Organic contamination and multi-biomarker assessment in watersheds of the southern Brazil: an integrated approach using fish from the Astyanax genus.

We aimed to examine the responses of pollution biomarkers in feral fish from Astyanax genus collected at three hydrographic regions in southern Brazil and the capacity of these tools to differentiate between various levels of contamination. To achieve this, levels of organochlorine pesticides (liver), as well as the biomarkers AChE (muscle and brain), TBARS (liver), and EROD (liver) were assessed. Collections were conducted in four municipalities (Alegrete, Caraá, Lavras, and Santa Vitória) during 1 year, encompassing winter and summer. Fish from Alegrete were the most contaminated overall, but animals sampled in Caraá, and Lavras also displayed elevated levels of current-use pesticides. Elevated levels of endosulfans, DDTs, HCHs, and current-use pesticides were accompanied by elevated levels of TBARS in the liver. Conversely, fish from Santa Vitória exhibited the highest levels of PAHs, accompanied by elevated levels of EROD in the liver and reduced levels of AChE in muscle and brain. TBARS proved to be a reliable biomarker for assessing impacts arising from pesticide accumulation, while EROD and AChE served as valuable indicators of impacts resulting from PAHs accumulation. Ultimately, the results obtained in this study demonstrate the reliable use of the proposed biomarkers for tracking biological impacts stemming from aquatic pollution using feral Astyanax as biomonitoring species.

The microbial profile of rivers and lagoons three years after the impact of the world's largest mining disaster (Fundão dam, Brazil).

The collapse of the Fundão tailings dam (Minas Gerais, Brazil) was the largest environmental disaster in Brazil's history and in the world mining industry. This disaster carried approximately 55 million m3 of iron ore tailings along the rivers and the lagoons of the Doce river basin. Although multiple studies assessed the impact on microbial communities in those rivers and lagoons right after the dam rupture, it is not known whether the microbiome in those environments remains impacted years after the disaster. Assessing the microbiome is very important to evaluate impacts and evaluate the health of the environment, due to the several ecological roles played by microorganisms. Here, we evaluated the impact of the dam failure on water and sediment bacteriome and archaeome by high-throughput next-generation sequencing. Samples were taken from two rivers and six lagoons during the dry and rainy seasons approximately three years post disturbance. The results showed a large number and abundance of microbial groups associated with the presence of heavy metals and mine tailings sediments. Some of these microorganisms were also reported in large abundance in the impacted rivers shortly after the Fundão dam rupture. Among the most abundant microorganisms in the Doce River, we can highlight the bacteria hgcI clade and the archaea Nitrososphera sp. in the water, and the bacteria Anaerolineaceae sp. in the sediment. These results suggest that the microbiome of the rivers and the lagoons in the Doce river basin remains severely impacted by the Fundão tailings dam failure even three years after the disaster. The presence of those microorganisms can also help to assess the occurrence of the Fundão dam sediment in other environments.

Ecotoxicological impacts of the Fundão dam failure in freshwater fish community: Metal bioaccumulation, biochemical, genetic and histopathological effects.

This study investigates the ecotoxicological impacts of the Fundão dam rupture, one of the major environmental disaster that occurred in Brazil and in the world mining industry history, through multi-biomarkers responses and metals bioaccumulation in the fish community of different trophic levels. Specimens of the fishes (omnivorous/herbivorous and carnivorous) were collected along the Doce River channel and its affluent Guandú River, and in different lakes and coastal lagoons adjacent to the river channel, in the Espirito Santo State, Southeast of Brazil. Four sampling collections were carried out over two years (2018 to 2020, during dry and rainy seasons). For both trophic groups the biomarkers responses indicated physiological alterations related to metals exposure and showed strong seasonal variations. The principal component analysis and integrated biomarker response index showed that DNA damage and lipid peroxidation were more associated with dry season 2 (Sep/Oct 2019) and the oxidative damage in proteins, metallothioneins concentration and the activity of superoxide dismutase in the gills showed a greater association with rainy season 2 (Jan/Feb 2020). On the other hand, the enzymes of energy metabolism, catalase and histological damage in the liver and the gills, were more associated with the dry and rainy campaigns of the first year of monitoring. The multivariate approach also suggested a temporal intensification in the bioaccumulation of metals and biological effects in the lacustrine environments. Thus, these results demonstrate that the release of mineral residues from the rupture of the Fundão mine dam affects the health status of the fish from the Doce River basin, provoking metals bioaccumulation, hepatic and branchial damage in the fish besides inducing of enzyme activity related to metal contamination, even four years after the rupture.

Impacts of tailings of Fundão dam (Brazil) rupture on marine fish: Metals bioaccumulation and physiological responses.

This study evaluated the impacts of the mining tailings after the rupture of the Fundão dam on fish communities on the Atlantic Ocean southeast coast. Four sample collections were carried out over two years (2018-2020), in seasonal periods. Omnivorous/herbivorous and carnivorous fish were collected for analysis of metal bioaccumulation, multibiomarkers of environmental contamination and histopathology. Metal bioaccumulation was stronger correlated in carnivorous fish in the dry-2018 collection, besides higher activity of antioxidant enzymes, energy metabolism and higher morphological damage; however, there was less oxidative damage and less metallothioneins induction, and these variables were strongly associated with the wet-2020 collection. In a temporal view, it was possible to observe a reduction in metal levels in fish, except in the mouth of the Doce River. These events can be explained by seasonal natural events, which tend the resuspension and boost metal levels, mainly in the mouth region during the rainy season.

Can the insecticide Imidacloprid affect the health of the Neotropical freshwater fish Astyanax altiparanae (Teleostei: Characidae)?

Female juveniles of the Neotropical fish Astyanax altiparanae were exposed for 96 h to four treatments containing the active ingredient from Imidacloprid® commercial formulation (IMI 1, IMI 2, IMI 3, and IMI 4) and to a control treatment (only dechlorinated tap water). Glutathione content, glutathione S-transferase activity, lipid peroxidation (LPO) and protein carbonylation levels, acetylcholinesterase (AChE) activity, and frequency of micronuclei and erythrocyte nuclear abnormalities (ENA) were measured in the fish. The muscle and gills were the most affected organs; their antioxidant defense was not enough to prevent oxidative damage (LPO) in the IMI 2 and IMI 4 treatment fish. IMI also inhibited AChE activity in the muscle (IMI 3 and IMI 4) and increased ENA frequency (IMI 4). IMI can affect the health of A. altiparanae in environmentally relevant concentrations, causing oxidative damage in different organs, neurotoxic effects in the muscle, and genotoxicity.

An integrated approach in subtropical agro-ecosystems: Active biomonitoring, environmental contaminants, bioaccumulation, and multiple biomarkers in fish.

Aquatic contamination in agricultural areas is a global problem, characterized by a complex mixture of organic and inorganic pollutants whose effects on biota are unpredictable and poorly investigated. In this context, in the present study, the Neotropical fish Prochilodus lineatus was confined in situ for 120 days in two sites with different levels of anthropic impact: 1) a fish hatchery station, within the State University of Londrina (reference site - REF) and 2) an agro-ecosystem area in one of the most productive regions of southern Brazil (experimental site - EXP). We evaluated multiple biomarkers at different levels of biological organization, such as biotransformation and antioxidant enzymes, oxidative damages, DNA damages and liver histopathology. We also evaluated the occurrence of 22 organochlorine pesticides (OCPs) and 6 trace metals in water and sediment; and 33 current-use pesticides (CUPs) in the water; besides the presence of OCPs in the liver and metals in different tissues of the confined fish. The chemical analysis confirmed that the two environments presented different levels of contamination. We verified a distribution gradient of data in the principal component analysis (PCA), separating the REF fish to one side and the fish at the agricultural area (EXP) to the other side. In general, the biomarker responses were more altered in fish from the EXP than fish from the hatchery station; and this fish presented a greater accumulation of endosulfan (an increase of 18× compared to basal value) and showed oxidative, genetic, and histological damage. Through the Biomarkers Response Index (BRI), we found that the EXP fish demonstrated a decrease in health status compared with the REF fish during the confinement time, due to their exposure to a higher concentration of contaminants. In conclusion, the use of multiple biomarkers at different response levels is an important tool for environmental monitoring.

The pyrethroid λ-cyhalothrin induces biochemical, genotoxic, and physiological alterations in the teleost Prochilodus lineatus.

The λ-cyhalothrin (CL) is a globally used pyrethroid insecticide that has been detected in different water bodies worldwide. However, studies on the effects of CL on freshwater fishes are still incipient. In this context, we evaluated the acute effects of a commercial formulation containing CL (Karate Zeon® CS 50) in juveniles of the teleost Prochilodus lineatus exposed for 96 h to four concentrations of the active ingredient (5, 50, 250 and 500 ng.L-1). Biochemical, physiological, and genotoxic biomarkers were evaluated in different organs of the fish. Exposure to CL induced significant changes in the enzymatic profiles of P. lineatus, with specific alterations in biotransformation enzymes and antioxidant defence in different tissues. Lipid peroxidation was observed in fish gills and kidney. Increases in esterases were observed in the liver of fish exposed to all CL concentrations evaluated, whereas acetylcholinesterase activity decreased in the muscles of fish at all concentrations. CL also promoted osmoregulatory disorders, with decreases in calcium and magnesium gill ATPases, with consequent hypocalcaemia, in addition an increase in sodium-potassium ATPase activity was observed in the gills of fish exposed to the highest CL concentration, probably in order to compensate a reduction in plasma sodium. Besides, increases in DNA damage were observed in the erythrocytes of fish exposed to all CL concentrations. Thus, despite the low CL concentrations and the short exposure time, this pyrethroid caused hematological adjustments, oxidative stress, osmoregulatory disorders, and DNA damage in P. lineatus, showing that the species is highly sensitive to the deleterious effects of CL.

DNA damage and oxidative stress induced by imidacloprid exposure in different tissues of the Neotropical fish Prochilodus lineatus.

Imidacloprid (IMI), a systemic neonicotinoid insecticide widely used in worldwide scale, is reported in freshwater bodies. Nevertheless, there is a lack of information about IMI sublethal effects on freshwater fish. Thus, the aim of this study was to identify the potential hazard of this insecticide to the South American fish Prochilodus lineatus exposed for 120 h to four IMI concentrations (1.25, 12.5, 125, and 1250 µg L-1). A set of biochemical, genotoxic and physiological biomarkers were evaluated in different organs of the fish. IMI exposure induced significant changes in the enzymatic profiles of P. lineatus, with alterations in the activity of biotransformation and antioxidant enzymes in different tissues. Redox balance of the tissues was affected, since oxidative damage such as lipoperoxidation (LPO) and protein carbonylation (PCC) were evidenced in the liver, gills, kidney and brain of fish exposed to different IMI concentrations. Fish exposed to all IMI concentrations showed decreased blood glucose indicating an increase of energetic demand. DNA damage was evidenced by the comet test, in the erythrocytes of fish all the concentrations evaluated. We integrated these results in the Integrated Biomarker Response (IBR) index, which evidenced that the organs most affected by IMI exposure were the liver and kidney, followed by the gills. Our results highlight the importance of investigating different target tissues after IMI exposure and show the sublethal effects of IMI in some of them; they also warn to the possible consequences that fish living in freshwater ecosystems can suffer due to IMI exposure.

A comparative approach using biomarkers in feral and caged Neotropical fish: Implications for biomonitoring freshwater ecosystems in agricultural areas.

The aim of this study was to investigate the responses of biomarkers in feral and caged fish and the capacity of these biomarkers to discriminate contamination levels along a stream located in an agricultural area in Southern Brazil. Specimens of the Neotropical fish, Astyanax altiparanae, were confined for 168h in three lakes along the stream. Additionally, during the weeks of in situ exposure, wild specimens of this species were collected from the same sites. Biochemical biomarkers were analyzed, such as phase I biotransformation enzyme 7-ethoxyresorufin-O-deethylase (EROD) and phase II biotransformation enzyme glutathione S-transferase, and we also determined hepatic and branchial levels of non-protein thiols (NPSH), oxidative damage such as lipid peroxidation (LPO), and acetylcholinesterase (AChE) activity in muscle and brain. Genetic biomarkers such as DNA breaks (comet assay), frequency of micronuclei (MN) and erythrocytic nuclear abnormalities (ENA) were also examined. The results indicate that the most sensitive biomarkers for discriminating contamination levels are DNA breaks, LPO and AChE activity. Similar results were obtained for both caged and feral fish. The biomarkers that reflect the results of cumulative events, such as ENA, were more discriminative for chronically exposed specimens (feral fishes). Analyzing biomarkers using an integrated response index showed that both approaches (using feral and caged A. altiparanae) were effective for discriminating contamination levels along the stream, corroborating the results of chemical analyses for selected pesticides. Taken together, these results highlight the importance of biomarker selection and show that both approaches (caged and feral fish) are satisfactory for evaluating water quality in streams impacted by agricultural activities.

Metals bioaccumulation and biomarkers responses in the Neotropical freshwater clam Anodontites trapesialis: Implications for monitoring coal mining areas.

As one of the most impactful industries, coal mining can promote several alterations at surrounding environment. In surface water, elevated concentrations of metals like Mn, Zn, Fe and Al are often observed. Thus, the aim of this study was to investigate the bioaccumulation and the sub-lethal effects of these metals on various organs of the Neotropical bivalve Anodontites trapesialis confined along a stream located near a coal mine, in order to assess a set of biomarkers that could be used for effectively monitoring coal mining areas. Clams were caged, for 96h, at two sites located upstream (Up1 and Up2) and two sites downstream (Dw1 and Dw2) from the mine. Metals bioaccumulation was determined in gills, mantle, digestive gland, muscle and hemolymph and the following biomarkers were measured in A. trapesialis tissues: total antioxidant capacity against peroxyl radicals, metallothionein content, lipid peroxidation (LPO), proteins carbonylation, glutathione S-transferase activity, superoxide dismutase activity and acetylcholinesterase (AChE) activity. The results showed that Al and Fe bioaccumulation in the gills and hemolymph, Al bioaccumulation in the mantle and muscle, increased LPO in the gills (Dw1 and Dw2) and mantle (Dw1), as well as reduced AChE activity in the muscle (Dw1 and Dw2) should be considered effective biomarkers for monitoring coal mining areas. A. trapesialis proved to be an efficient biological model, considering that biomarkers responses were observed in the clams after only 96h of confinement at Dw sites, accordingly this species could be a good candidate for monitoring Neotropical freshwaters.

Multiple biomarker responses in Prochilodus lineatus subjected to short-term in situ exposure to streams from agricultural areas in Southern Brazil.

In order to assess the quality of streams susceptible to contamination by pesticides we apply biochemical and genotoxic biomarkers in the Neotropical fish Prochilodus lineatus submitted to in situ tests. Fish were caged, for 96 h, in two streams located in areas with intensive use of pesticides, the Apertados (AP) and the Jacutinga (JC), and in a small stream (Godoy stream ­ GD) found inside a forest fragment adjacent to a State Park. Biochemical parameters, such as biotransformation enzymes 7-ethoxyresorufin-O-deethylase (EROD) and glutathione-S-transferase (GST), non-protein thiols (NPSH), lipoperoxidation (LPO), protein carbonylation (PCO) and acetylcholinesterase (AChE) were evaluated in various fish organs, as well as genotoxic biomarkers (damage to DNA and occurrence of micronuclei and erythrocyte nuclear abnormalities). Samples of water and sediment were collected for analysis of metals (Cu, Cr, Pb, Ni, Mn, Cd and Zn), organochloride pesticides, and triazine and glyphosate herbicides. We observed an increase in liver GST activity in fish at AP and gill GST activity in fish at JC. An increase in liver LPO was also observed in fish exposed to AP and JC. The same animals also exhibited increased DNA damage and erythrocyte nuclear abnormalities (ENAs) compared to the fish kept in GD. A number of compounds showed concentrations higher than the permitted levels, in particular, dichlorodiphenyltrichloroethane (DDT), its metabolites dichlorodiphenyldichloroethylene (DDE) and dichlorodiphenyldichloroethane (DDD), hexachlorocyclohexanes (HCH), heptachloride, diclofluanid and aldrins. These pesticides were detected at higher concentrations in water and sediment samples from AP, followed by JC and GD. The Integrated Biomarker Response Index (IBR) indicated that AP and JC (AP: 21.7 > JC: 18.5 > GD: 12.6) have the worst environmental quality. Integrated biomarker analysis revealed that the alterations observed related well with the levels of environmental contaminants, demonstrating the effectiveness of this biomonitoring approach.

Integrated biomarker response index using a Neotropical fish to assess the water quality in agricultural areas

Aquatic ecosystems in areas with intense agricultural activity are subject to pesticide contamination, which may compromise the health of the fish. In order to verify the quality of the water and the possible effects of pesticides on fish, a method that combines different biomarker responses into an index named "integrated biomarker response" (IBR) was applied using the biological alterations in the Neotropical fish Astyanax altiparanae. Fish were maintained in situ at five sites along a stream that runs in an agricultural area and in a stream within a forest fragment, considered a reference site. After seven days of exposure the following alterations were observed in fish confined at experimental sites: increased activity of glutathione-S-transferase (GST) and catalase (CAT) and increase in the content of reduced glutathione (GSH) in liver and gills, reduction of acetylcholinesterase (AChE) activity in the brain and muscle, increase in the occurrence of DNA strand breaks and in the frequency of micronuclei (MN) and nuclear abnormalities (ENA) in erythrocytes. The IBR highlighted three sites as the most affected, as the animals confined at these sites showed greater variations in biological responses. The biomarkers most important for the IBR results were GST, AChE, DNA breaks and ENA. Ecossistemas aquaticos inseridos em areas com intensa atividade agrmcola estco susceptmveis ` contaminagco por pesticidas, os quais podem comprometer a sazde dos peixes. A fim de verificar a qualidade da agua e os possmveis efeitos de pesticidas sobre peixes, um mitodo integrando o uso de diferentes biomarcadores, denominado mndice integrado de respostas de biomarcadores (IBR), foi aplicado utilizando-se as alteragues biolsgicas avaliadas em varios srgcos do peixe neotropical Astyanax altiparanae. Os peixes foram confinados in situ em cinco pontos ao longo de um ribeirco localizado em area de produgco agrmcola e em um csrrego dentro de um fragmento florestal, considerado um local de referjncia. Apss sete dias foram observadas as seguintes alteragues nos peixes confinados nos pontos experimentais: aumento da atividade da glutationa-S-transferase (GST) e catalase (CAT) e aumento do contezdo de glutationa-reduzida (GSH) em fmgado e brbnquias, redugco da atividade da acetilcolinesterase (AChE) no mzsculo e cirebro, aumento de quebras no DNA e na frequjncia de micronzcleos (MN) e alteragues nucleares (ENA) em eritrscitos. O IBR destacou trjs dos pontos estudados como os mais afetados pela contaminagco agrmcola, uma vez que foram observados nos animais confinados nestes locais maiores variagues nas respostas biolsgicas. Os biomarcadores mais significativos para os resultados de IBR foram a GST, AChE, quebras no DNA e ENA.