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Chemical exposure during the early life stages of development may have long lasting effects on organisms that are rarely studied. The present work intended to evaluate the effect of embryonic exposure to the pesticide carbaryl on adult fish behavior. Zebrafish (Danio rerio) embryos were exposed, for 4 days, to sublethal concentrations of carbaryl (0.01, 0.1 and 1.0 mg/L) plus a control and then kept in standard cultivation conditions until adulthood. A battery of behavioral tests was then performed to assess anxiety-like behavior (locomotor activity, thigmotaxis and novel tank diving test), social behavior, and feeding. Developmental exposure of zebrafish to sublethal concentrations of carbaryl produced important behavioral alterations in the adulthood. Main effects included decreased locomotion/hypoactivity (increase in slow movements and decrease of medium and rapid movements), especially in the light periods. Moreover, spatial pattern also changed: while during dark periods control fish increased activity in the outer zone of the tank, this was not observed in exposed fish. Overall, this demonstrated the importance of life stage exposure, clearly demonstrating long lasting effects of a (chemical) stress event at embryonic stages. This data supports the need of considering this scenario in environmental risk evaluations. Further work should focus on the mechanistic effects of developmental disruption responsible for the effects observed. • The effect of embryonic exposure to the pesticide carbaryl was assessed on adult fish behavior. • Exposure at embryonic stages affected adult behavior. • Exploratory behavior, feeding and social behavior may reflect embryonic exposure to pesticides. [ABSTRACT FROM AUTHOR]

Abstract Oxytetracycline (OTC) is a broad-spectrum antibiotic widely used in livestock production. Like many other pharmaceuticals, OTC is not completely metabolized by the organism and thus, increasing amounts of the compound are being detected in the aquatic environment. The assessment of the environmental risk of pharmaceuticals is hindered by their very low concentrations and specific modes of action and thus relevant exposure scenarios and sensitive endpoints are needed. Thus, this work aimed to study the long-term effect of OTC exposure in zebrafish (at behavior and biochemical levels) and associated bacterial communities (fish gut and water bacterial communities). Results revealed that at behavioral level, boldness increase (manifested by increased exploratory behavior of a new environment) was observed in fish exposed to low OTC concentrations. Moreover, changes in fish swimming pattern were observed in light periods (increased stress response: hyperactivity and freezing) probably due to photo-sensibility conferred by OTC exposure. Effects at biochemical level suggest that long-term exposure to OTC interfere with cellular energy allocation mainly by reducing lipids levels and increasing energy consumption. Moreover, evidences of oxidative damage were also observed (reduced levels of TG, GST and CAT). The analysis of water and gut microbiome revealed changes in the structure and diversity of bacterial communities potentially leading to changes in communities' biological function. Some of the effects were observed at the lowest concentration tested, 0.1 μg/L which is a concentration already detected in the environment and thus clearly demonstrating the need of a serious ecotoxicological assessment of OTC effects on non-target organisms. Graphical abstract Image 1 Highlights • Zebrafish exposed to oxytetracycline present different feeding behavior pattern. • Oxytetracycline induced fish boldness and hyperactivity in light periods. • Energetic reserves and oxidative stress enzymes decrease after long-term exposure. • Bacterial communities of fish gut and exposure water were affected by oxytetracycline. [ABSTRACT FROM AUTHOR]

Abstract Climate change is expected to alter the dynamics of water masses, with consequent changes in water quality parameters such as dissolved organic carbon (DOC) concentration. DOC levels play a critical role in the fate of organic chemicals, influencing their bioavailability and toxicity to aquatic organisms. This study aimed to evaluate the effects of DOC, particularly humic acids (HA), in the toxicity of gemfibrozil (GEM) - a human pharmaceutical frequently detected in surface waters. Lethal and sublethal effects (genotoxic, biochemical and behavioural alterations) were evaluated in zebrafish embryos exposed to several concentrations of GEM and three HA levels, in a full factorial design. HA significantly increased GEM LC 50 values, mainly in the first 72 h of exposure, showing a protective effect. At sublethal levels, however, such protection was not observed since HA per se elicited adverse effects. At a biochemical level, individual exposure to HA (20 mg/L) elicited significant decreases in cholinesterase and glutathione S -transferase activities. Regarding behaviour, effects of individual exposure to HA appear to surpass the GEM effects, reducing the total distance moved by larvae. Both GEM and HA significantly increased DNA damage. Hence, this study demonstrated that abiotic factors, namely HA, should be considered in the assessment of pharmaceuticals toxicity. Moreover, it showed that lethality may not be enough to characterize combined effects since different patterns of response may occur at different levels of biological organization. Testing sublethal relevant endpoints is thus recommended to achieve a robust risk assessment in realistic scenarios. Highlights • Humic acids (HA) increased gemfibrozil LC 50 to zebrafish embryos. • The HA protective role observed at lethal level was not verified at sublethal levels. • HA per se affected biochemical, behavioural and genotoxic parameters. • Testing endpoints at different levels of biological organization is recommended. • Dissolved organic carbon should be considered when assessing the toxicity of pharmaceuticals. [ABSTRACT FROM AUTHOR]

Pharmaceuticals are emerging contaminants as their worldwide consumption increases. Fibrates such as gemfibrozil (GEM) are used in human medicine to reduce blood concentrations of cholesterol and triacylglycerol and also are some of the most frequently reported pharmaceuticals in waste waters and surface waters. Despite some studies have already demonstrated the negative impact in physiological and/or reproductive endpoints in adult fish, data on survival and behavioral effects in fish larvae are lacking. This study aimed to assess the effects of GEM on zebrafish eleutheroembryo development and locomotor behavior. A fish embryo toxicity (FET) test was undertaken to evaluate GEM acute toxicity by exposing embryos to 0, 6.58, 9.87, 14.81, 22.22, 33.33 and 50 mg/L. Developmental endpoints such as hatching success, edemas and malformations were recorded. A second test was undertaken by exposing embryos to 0, 1.5, 3 and 6 mg/L in order to evaluate the effects of GEM on 120 and 144 h post fertilization (hpf) larvae locomotor activity by video tracking, using a Zebrabox ® (Viewpoint, France) device. From the data recorded, several parameters such as total swimming distance (TSD) and total swimming time (TST) in each 120 s integration time were calculated. Data showed that this compound has a moderate toxic effect on fish embryo development, affecting both survival and hatching success with a calculated 96 h LC 50 of 11.01 mg/L and no effects at the developmental level at 6 mg/L. GEM seems to impair locomotor activity, even at concentrations where developmental abnormalities were unperceived, at concentrations as low as 1.5 mg/L. Both TSD and TST were sensitive to GEM exposure. These effects do not seem to be independent of the developmental stage as 120 hpf larvae seem to present a development bias with repercussions in locomotor behavior. This study highlights the need to include behavioral endpoints in ecotoxicological assays as this seems to be a more sensitive endpoint often disregarded. [ABSTRACT FROM AUTHOR]

This study aimed to assess the effect of ultraviolet radiation (UVR) and chemical stress (triclosan-TCS; potassium dichromate-PD; prochloraz-PCZ) on bacterial communities of zebrafish ( Danio rerio ) embryos (ZEBC). Embryos were exposed to two UVR intensities and two chemical concentrations not causing mortality or any developmental effect (equivalent to the No-Observed-Effect Concentration-NOEC; NOEC diluted by 10-NOEC/10). Effects on ZEBC were evaluated using denaturing gradient gel electrophoresis (DGGE) and interpreted considering structure, richness and diversity. ZEBC were affected by both stressors even at concentrations/doses not affecting the host-organism (survival/development). Yet, some stress-tolerant bacterial groups were revealed. The structure of the ZEBC was always affected, mainly due to xenobiotic presence. Richness and diversity decreased after exposure to NOEC of PD. Interactive effects occurred for TCS and UVR. Aquatic microbiota imbalance might have repercussions for the host/aquatic system, particularly in a realistic scenario/climate change perspective therefore, future ecotoxicological models should consider xenobiotics interactions with UVR. [ABSTRACT FROM AUTHOR]

Oxytetracycline (OTC) is a broad-spectrum antibiotic widely used in aquaculture, resulting in contamination of aquatic environments. In a previous study, we observed significant effects of OTC sublethal concentrations in zebrafish, its microbiome and the water bacterial community. Here we assessed the extent to which these effects are reversible after a recovery period. Zebrafish adults were exposed to OTC (10,000 μg/L) via water exposure. Effects were analyzed at 5 days (5 dE) and 2 months (2 mE) of exposure and recovery was assessed at 5 days (5dPE) and 1 month (1mPE) after exposure Impacts were observed in fish energetic reserves and in fish and water microbiomes structure, being significant even at 5 dE. At energetic reserves level, the effect in cellular energy allocation (CEA) was dependent on the exposure time: initially CEA increased while after 2 mE CEA decreased. At microbiome level, diversity was not affected but the richness of the water microbiome significantly decreased at 2 mE. Regarding the post-exposure period, at CEA level, organisms seem to recover. In water and gut microbiomes OTC effects were also attenuated after exposure ceases, indicating a recovery. Even so, the structure of water exposed community remained significantly different towards the control, while richness of this community significantly increased at 1mPE. During exposure the relative abundance of 11 and 16 genera was significantly affected in the gut and water microbiomes, respectively, though these numbers decreased to 4 and 8 genera in the post-exposure period. At functional level during exposure 12 and 13 pathways were predicted to be affected in zebrafish gut and water microbiomes respectively, while post-exposure few pathways remained significantly affected. Hence, our results suggest a recovery of the fish fitness as well as of the water and intestine microbiomes after exposure ceases. Even so, some of the effects caused by OTC remain significant after this recovery period. Image 1 • Zebrafish adults were exposed to OTC for 2 months and allowed to recover for 1 month. • OTC effects were observed after 5 days and 2 months of exposure. • After a post-exposure period, fish recovered at energetic reserves level. • Zebrafish gut microbiome also recovered at structural and functional levels. • Water microbiome also recovered but OTC effects were still significant after 1 month. Oxytetracycline impacts both fish energetic reserves and fish gut and water microbiome. After exposure ceased, a re-adjust of fish fitness and water microbiome occurs. [ABSTRACT FROM AUTHOR]

Pantanal is a biome characterized by an extraordinary diversity and abundance of wildlife and houses several endemic species such as the freshwater shrimp Macrobrachium pantanalense . However, the increase in agriculture and husbandry activities in the region has contributed with residues of pesticides reaching aquatic systems. The main objective of this study is to assess the sensitivity of the endemic shrimp M. pantanalense compared with other freshwater species: the shrimp M. amazonicum , the crustacean Daphnia similis and the fish Danio rerio . The sensitivity of these organisms was assessed through acute exposure to copper and cypermethrin (through the formulation Barrage ® , widely used in Pantanal). For copper the species sensitivity decreased in the following order: D. similis (48 h-EC 50 0.051 mg/L) > M. pantanalense > D. rerio > M. amazonicum (48 h-LC 50 26.34 mg/L). Copper caused reduced length of shrimps and zebrafish and reduced heartbeat of zebrafish embryos. For cypermethrin the species sensitivity decreased in the following order: M. pantanalense (96 h-LC 50 0.05 μg/L) > M. amazonicum > D. similis > D. rerio (144 h-LC 50 1680 μg/L). Major effects of cypermethrin included reduced length of shrimps and zebrafish, as well as early hatching and increased incidence of developmental deformities in zebrafish embryos. This study highlights the importance of using endemic species for risk evaluations in sensitive biomes such as Pantanal. Moreover, it emphasizes the importance of testing pesticides toxicity as commercial formulations. Furthermore, we suggest that the endemic shrimp species M. pantanalense can be successfully used as a test species in ecotoxicology. [ABSTRACT FROM AUTHOR]

Carbendazim is a widely used broad spectrum benzimidazole fungicide; however, its effects to non-target aquatic organisms are poorly studied. The aim of this study was to investigate the toxic effects of carbendazim to zebrafish early life stages at several levels of biological organization, including developmental, biochemical and behavioural levels. The embryo assay was done following the OECD guideline 236 and using a concentration range between 1.1 and 1.8 mg/L. Lethal and developmental endpoints such as hatching, edemas, malformations, heart beat rate, body growth and delays were assessed in a 96 h exposure. A sub-teratogenic range (from 0.16 to 500 μg/L) was then used to assess effects at biochemical and behavioural levels. Biochemical markers included cholinesterase (ChE), glutathione-S-transferase (GST), lactate dehydrogenase (LDH) and catalase (CAT) and were assessed at 96 h. The locomotor behaviour was assessed using an automated video tracking system at 120 h. Carbendazim (96 h-LC 50 of 1.75 mg/L) elicited several developmental anomalies in zebrafish embryos with EC 50 values ranging from 0.85 to 1.6 mg/L. ChE, GST and LDH activities were increased at concentrations equal or above 4 μg/L. The locomotor assay showed to be extremely sensitive, detecting effects in time that larvae spent swimming at concentrations of 0.16 μg/L and thus, being several orders of magnitude more sensitive that developmental parameters or lethality. These are ecological relevant concentrations and highlight the potential of behavioural endpoints as early warning signs for environmental stress. Further studies should focus on understanding how the behavioural disturbances measured in these types of studies translate into fitness impairment at the adult stage. [ABSTRACT FROM AUTHOR]

• Primary cardiac lymphomas are extremely rare entities, usually behave aggressively and are associated with high mortality rates. • The use of several imaging tests may aid in the aetiological assessment and allow the detection of a possible biopsy site. • In HIV positive patients, the presence of a cardiac mass should raise the suspicion for high-grade B-cell lymphomas and proper treatment may significantly improve survival rates. A 65-year-old man with previous history of smoking, controlled HIV infection, treated hepatitis B infection, and type III cryoglobulinemia, was admitted due to right heart failure symptoms and significant weight loss. Despite being haemodynamically stable, he had periods of 1:1 conduction atrial flutter and presented with respiratory alkalosis and metabolic acidosis, as well as acute kidney and hepatic dysfunction, elevated D-dimer and cardiac markers. He underwent imaging with chest computed tomography and echocardiogram that confirmed pulmonary embolism and most notably revealed a significant sized cardiac mass causing almost complete obstruction of the right chambers, with no cleavage plane with the myocardial walls and tricuspid valve. Cardiac magnetic resonance was highly suggestive of malignancy. Cardiac surgery for mass excision and endomyocardial biopsy for diagnosis were considered, but the patient died with obstructive shock unresponsive to medical treatment. The autopsy revealed a primary unspecified diffuse large B-cell lymphoma. [ABSTRACT FROM AUTHOR]

The Pantanal (Brazil) is a wetland region characterized by seasonal flooding. Hydrological cycles influence the water physicochemical parameters, causing seasonal variations in pH and nitrites. Thus, the objective of this study was to evaluate the effects of varying pH and nitrite concentrations on the toxicity of the cypermethrin-based pesticide Barrage®, considering both lethal (mortality) and sublethal endpoints (growth and development). Larvae of the endemic shrimp Macrobrachium pantanalense and of the estuarine Amazonian congener Macrobrachium amazonicum were exposed to cypermethrin (through Barrage®) under several pH levels (6.5, 7.5 and 8.5) or nitrite concentrations (0.1, 0.2 and 0.4 mg/L). The pH had direct effects on all the tested endpoints for both species. For M. pantanalense , the lethal effects of the cypermethrin formulation were more pronounced at low pH (96-h LC 50 = 0.004 μg/L at pH 6.5, and 0.146 μg/L at pH 8.5). For M. amazonicum, an opposite response was observed, with increased toxicity of the formulation at high pH (96-h LC 50 = 0.110 μg/L at pH 6.5 and 0.044 μg/L at pH 8.5). Variations in pH also seemed to modify the sublethal effects of the formulation on larval growth and development of M. pantanalense. Nitrite concentrations affected larval growth of both species, modifying also the effects of the cypermethrin formulation on the larval development of M. amazonicum. This work shows the importance of considering abiotic factors for risk assessment either due to possible direct effects on the physiology of organisms and/or due to interactions with other stressors, particularly in fragile biomes such as Pantanal. • Barrage (B) is a cypermethrin-based pesticide widely used in Pantanal (Brazil). • Combined effects of B allied to pH or nitrites were tested using Macrobrachium sp. • pH had marked direct effects on mortality, growth and development of shrimp larvae. • pH and nitrites changed the sublethal toxicity of B (growth and larvae development). • Dissimilar responses among M. pantanalense and M. amazonicum. [ABSTRACT FROM AUTHOR]

Abstract In recent years, the need for the development of alternative test methods for the conventional acute fish toxicity test (AFT) with adult fish has often been discussed. In addition, concerns have been raised on the potential risks related with environmentally realistic pesticide mixtures since risk evaluations have traditionally been based on individual pesticides. The insecticide/acaricide abamectin and the fungicide difenoconazole are the main pesticides that are intensively used in Brazilian strawberry crop and are hence likely to occur simultaneously in edge-of-field waterbodies. The aim of the present study was therefore to evaluate the lethal and sublethal toxicity of single and mixture exposures of these pesticides to zebrafish early life stages (embryos and juveniles). By comparing the derived toxicity data of the individual compounds with that previously determined for zebrafish adults, the order of life stage sensitivity was juvenile > adult > embryo. The pesticide mixture revealed a dose-level dependent deviation of the independent action model, with antagonism at low dose levels and synergism at high dose levels. Sublethal parameters (especially those related with locomotion) were considerably more sensitive than lethality. Subsequently, the inclusion of sublethal parameters may greatly improve the sensitivity of FET tests and hence its suitability as a substitution of adult fish testing in risk assessment evaluations. Highlights • The order of zebrafish life stage sensitivity was juvenile > adult > embryo. • The mixture revealed antagonism at low dose levels and synergism at high dose levels. • Sublethal parameters (related with locomotion) are more sensitive than lethality. [ABSTRACT FROM AUTHOR]