the liver seems to be compromised and damage is observed. These pollutant-adapted individuals inhabiting contaminated sites may exhibit altered biochemical parameters that confer increased tolerance in various tissues other than the liver. The skeletal muscle tissue of C. russula might be an alternative tissue that allows the survival of organisms inhabiting historically polluted sites due to the detoxification of redistributed metals. Organisms from two heavy metal mine populations and one population derived from an unpolluted site were used to determine the detoxification activities, antioxidant capacity, and oxidative damage, as well as cellular energy allocation parameters and acetylcholinesterase activity (a biomarker of neurotoxicity). Muscle biomarkers differ between shrews from polluted sites and shrews from the unpolluted location, with the mine animals showing: (1) a decreased energy consumption concomitant with increased energy reserves and total available energy; (2) reduced cholinergic activity, suggesting an impairment of neurotransmission at the neuromuscular junction; (3) an overall decrease in detoxification capacity and enzymatic antioxidant response and a higher level of lipid damage. Also, some of these markers differed between females and males. These changes may have resulted from a decreased detoxifying capacity of the liver and could potentially bring about significant ecological effects for this highly active species. Heavy metal pollution induced physiological changes in Crocidura russula showing that skeletal muscle may serve as a backup sink organ allowing rapid species adaptation and evolution. published

Background: Most European rivers have not yet achieved “good” ecological status. In addition, the presence and abundance of microplastics (MPs) in freshwater is a matter of great concern to the scientific community. Methodology: This study assesses the ecological status of four sampling sites of Selho (S1–S4) and Costa-Couros (C1–C4) rivers (Guimarães, Portugal), and the abundance of MPs in sediments and benthic macroinvertebrates (Oligochaeta) from those sites. Results: All sites of both rivers under higher pressure did not reach a “good” ecological status (S2–S3, C2–C4) based on the macroinvertebrate community. High levels of nutrients were observed at all sites in both rivers (except C2), especially phosphorus. In the Oligochaeta’s gut of Costa-Couros river, the high number of MPs seems to be associated with their weight (95.25 ± 25.61 to 1069.00 ± 385.30 MPs g/fresh weight), suggesting the existence of malnutrition and digestive disorders, whereas the highest number of MPs in gut (134.00 ± 30.36 to 558.6 ± 100.70 MPs g/fresh weight) were found for the heaviest organisms of Selho. Conclusions: Thus, sites with higher ecological status do not necessarily have lower abundance of MPs. In the sediments, urbanization seems to be the main driver for MP contamination. MP contamination is pervasive across the sediments and Oligochaeta’s gut in both rivers. Since MPs have the potential to cause harm to environmental and human health, it is essential to monitor not only the ecological status of freshwaters, but also emerging pollutants such as MPs., This study was financially supported by the Guimarães City Council, by the Ministry of the Environment and Climate Action (Portugal) through the project “Aqualastic: Educar, Reduzir e Valorizar”, by the Foundation for Science and Technology (FCT) and the POPH/FSE (Human Potential Operational Program/European Social Fund) through funds of CESAM (UIDP/50017/2020; UIDB/50017/2020; LA/P/0094/2020), and by the project comPET (POCI-01-0145-FEDER-030361).

Microplastics (MPs) are ubiquitous and persistent contaminants, particularly in river sediments reaching high abundances with potential to impair ecosystem functions and services of great importance to humankind. Thus, monitoring programmes and public awareness are pivotal to implement mitigation and remediation strategies. This study evaluated the abundance of MPs in freshwater sediments and in benthic macroinvertebrates collected at four sites with different anthropogenic pressures of Costa/Couros river (Guimarães - Portugal), in order to alert, raise awareness and educate local citizens for (micro)plastic pollution. Results showed higher levels of MPs in the sediments of sites with more anthopogenic preassure. High number of MPs was also observed inside macroinvertebrates’ gut from all sites, particularly in low weight organisms indicating malnutrition and digestive disorders. Such results were incorporated into public awareness campaigns to increase the citizens' knowledge and understanding about MPs prevalence and threats to the environment, human health and the economy, as well as spread individual measures and actions that can reduce MP contamination in aquatic ecosystems. This educational study also promoted the artistic creation in public spaces to raise awareness about plastic pollution.

Freshwater provides valuable services and functions to humankind. However, macroinvertebrates that underpin the delivery of many of those ecosystem services and functions are under an additional threat caused by microplastic pollution. Chironomids are one of the most abundant groups of macroinvertebrates in these environments and the most sensitive to microplastics. This investigation addressed the effects of polyurethane (PU-MPs; 7.0-9.0 µm) on the chironomid

Thiamethoxam (TMX) is a systemic neonicotinoid that acts as a partial agonist of the nicotinic acetylcholine receptors (nAChRs). However, target species have shown resistance to formulations based on such neonicotinoids, which can also be expected for non-target insects. This research aimed to study the effects of a formulation based on TMX [Cruiser® 350 FS (CRZ)] on the life traits of Chironomus xanthus filial generation (F1) and compare it with the parental generation (P). Environmentally relevant concentrations of CRZ significantly decreased larvae growth P generation , also slowing and decreasing their emergence. Larvae of the F1 generation were less sensitive than their parents, suggesting that the progeny were able to thrive and perform basic physiological functions better than the parental generation. Our results highlight that insect resistance to neonicotinoids may be associated with the better performance of the filial generation, which is related to the change in affinities of the active ingredient for the sub-units constituting the nAChRs subtypes of F1 organisms, inherited from P organisms that were able to survive and reproduce. Moreover, further studies using biochemical and omics tools should be performed to disentangle the specific changes occurring at the nAChRs throughout insect development.

A variety of veterinary and human medicinal products (VHMPs) are found in groundwater, an often-neglected habitat inhabited by species with unique traits, stygobitic species. It is crucial to understand the effect of VHMPs on stygobitic species because they may respond differently to stressors than surface species. Our hypothesis is that groundwater species may be more susceptible to environmental contaminants due to less plasticity in their detoxification response and acquisition of energy because subterranean habitats are more stable and isolated from anthropogenic activities. We performed a battery of biomarkers associated with important physiological functions on the stygobitic asellid crustacean Proasellus lusitanicus, after a 14-day exposure to acetaminophen, a commonly used pharmaceutical and pollutant of groundwaters. Our results show a decrease in total glutathione levels and an increase in glutathione S-transferase activity, suggesting a successful detoxification response. This helps explaining why acetaminophen did not cause oxidative damage, as well as had no effect on cholinesterase activity nor in aerobic production of energy. This study shows the remarkable capacity of P. lusitanicus to tolerate sublethal concentrations of VHMP acetaminophen. Most ecotoxicological studies on stygobitic species focused on the lethal effects of these compounds. The present study focuses on consequences at sublethal concentrations. Future studies should assess the stress levels induced to better predict and estimate the impacts of contaminants on groundwater ecosystems.

Bio-insecticides have been increasingly used worldwide as ecofriendly alternatives to pesticides, but data on their effects in non-target freshwater organisms is still scarce and limited to insects. The aim of this study was to determine the lethal and sub-lethal effects of the bio-insecticides Bac Control (based on Bacillus thuringiensis kurstaki—Btk) and Boveril (based on Beauveria bassiana—Bb) on regeneration, behavioral, and reproductive endpoints of the freshwater planarian Girardia tigrina. The estimated LC50–48h were > 800 mg a.i./L for Btk and 60.74 mg a.i./L for Bb. In addition, exposure to Btk significantly decreased locomotion and feeding activities of planarians (lowest observed effect concentration (LOEC) of 12.5 mg a.i./L Btk) and fecundity rate (LOEC = 3.12 mg a.i./L Btk), whereas exposure to Bb significantly delayed regeneration (LOEC = 0.75 mg a.i./L Bb) and decreased fecundity rate (1.5 mg a.i./L Bb) of planarians. Thus, both bio-insecticides induced deleterious sub-lethal effects on a non-insect freshwater invertebrate species. However, only Bb-based formulation affected the survival, fecundity rate, and regeneration at concentrations below the maximum predicted environmental concentration (PEC = 247 mg/L). Thus, care should be taken when using such formulations as alternatives to chemical insecticides near aquatic ecosystems.

Mercury (Hg) is a non-essential metal that can have toxic effects on the fitness of organisms and tends to bioaccumulate with age and to biomagnify in higher trophic levels. Few studies have assessed oxidative stress and neurotoxicity in deep-water sharks. This study evaluated early ontogenetic changes and physiological effects (antioxidant defences, oxidative damage, aerobic metabolism and neurotransmission functions) of Hg accumulation in the white muscle and brain tissues of the velvet belly lantern shark Etmopterus spinax from the southern Iberian coast (NE Atlantic). Results suggested that the low mercury concentrations observed may induce acute effects in E. spinax before they reach sexual maturity. We found different Hg concentrations in E. spinax: [Hg] males > [Hg] females; [Hg] muscle > [Hg] brain. Females appeared to have higher redox capability translated into higher activities and levels of antioxidant defences than males. However, higher levels of oxidative damage were also observed in females. Whilst the mechanisms underlying these effects remain unknown, these results suggest differences in mercury accumulation between tissues and sex, and potentially deleterious effects on oxidative stress status and neurophysiology of E. spinax, potentially impairing swimming performance and reproduction, which could subsequently impact on the health of both individuals and population. info:eu-repo/semantics/publishedVersion

Bio-insecticides have been increasingly used worldwide as ecofriendly alternatives to pesticides, but data on their effects in non-target freshwater organisms is still scarce and limited to insects. The aim of this study was to determine the lethal and sub-lethal effects of the bio-insecticides Bac Control (based on Bacillus thuringiensis kurstaki - Btk) and Boveril (based on Beauveria bassiana - Bb) on regeneration, behavioral and reproductive endpoints of the freshwater planarian Girardia tigrina. The estimated LC50 − 48h were > 800 mg a.i./L for Btk and 60.74 mg a.i./L for Bb. In addition, exposure to Btk significantly decreased locomotion and feeding activities of planarians (lowest observed effect concentration (LOEC) of 12.5 mg a.i./L Btk) and fecundity rate (LOEC = 3.12 mg a.i./L Btk), whereas exposure to Bb significantly delayed regeneration (LOEC = 0.75 mg a.i./L Bb) and decreased fecundity rate (1.5 mg a.i./L Bb) of planarians. Thus, both bio-insecticides induced deleterious sub-lethal effects on a non-insect freshwater invertebrate species. However, only Bb-based formulation affected the survival, fecundity rate and regeneration at concentrations below the maximum predicted environmental concentration (PEC = 247 mg/L). Thus, care should be taken when using such formulations as alternatives to chemical insecticides near aquatic ecosystems.

Despite the increasing interest for biochar as a soil amendment, a knowledge gap remains on its impacts on non-target soil and aquatic species. We hypothesised that biochar particle size and application rate can play a role in the toxicity to biota. Pine woodchip biochar was incorporated in a clean soil at three particle size classes: small (0.5 mm), medium (1-2 mm), and large (4 mm), and at two concentrations: 1% and 6% w/w. A laboratory screening with earthworm Eisenia andrei avoidance behaviour bioassay was carried out to test the most adequate application rates, particle sizes and soil-biochar pre-incubation period. Thereafter, a 28-day greenhouse microcosm experiment was conducted as an ecologically more representative approach. Survival, vertical distribution and weight changes of E. andrei, and bait-lamina consumption were recorded. Soil leachates from the microcosms were collected to evaluate their impact on Daphnia magna immobilisation and Vibrio fischeri (Microtox®) bioluminescence. A feeding experiment with E. andrei was also performed to address earthworm weight changes and to conduct a screening of PAH-type metabolites in their tissue. The 6%0.5 mm treatment pre-incubated for 96 h induced significant avoidance of the earthworms. Significantly lower bait-lamina consumption was observed in microcosms for the 6%0.5 mm treatment. Moreover, particle size was a statistically significant factor regarding the loss of weight in the feeding experiment and higher concentration of naphthalene-type metabolites detected in E. andrei tissue, when exposed to0.5 mm biochar particles. The leachates had no adverse effects on the aquatic species. The results suggest that particles0.5 mm of pine woodchip biochar can pose sub-lethal effects on soil biota.

The degradation of dyes can generate harmful by-products, thereby requiring the need to evaluate the toxicity to aquatic organisms. This study aims to evaluate the chronic ecotoxicity of methylene blue dye degraded by the Fenton process using the non-target planarian Girardia tigrina as a sensitive bioindicator of environmental contamination. The bioassays evaluated the lethality of several concentrations of the untreated and degraded dye methylene blue (MB), as well as, their sub-lethal effects on locomotion, feeding, regeneration, and reproduction. In both acute and chronic tests, the degraded dye had a stronger toxic effect when compared to the untreated dye. This negative effect after treatment was mainly associated with the presence of residual hydrogen peroxide and iron (and consequently the hydroxyl radical formed). We conclude that the utilization of the Fenton process using less oxidizing agents should be considered as important alternatives for the protection of aquatic ecosystems, without compromising the efficient removal of MB.

13 páginas The present study aims to use behavioral responses of the freshwater planarian Girardia tigrina to assess the impact of anthropogenic activities on the aquatic ecosystem of the watershed Araguaia-Tocantins (Tocantins, Brazil). Behavioral responses are integrative and cumulative tools that reflect changes in energy allocation in organisms. Thus, feeding rate and locomotion velocity (pLMV) were determined to assess the effects induced by the laboratory exposure of adult planarians to water samples collected in the region of Tocantins-Araguaia, identifying the sampling points affected by contaminants. Furthermore, physicochemical and microbiological parameters, as well as the presence of inorganic compounds (dissolved aluminum, total barium, total chloride, dissolved iron, total fluoride, total manganese, nitrates, nitric nitrogen, total sulfate, total zinc) and surfactants, were determined on each specific sampling point. The behavioral biomarkers (feeding rate and pLMV) of the freshwater planarians were significantly decreased when organisms were exposed to water samples from four municipalities (Formoso do Araguaia, Lagoa da Confusão, Gurupi and Porto Nacional), sites of the Tocantins-Araguaia hydrographic region—TAHR. Both behavioral biomarkers decreased up to ~37–39% compared to organisms in ASTM medium only. Our results showed that these behavioral biomarkers can be used for fast screening monitoring of environmental samples of freshwater ecosystems, since a decrease in feeding rate and locomotor activity was observed in sites impacted by anthropogenic activities. However, the absence of effects observed in some sampling points does not represent the absence of contamination, since several other classes of contaminants were not determined. In these negative results, the absence of deleterious effects on behavioral biomarkers might only be indicative that the potential presence of contaminants on such sites does not significantly affect the performance of planarians. This fast screening approach seems to be useful to determine contaminated sites in freshwater ecosystems for biomonitoring purposes. This knowledge will help to develop biomonitoring programs and to decide appropriate sampling sites and analysis. Abstract. 1. Introduction. 2. Material and Methods. 3. Results. 4. Discussion. 5. Conclusions. References

Acetylcholinesterase (AChE) is a useful biomarker for organophosphate and carbamate pesticides exposure. The inhibition of this enzyme has been associated with neurotoxicity and alterations at higher levels of biological organization, such as behavior and development impairments. In this chapter, we describe the methodologies for analyses of AChE activity in pools of 96 h of embryos of zebrafish (Danio rerio) using a spectrophotometric method adapted to 96-well microtiter plates.

Fish embryo toxicity (FET) test using zebrafish (Danio rerio) has been established as an alternative assay to animal experimentation. The FET assay enables the assessment of multiple morphological endpoints during the development of zebrafish early life stages, showing high impact to the field of ecotoxicology on risk assessment of chemicals and pollutants. Moreover, it is also applied to screening drug-induced toxicity and human diseases, due to the high genetic and physiological orthology between zebrafish and humans. Here, we describe FET test, with all steps and several adaptations involved in the methodological procedures. To demonstrate the efficiency of this method, results using the reference substance 3,4-dichloroaniline (DCA) were included to demonstrate sublethal and teratogenic malformations on zebrafish embryos. Thus, there is a strong tendency for using FET tests as a replacement strategy of traditional tests in toxicology and ecotoxicology.

Acetylcholinesterase (AChE) is a useful biomarker for organophosphate and carbamate pesticides exposure. The inhibition of this enzyme has been associated with neurotoxicity and alterations at higher levels of biological organization, such as behavior and development impairments. In this chapter, we describe the methodologies for analyses of AChE activity in pools of 96 h of embryos of zebrafish (Danio rerio) using a spectrophotometric method adapted to 96-well microtiter plates.

Fish embryo toxicity (FET) test using zebrafish (Danio rerio) has been established as an alternative assay to animal experimentation. The FET assay enables the assessment of multiple morphological endpoints during the development of zebrafish early life stages, showing high impact to the field of ecotoxicology on risk assessment of chemicals and pollutants. Moreover, it is also applied to screening drug-induced toxicity and human diseases, due to the high genetic and physiological orthology between zebrafish and humans. Here, we describe FET test, with all steps and several adaptations involved in the methodological procedures. To demonstrate the efficiency of this method, results using the reference substance 3,4-dichloroaniline (DCA) were included to demonstrate sublethal and teratogenic malformations on zebrafish embryos. Thus, there is a strong tendency for using FET tests as a replacement strategy of traditional tests in toxicology and ecotoxicology.

The activation of insects’ immune system due to the ingestion of microplastics (MPs) has only been evidenced by the upregulation of specific genes. The activation of phenoloxidase (PO) system is one of the primary responses involved in insects' innate immunity when facing parasites and pathogens, and ingestion of MPs can trigger a similar process. This study aimed at addressing the activities of basal PO and total PO (PO+ prophenoloxidase – proPO), in Chironomus riparius larvae (a model species in ecotoxicology) exposed to sediments spiked with polyethylene microplastics (PE-MPs; size-range 32–63 µm; concentrations: 1.25; 5; to 20 g kg–1) for 48 h. The ingestion of PE-MPs by larvae triggered a significant increase of basal PO activity at 5 and 20 g PE-MPs kg–1, by 26% and 29%, respectively, whereas total PO increased significantly in the latter (+48%), suggesting de novo synthesis of proPO by organisms. Considering the particle size, the immune response's activation is probably linked to damage in the epithelial cells of the gut lumen. This research work provides the first evidence on the activation of the insect's innate immune system after ingestion of MPs and underlines the PO activity as a good indicator of the immune response induced by MPs' ingestion.

Freshwater planarians have been gaining relevance as experimental animals for numerous research areas given their interesting features, such as high regeneration potential, shared features with the vertebrate's nervous system or the range of endpoints that can be easily evaluated in response to contaminants. Ecotoxicological research using these animals has been steadily increasing in the past decades, as planarians' potentialities for this research area are being recognized. In this work, we used polycyclic aromatic hydrocarbons (PAHs) as model contaminants and evaluated effects of exposure to phenanthrene, pyrene and benzo[a]pyrene (B[a]P) in planarians. The freshwater planarian Girardia tigrina was chosen and mortality, cephalic regeneration (during and post-exposure), behavioral endpoints and presence of PAHs in tissues, were evaluated. Mortality was only observed in planarians exposed to phenanthrene, with an estimated LC50 of 830 μg L−1. Results indicate that planarian behavioral endpoints were very sensitive in response to sub-lethal concentrations of PAHs, showing a greater sensitivity towards B[a]P and pyrene. Briefly, post-exposure locomotion and post-exposure feeding were significantly impaired by sub-lethal concentrations of all compounds, whereas regeneration of photoreceptors was only significantly delayed in planarians exposed to pyrene. Moreover, levels of PAH-type compounds in planarian tissues followed a concentration-dependent increase, showing uptake of compounds from experimental solutions. The present results highlight the importance of studying alternative and complementary endpoints, such as behavior, not only because these may be able to detect effects at lower levels of contamination, but also due to their ecological relevance. The simplicity of evaluating a wide range of responses to contaminants further demonstrates the utility of freshwater planarians for ecotoxicological research.

In the coming decades, and despite advances in the selection of resistant strains and the production of triploid organisms, the temperature could seriously affect salmonid aquaculture. Lower environmental tolerance has been hinted at for triploids, but the physiological mechanisms leading to such differences, and whether they are translated to the individual level, are poorly understood. This study aimed to evaluate the effects of seasonal variations on the humoral and immune status in the blood (peripheral blood leukocytes) and plasma (antiprotease, lysozyme and peroxidase activities), the oxidative stress (catalase, glutathione-S-transferase, total glutathione and lipid peroxidation) balance in the liver, and the energy budget (sugars, lipids, proteins and energy production) in the liver and muscle of diploid and triploid Oncorhynchus mykiss. Leukocytes&rsquo, numbers changed with the water temperature and differed between fish ploidies. Peroxidase activity was increased in the summer, but lysozyme and antiprotease activities were increased in the winter. Concomitantly, antioxidant defenses were significantly altered seasonally, increasing oxidative damage at higher temperatures. Moreover, warmer waters induced a reduction in the energy production measured in the liver. Differences in feed efficiency, which have been previously reported, were confirmed by the low lipid and protein contents of the muscle of the triploids. In sum, the inherent trade-offs to deal with the seasonal changes culminated in the higher growth observed for diploid fish.

Riverine sediments are major sinks of microplastics from inland anthropogenic activities, imposing a threat to freshwater benthic invertebrates. This study investigated the ingestion of three size-classes (SC) of irregularly shaped polyethylene microplastics (PE-MPs; SC I: 32-63 μm; II: 63-250 μm; III: 125-500 μm) after 48 h by dipteran larvae (detritivore/collector) Chironomus riparius, and the consequent effects on neurotransmission, energy allocation and oxidative stress. The tested PE-MPs concentrations (1.25; 5; 20 g kg-1) were within the range of concentrations reported in riverbanks from highly urbanised areas (1 - 9 g kg-1), except for 20 g kg-1 representing the worst-case scenario. After exposure to SC I, larvae presented high amounts (up to ∼2400 particles/organism) of PE-MPs in their guts, with an average size-range of 30-60 μm. In the SC II and III, larvae presented PE-MPs of higher diameter (up to 125 μm) and a visible gut obstruction. The high number of particles in the larval gut (SC I) and/or difficulties for their egestion (SC I, II and III) induced oxidative damage and reduced aerobic energy production. In addition, larvae exposed to SC II and III revealed depletion in their total lipid reserves as a consequence of lacking nutrients, and the ones exposed to SC III presented a decrease in their detoxification capacity. These results highlight that freshwater detritivores with low selective feeding behaviour (e.g., chironomids) are more prone to ingest microplastics, with potentially adverse effects on cellular metabolism, redox status and antioxidant-detoxification defences. These harmful effects at lower levels of the biological organisation may ultimately affect organisms' physiology and fitness. published

Made available in DSpace on 2022-04-29T08:33:02Z (GMT). No. of bitstreams: 0 Previous issue date: 2022-01-15 European Commission Fundação para a Ciência e a Tecnologia Ministério da Ciência, Tecnologia e Ensino Superior Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) European Regional Development Fund Centro de Investigação em Materiais Cerâmicos e Compósitos Centro de Estudos Ambientais e Marinhos, Universidade de Aveiro Microplastics and nanomaterials are applied in a myriad of commercial and industrial applications. When leaked to natural environments, such small particles might threaten living organisms' health, particularly when considering their potential combination that remains poorly investigated. This study investigated the physiological and biochemical effects of polyethylene (PE; 64-125 μm in size, 0.1, 1.0, and 10.0 mg·L−1) single and combined with an engineered nanomaterial applied in antifouling coatings, the copper-aluminium layered double hydroxides (Cu-Al LDH; 0.33, 1.0, and 3.33 mg·L−1) in the flatfish Solea senegalensis larvae (8 dph) after 3 h exposure, in a full factorial design. Particles ingestion, histopathology, and biochemical biomarkers were assessed. Fish larvae presented

The timing of metamorphosis has recently been referred as a more suitable proxy to evaluate postlarval quality than size. Indeed, while displaying a larger size at settlement, late settlers that originate from larvae that have delayed metamorphosis commonly display poorer growth performances This delay in metamorphosis is possibly at the expenses of larval endogenous energetic reserves that, once allocated to facilitate the delayed transition to benthic life, will no longer be available to help fuelling early juvenile somatic growth. To further advance our knowledge on this topic, we evaluated the physiological status (energy reserves and allocation, aerobic and anaerobic energy consumption), along with biochemical responses related with detoxification processes, antioxidant defences, oxidative damage, neuromotor activity in early settlers (ES), middle settlers (MS) and late settlers (LS) of postlarvae of the marine ornamental shrimp Lysmata seticaudata. Our results revealed that LS postlarvae presented a higher weight compared to MS and ES, likely related with a lower metabolism and neuromotor activity. Yet, the low metabolism allied with diminished detoxification and antioxidant capacities seemed to result in an increased oxidative stress condition that may negatively condition the growth performance of LS postlarvae. Conversely, ES postlarvae presented a lower weight, likely because of high metabolic costs associated with increased neuromotor activity, detoxification, and antioxidant capacities to avoid oxidative damage. The present study highlights how the physiological, metabolic and biochemical status of L. seticaudata postlarvae is shaped by the timing of their metamorphosis, as well as how this event will shape their early benthic life and confirms that a larger size or weight at metamorphosis may not be good proxies to select premium seedlings for grow-out. Overall, metamorphosis is not a new beginning and does not reset larval history. Crustacean farmers should avoid decoupling larviculture history from grow-out, as only by knowing larval performance to metamorphosis will it be possible to enhance survival and growth performances to commercial size.

Made available in DSpace on 2018-12-11T16:54:28Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-08-25 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Federación Española de Enfermedades Raras Fundació Catalana de Trasplantament Centro de Estudos Ambientais e Marinhos, Universidade de Aveiro Dyes have been used for more than twenty thousand years in textile, pharmaceutical, food, cosmetic, and photographic industries, among others. Despite their importance in these applications, dyes can be toxic and resistant to many degradation processes used in wastewater treatment plants. Although a large number of dyes have been released in the environments in high amounts, studies into the environmental toxicity of these substances are still scarce. The aim of this study was to evaluate the potential toxic effects of textile dyes Disperse Red 60, Disperse Red 73 and Disperse Red 78 in zebrafish early life stages. To this end, biochemical biomarkers were selected to evaluate non-enzimatic antioxidant (Total Glutathione), antioxidant enzymes (Glutathione S-transferase and Catalase), oxidative stress (lipid peroxidation), neurotransmission (acetylcholinesterase) and energetic metabolism parameters (energy available and energy consumed) after 96 h exposure to these dyes. Our results demonstrated that these disperse dyes induce biochemical alterations in zebrafish embryos at environmental realistic concentrations and that the discharge of these disperse dyes into water bodies should be carefully evaluated. The selected biomarkers were sensitive as early-warning endpoints of disperse dyes toxicity on zebrafish embryos. Implications for risk assessment and indications for future research are discussed. School of Pharmaceutical Sciences of Ribeirão Preto University of São Paulo Department of Biology and CESAM University of Aveiro CENSE Department of Environmental Sciences and Engineering Faculty of Sciences and Technology New University of Lisbon, Quinta da Torre NEEA/CRHEA/SHS São Carlos Engineering School University of São Paulo Institute of Chemistry-State University “Julio de Mesquita Filho”-UNESP Faculty of Sciences and CESAM University of Lisboa, Campo Alegre National Institute of Science and Technology for Detection Toxicological Evaluation and Removal of Emerging and Radioactive Contaminants (INCT-DATREM) Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq) Institute of Chemistry-State University “Julio de Mesquita Filho”-UNESP CNPq: 142379/2014-0 FAPESP: 2014/14621-0 FAPESP: 2016/03281-9 FAPESP: 2017/05543-3 CAPES: 88881.134597/2016-01 Federación Española de Enfermedades Raras: PT2020 Fundació Catalana de Trasplantament: SFRH/BPD/109199/2015 Centro de Estudos Ambientais e Marinhos, Universidade de Aveiro: UID/AMB/50017

Dyes are widely distributed worldwide, and can be found in wastewaters resulting from industrial or urban effluents. Dyes are of particular concern as contaminants of the aquatic environment, since their toxicity remain poorly understood. Thus, the current study was designed to assess the effects induced by the synthetic azo dye Basic Red 51 (BR51) and by the natural naphthoquinone dye erythrostominone (ERY) on zebrafish early life stages (Danio rerio) at different biological organization levels, i.e., studying how changes in biochemical parameters of important physiological functions (neurotransmission and cellular energy allocation) may be associated with behavior alterations (swimming activity). This approach was also used to assess the effects of ERY after its photodegradation resulting in a colorless product(s) (DERY). Results showed that after 96 h exposure to BR51 and Ery, zebrafish embryos consumed less energy (LOEC = 7.5 mg/L), despite the unaltered levels of available energy (carbohydrates, lipids and proteins). Hence, cellular energy allocation (CEA) was significantly increased. On the other hand, only ERY decreased the acetylcholinesterase activity (LOEC = 15 mg/L). Despite that, zebrafish larvae exposed to both dyes until 144 h were less active. In contrast, DERY did not affect any parameter measured. These results indicate an association between a decrease consumption of energy and decrease swimming activity resulting from an environmental stress condition, independently of the neurotoxicity of the dyes. Degradation of ERY by light prevented all toxic effects previously observed, suggesting a cheap, fast and easy alternative treatment of effluents containing this natural dye. All tools assessed in our current study were sensitive as early-warning endpoints of dyes toxicity on zebrafish early life stages, and suggest that the CEA assay might be useful to predict effects on locomotor activity when cholinergic damage is absent.

Pyrethroid insecticides have been used for decades and their worldwide market continues to increase, despite their high toxicity to non-target insects. Recent studies reveal that it is essential to investigate the secondary mechanisms of action of type II pyrethroids to understand their cellular effects on invertebrates. The aim of this study was to evaluate the lethality, behaviour and physiological alterations and energetic costs in caddisfly larvae exposed to environmentally relevant concentrations of esfenvalerate (ESF). ESF caused both mortality and feeding inhibition of exposed caddisfly larvae: nominal ESF 96 h LC50 was 2.29 μg/L; feeding activity was impaired at concentrations equal or above 0.25 μg/L. At the cellular level, glutathione-S-transferase (GST) activity was increased on caddisfly larvae exposed to 0.25 and 0.5 μg/L ESF, which might contribute to prevent oxidative damage since levels of lipid peroxidation (LPO) were not altered. The energy budget of exposed caddisfly larvae was impaired by exposure to 0.25 μg/L ESF since sugar and protein contents decreased, while a decline of energy consumption was observed. The analysis of feeding, energy reserves and consumption data through structural equation modelling (SEM) allowed to quantify the direct and indirect effects of ESF exposure on bioenergetics of caddisfly larvae. SEM analysis showed a strong negative direct influence of ESF onto feeding activity, sugars content and energy consumption, highlighting a significant positive relationship between sugars and protein contents. These results show that energy expenditure is related to oxidative defense mechanisms induced by ESF stress that may lead to deleterious effects on growth and development.

Despite the frequent detection of organic ultraviolet-filters (UV-filters) in freshwater sediments, there is a lack of ecotoxicological data undermining a correct risk assessment for these emerging contaminants. The present study assessed the effects of three of the most commonly used UV-filters (benzophenone-3 - BP3; 3-(4-methylbenzylidene)camphor - 4-MBC and octocrylene - OC) on Chironomus riparius life history and biochemical responses. Standard ecotoxicological assays confirmed that all compounds impaired growth of C. riparius larvae and induced developmental effects such as delayed emergence and a reduction of imagoes weight. Concerning the biochemical responses analysed no evidences of oxidative damage in lipids or neurotoxicity (tested assessing acetylcholinesterase activity) were observed for any of the tested compounds. However, 4-MBC exposure induced a decrease in catalase activity and an increase in glutathione-S-transferase activity at 14.13mg/Kg while OC exposure caused an increase in total glutathione levels at 0.23 and 18.23mg/Kg. Exposure to all UV-filters tested, increased energy consumption on C. riparius with significant differences above 1.00mg/Kg for BP3, 0.09mg/Kg for 4-MBC and 2.13mg/Kg for OC. These results suggest that environmental relevant concentrations of UV-filters can cause deleterious effects to aquatic benthic species, such as C. riparius, and call for further research concerning effects of organic UV-filters on natural invertebrate communities and ecosystem functioning.

Physiological responses allow populations to cope with metal contamination and can be involved in the evolution of tolerance under historical metal contamination scenarios. Here we investigate physiological aspects that might be underlying the heritable high tolerance to cadmium (Cd) in two Chironomus riparius populations collected from historically metal contaminated sites in comparison to two populations from reference sites. To evaluate differences in the physiological response to short-term Cd exposure, protein expression profiles, metallothioneins [MTs] and several antioxidant defences such as total glutathione (GSHt), catalase (CAT) and glutathione-S-transferases [GSTs], were measured in all four populations reared for at least 8 generations under laboratory clean conditions. Cd-induced oxidative damage in lipids and energy related parameters (energy consumption and energy reserves) were also assessed. Results showed two major gradients of protein profiles according to Cd concentration and population tolerance. Furthermore, Cd-tolerant populations showed higher baseline levels of MTs and GSHt while Cd-sensitive populations, collected from reference sites, showed significant induction of GSHt levels with Cd exposure that were nonetheless insufficient to avoid increased oxidative damage to lipids. Cd exposure had no clear effects on the antioxidant enzymes or energy reserves but triggered a general increase in energy consumption. Finally, energy consumption was higher in Cd-tolerant populations across experimental conditions. Altogether, results demonstrate that inherited Cd-tolerance in these midge populations is related, at least in part, with different constitutive levels and plasticity of different defence mechanisms confirming the validity of using multiple physiological traits when studying evolution of tolerance.

The insecticide thiamethoxam (TMX) is a systemic neonicotinoid widely used for pest control in several agricultural crops. TMX mimics the action of acetylcholine causing uncontrolled muscular contraction eventually leading to insect death. TMX is being found in freshwater ecosystems at concentrations of up to 225µg/L. Still, chronic toxicity data for freshwater invertebrates is limited. Therefore, the aim of this study was to evaluate the acute and chronic effects (at organismal and biochemical levels) of TMX on the freshwater insect Chironomus riparius. C. riparius life history responses were significantly affected by TMX exposure, namely with a decrease in growth and delay in emergence. Concerning the biochemical responses, after a short exposure (48h) to TMX, our results showed that low concentrations of TMX significantly reduced CAT activity and LPO levels of C. riparius. No effects were observed in AChE, GST and ETS activities. Effects in terms of survival, development rates and biochemical responses of C. riparius exposed to low concentrations of TMX observed in this study suggest potential deleterious effects of this neonicotinoid on aquatic insects inhabiting freshwaters environments near agricultural areas.

Microplastics (MPs) are emerging contaminants of freshwater ecosystems. Once in aquatic systems, most of these plastic particles undergo processes of fragmentation, biofouling, and sedimentation, resulting in increased concentrations of smaller sized and irregularly-shaped particles in the sediment. High levels of MPs in freshwater sediments can denote a potential threat to benthic and sediment-dwelling organisms such as dipteran larvae. This study evaluates the ecotoxicological effect of three pools of irregularly-shaped polyethylene (PE) microplastics (pools containing 90% of the particles within 32–63 μm (size-class A), 63–250 μm (size-class B) and 125–500 μm (size-class C)), with concentrations ranging from 1.25 to 20 g Kg−1 sediment, on the dipteran Chironomus riparius life-history traits. After ten days of exposure, larvae ingested PE particles typically in the 32–63 μm range, even when 90% of the particles possessed higher size (i.e., in size-classes B and C) and the larvae mandible allowed the ingestion of such bigger-sized particles. Thus, the number of ingested particles was higher in size-class A, followed by B and C, and led to a significant reduction with similar magnitude on larval growth (Lowest Observed Effect Concentrations (LOEC) = 2.5 g Kg−1 sediment DW) and a significant delay on imagoes emergence (e.g., LOEC = 1.5 g Kg−1 sediment DW for females). The results from this study show that the ingestion and persistence of small-sized polyethylene microplastics caused significant impairments on life-history traits of C. riparius. Considering their role on freshwater food-webs and the potential persistence of small-sized PE particles in their larval gut, these results also point for the potential adverse effects of small-sized microplastics at the community and ecosystem level. This study was supported by FCT through national funds (PIDDAC), and the cofunding by the FEDER, within the PT2020 Partnership Agreement and Compete 2020 within the research project ComPET (POCI-01-0145-FEDER-030361) and CESAM (UID/AMB/50017). Also, by FCT and FSE/POCH, via a Post-Doc grant to A.L. Patricio Silva (SFRH/BPD/114870/2016), a Ph.D. grant to Carlos Silva (SFRH/BD/128134/2016) and a research contract under the program “Investigador FCT2015” to João L.T. Pestana (IF/01420/2015). published

Microbial insecticides are being used as ecologically-friendly alternatives to traditional insecticides. However, their effects have been poorly investigated on non-target freshwater species, with exception of a few insect species. Moreover, combined effects of microbial insecticides with other environmental stressors, such as salinity, have never been investigated. Thus, our goal was to assess the effects of Bac-Control® (based in Bacillus thuringiensis - Btk) and Boveril® (based in Beauveria bassiana - Bb) with increasing salinities (NaCl) on freshwater planarian Girardia tigrina. It has been reported that increased salinity levels affect freshwater organisms compromising their survival by triggering adaptation processes to cope with osmotic stress. Our results showed delayed regeneration, decreased locomotion and feeding on planarians exposed to NaCl, whereas their sexual reproduction was not affected. Both microbial insecticides impaired feeding, locomotor activity, regeneration, and sexual reproduction of planarians. Planarians exposed to microbial insecticides compromised their progeny. Therefore, microbial insecticides might not be ecologically friendly alternatives to chemical insecticides. Interestingly, harmful effects of microbial insecticides with increasing salinities showed an inadequate response of planarians to cope with induction of their immune response and osmoregulation.

Caddisflies have been included in ecotoxicological studies because of their sensitivity and ecological relevance. The present study aimed to assess the sublethal effects of an anthranilic diamide insecticide, chlorantraniliprole (CAP), to Sericostoma vittatum. Used worldwide, CAP is a persistent compound that has been found in surface waters at concentrations from 0.1 μg/L to 9.7 μg/L. It targets the ryanodine receptors, and the present ecotoxicological assessment focused on biomarkers related to neurotransmission, biotransformation, oxidative stress damage, and endpoints related to energy processing (feeding, energy reserves, and cellular metabolism). Six days of exposure trials revealed that feeding activity was significantly decreased in S. vittatum larvae exposed to 0.9 μg/L CAP. Concomitantly, a reduction in cellular metabolism and a significant decrease in protein content were also observed in caddisfly larvae exposed to CAP, suggesting metabolic depression. The results show that sublethal concentrations of CAP can cause detrimental sublethal effects on S. vittatum total glutathione content at concentrations as low as 0.2 μg/L. Bioenergetics can be used to assess physiological effects of contaminants, and the present results show that exposure to low, environmentally relevant, concentrations of CAP alter energy acquisition and metabolism in nontarget aquatic insects with potential population level effects. Environ Toxicol Chem 2017;36:1584-1591. © 2016 SETAC.

Integration of conventional wastewater treatments with advanced oxidation processes (AOPs) has become of great interest to remove pharmaceuticals and their metabolites from wastewater. However, application of these technologies generates reactive oxygen species (ROS) that may reach superficial waters through effluents from sewage treatment plants. The main objective of the present study was to elucidate if ROS present in real effluents after biological and then chemical (single ozonation, solar photolytic ozonation, solar photocatalytic ozonation (TiO2, Fe3O4) and solar photocatalytic oxidation (TiO2)) treatments induce oxidative stress in Daphnia magna. For this, the activity of two antioxidant enzymes (superoxide dismutase and catalase) and the level of lipid peroxidation were determined in Daphnia. The results of oxidative stress biomarkers studied suggest that D. magna is able to cope with the superoxide ion radical (O2·-) present in the treated effluent due to single ozonation by mainly inducing the antioxidant activity superoxide dismutase, thus preventing lipid peroxidation. Lethal effects (measured in terms of immobility) were not observed in these organisms after exposure to any solution. Therefore, in order to probe the ecological efficiency of urban wastewater treatments, studies on lethal and sublethal effects in D. magna would be advisable.

Stream ecosystems face ever-increasing pressures by the presence of emergent contaminants, such as, personal care products. N, N-diethyl-3-methylbenzamide (DEET) is a synthetic insect repellent that is being found in surface waters environments in concentrations up to 33.4 μg/L. Information concerning DEET's toxicity in the aquatic environment is still limited and focused only on its acute effects on model species. Our main objective was to assess the effects of DEET exposure to a caddisfly non-target species using sub-lethal endpoints. For that, we chose Sericostoma vittatum, an important shredder in Portuguese freshwaters that has been already used in different ecotoxicological assays. Besides acute tests, S. vittatum were exposed during 6 days to a gradient of DEET concentrations (8, 18 and 40.5 mg/L) to assess effects on feeding behaviour and biochemical responses, such as, lipid peroxidation levels (LPO), catalase and acetylcholinesterase (AChE) activities, and also assess effects on energy reserves and consumption. Acute tests revealed a 48 h-LC50 of 80.12 mg/L and DEET exposure caused feeding inhibition with a LOEC of 36.80 mg/L. Concerning the biochemical responses, DEET caused no effects in LPO nor on catalase activity. A non-significant decrease in AChE activity was observed. Regarding energetic reserves, exposure to DEET caused a significant reduction in S. vittatum carbohydrates levels. These results add important information for the risk assessment of insect repellents in the aquatic environment and suggest that reported environmental concentrations of DEET are not toxic to non-target freshwater insects.

Paraquat (PQ) is still used in several countries worldwide as an herbicide for weed control in agricultural production, ponds, reservoirs and irrigation canals. Thus, PQ is frequently found in surface water systems and is potentially toxic to aquatic organisms, since it can cause mitochondrial dysfunction altering in the redox state of cells. This study aimed to investigate the chronic effects of PQ to Chironomus riparius and Girardia tigrina, and compare their physiological strategies to cope with environmental stress. The mean emergence time was the most sensitive endpoint for Chironomids, with the lowest observed effect concentrations (LOEC) being 0.02 for males and 0.1 mg PQ L−1 for females. Moreover, PQ reduced the body weight of male and female imagoes, with LOECs of 0.5 and 2.5 mg PQ L−1, respectively. Paraquat also decreased the respiration rate (LOEC = 2.5 mg PQ L−1) and total glutathione (tGSH) content (LOEC = 0.5 mg PQ L−1). Thus, the aerobic production of energy was not affected and allowed chironomids to cope with oxidative stress induced by PQ, but with consequent physiological costs in terms of development rates and weight of adults. In planarians, PQ decreased the locomotion and feeding activity, and delayed photoreceptor regeneration (LOECs = 2.5 mg PQ L−1 for all endpoints). Despite increased aerobic energy production (LOEC = 0.5 mg PQ L−1), planarians were not able to cope with oxidative stress induced by the highest PQ concentrations, since lipid peroxidation levels were significantly increased (LOEC = 2.5 mg PQ L−1) concomitantly with a significant decrease of tGSH (LOEC = 2.5 mg PQ L−1). These results showed that planarians were unable to cope with oxidative stress induced by PQ with consequent impairments of behavior and regeneration despite an increased aerobic energy production.