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The microbiological diversity of cultivable bacteria was analyzed in an aerated facultative lagoon. The removal of specific compounds and measures of pollutant load was evaluated with isolated native bacteria, selected and identified in kraft cellulose effluent. The system was operated with an organic loading rate of 0.2 kgCODm-3d-1 for 60 days. Analyses of the fluorescence excitation-emission matrix, acute ecotoxicity, and microbiology were performed. Bioaugmentation tests were done to emphasize the removal of color, using promising species. The removals of biochemical oxygen demand, chemical oxygen demand, and total organic carbon in AFL were 94%, 51%, and 41%, respectively. Regarding color, removal was up to 4%, and the total phenolic compounds were not removed through biological treatment. The treatment also decreased turbidity by 94% and lignin derivatives by 12%. The bacteria identified through NCBI-BLAST and statistical similarity totaled 9 species in the cellulose effluent, three of which have the potential for color treatment: Bacillus cereus, Bacillus thuringiensis, and Paenibacillus sp. The Bacillus cereus combined with biomass removed color (69%), total phenolic compounds (37%), and compounds derived from lignin (53%). These species are promising for removing specific parameters combined with biomass from biological AFL treatment systems.

Herbicide mixture is used as an alternative to obtain different mechanisms of action acting on weeds, resulting in the frequent presence of pesticides in environmental compartments. As they are products used worldwide, this study evaluated effects of environmentally relevant concentrations of the analytical standards and commercial formulations of the herbicides atrazine (2 µg L− 1) and glyphosate (65 µg L− 1), in isolation and also in mixture (2 + 65 µg L− 1) on the microcrustacean Daphnia magna. Through chronic exposure (21 days) of two generations of organisms, effects on survival and reproductive capacity were observed, as well as responses regarding oxidative stress, determined through the analysis of biochemical biomarkers such as catalase and glutathione S-transferase. In the evaluation of the first generation of test organisms, no significant results related to biochemical biomarkers were observed, only effects over sexual maturation of organisms. However, in the second generation of exposed organisms, changes were observed in all parameters evaluated, with the mixture of herbicide active principles being the treatment responsible for more significant responses (p

Since 2008, Brazil has been the largest consumer of agrochemicals in the world, using pesticides to combat pests and vectors, impacting both microbiota and human health. Aiming at the degradation of the contaminants present in the environment, treatments by advanced oxidative processes are based on the synthesis of free radicals that allow the degradation of the pollutant. Among these processes are included UV/H 2 O 2 . By evaluating the ecotoxicity and phytotoxicity of the commercial Malathion® 500 CE product, using Aedes aegypti larvae and Lactuca sativa seed, the high toxicity of this formulation was observed. EC 50 values for A. aegypti being equal to 0.4 μg L -1 and for L. sativa equal to 550 μg L -1 . The commercial agrochemical degradation was carried out by UVC radiation and UV/H 2 O 2 , and the toxicity was evaluated after 30 and 120 minutes. After both treatments, the percentage of immobility was zero for A. aegypti , inferring the efficiency of the processes. However, for L. sativa, no treatment was able to remove or reduce initial toxicity.

Cylindrospermopsin (CYN) is a cyanotoxin that is cytotoxic to a wide variety of cells, particularly to the hepatocytes. In this study, the toxic effects of purified CYN were investigated in primary cultured hepatocytes of Neotropical fish Hoplias malabaricus. After isolation, attachment, and recovery for 72 h, the cells were exposed for 72 h to 0, 0.1, 1.0, 10, and 100 μg l−1 of CYN. Then, cell viability and a set of oxidative stress biomarker responses were determined. Catalase, superoxide dismutase, glucose-6-phosphate dehydrogenase, and glutathione S-transferase activities were not affected by exposure to CYN. Concentration-dependent decrease of glutathione reductase activity occurred for most CYN-exposed groups, whereas non-protein thiol content increased only for the highest CYN concentration. Lipid peroxidation, protein carbonylation, and DNA damage levels were not altered, but reactive oxygen species levels increased in the cells exposed to the highest concentration of CYN. Cell viability decreased in all the groups exposed to CYN. Thus, CYN may cause a slight change in redox balance, but it is not the main cause of cell death in H. malabaricus hepatocytes.

Malathion is a highly toxic organophosphate insecticide, being one of the most widely used in the world and is generally used for insect control in food production. Thus, ecotoxicological studies have been used to verify its toxic effects on aquatic organisms, such as Daphnia magna and biomarkers, as the comet assay. The comet assay is a microgel electrophoresis method for the detection and quantification of DNA strand breaks in individual cells. Cells were obtained from Daphnia magna after disaggregation of newborn organisms, exposed at concentrations of 0.23 μg L−1 and 0.47 μg L−1 for 48 h. Malathion has shown to cause damage to DNA of the exposed organisms. It was also observed the need of further studies to standardize the comet assay technique for Daphnia magna, once methodologies used present several differences.

Pyroligneous acid (PA) is a by-product of bio-oil, which is obtained by pyrolysis of the wood. This product has been tested for use in several areas, such as agriculture, as a promising green herbicide; however, there are few scientific data regarding its environmental impacts. For this study, an ecotoxicity testing battery, composed of Daphnia magna acute toxicity test, Allium cepa test and in vitro Comet assay with the rainbow trout gonad-2 cell fish line (RTG-2) were used to evaluate the acute toxicity and genotoxicity of PA obtained from fast pyrolysis of eucalyptus wood fines. The PA presented acute toxicity to D. magna (microcrustacea) with EC50 of 26.12 mg/L, and inhibited the seed germination (EC50 5.556 g/L) and root development (EC50 3.436 g/L) of A. cepa (higher plant). No signs of genotoxicity (chromosomal aberrations and micronuclei in A. cepa and primary DNA lesions in RTG-2 cells) were detected to this product. The acute toxicity and absence of genotoxicity may relate to the molecules found in the PA, being the phenolic fraction the key chemical candidate responsible for the toxicity observed. In addition, daphnids seem to be more sensitivity to the toxicity of PA than higher plants based on their EC50 values. This first ecotoxicological evaluation of PA from fast pyrolysis pointed out the need of determining environmental exposure limits to promote the safer agriculture use of this product, avoiding impacts to living organisms.

Astyanax is widely used in ecotoxicological bioassays and in biomonitoring studies because of the similarities that different species of the genus have regarding their ecological, morphological and behavioral characteristics. Due to these similarities, ecotoxicological studies are commonly conducted using any sampled Astyanax as if they were the same, i.e. without separating the species and using any gathering as a whole representative of the genus. This conduct is worrying since there are no studies showing the ecotoxicological relationship among the species. Therefore, this article addressed to this issue by inducing, through acute water exposure bioassays, the ecotoxicological responses of biomarkers of three species of Astyanax (A. altiparanae, A. bifasciatus and A. ribeirae). The antioxidant systems were induced by Cu contamination and were verified through the activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and carbonic anhydrase (CA), while the responses of biotransformation (activity of etoxiresorufin-O-deetilase (EROD) and glutathione S-transferase (GST)), the neurotoxicity (activity of acetylcholinesterase (AChE)) and liver histopathology were checked against the contamination by malathion. Also, for both bioassays, the damage to macromolecules (lipids (LPO) and DNA (comet assay in blood (ECA) and liver (HCA), micronucleus test (MNA) and polichromatic erythrocytes (PCE)) were evaluated. Overall, the study showed that, despite the non-induction or the unclear action of some of the enzyme systems, all three species of Astyanax were susceptible to the contamination of both xenobiotics, since damages to their macromolecules were observed. From these results, a cluster analysis was performed and was efficient in integrating all the data of the three species to display the ecotoxicological relationship between them, which was consistent and representative. With this paper, we hope to contribute in promoting the investigation of the interspecifc relationship of ecotoxicological responses of Astyanax by creating a database on the genus and by encouraging further studies with this and other genera.

The Comet Assay is a rapid and sensitive method for detection of breaks in the DNA strand into individual cells, commonly used at ecotoxicology. At this time, the method has been refined, but not yet fully standardized and variations on protocol are common. When environmental reviews are performed, the large number of samples required and the conditions of handling during transportation of samples to the laboratory are frequent problems. Until now, however, no truly effective method for samples preservation for this test was described. Therefore, we tested three stock solutions: fetal bovine serum (FBS), phosphate buffered saline (PBS) e Tris. Gill cells of Rhamdia quelen contaminated with insecticide Fipronil were maintained in these solutions, for 0, 24 and 48 hours to perform Comet Assay. It was found that fetal bovine serum was the solution that best preserved the DNA integrity of gill cells, followed by Tris and finally by PBS. We concluded in this study that fetal bovine serum is the solution that best conserve the gill cells and genetic material of R. quelen for a time until 48 hours in the absence of light and at 4oC.

The bioaccumulation of saxitoxins (STX) in the trophic chain, mainly in freshwater, are not completely known. This work aimed to elucidate the effects of STX on Hoplias malabaricus through trophic bioassay. The fish were fed once every five days with Astyanax sp. before being subjected to an intraperitoneal inoculation with the lysate of Cylindrospermopsis raciborskii culture containing 97% STX and 3% by neosaxitoxin and gonyautoxin during 20 days. The animal's liver was assessed using biomarkers as activities of superoxide dismutase (SOD), catalase (CAT), glutathione S-transferase (GST), and glutathione peroxidase (GPx), and concentrations of reduced glutathione (GSH) and lipoperoxidation (LPO) and protein carbonylation (PCO). In the blood was analyzed the genotoxic and hematological parameters. The hepatosomatic index and the relative condition factor did not show a significant difference between the exposed and control groups. The values of mean corpuscular hemoglobin concentration and mean corpuscular hemoglobin increased in the STX group. The hepatic tissue from both groups exhibited a typical pattern that have been already described for most teleost fish. The results suggested the generation of reactive oxygen species, with increased activity of GPx and concentrations of LPO and GSH; whereas the specific activity of SOD decreased. However, no changes were observed in the CAT, PCO, and DNA damage. Although the STX effects are known as neurotoxic, this cyanotoxin caused liver biochemical alterations that can be considered ecologically relevant.

The survey of the effects of toxic metals and its organic compounds are of critical importance because these compounds tend to accumulate in aquatic environments. In the present work, the genotoxic potential of methylmercury, an organomercurial compound with high toxicity and present in large amounts in fish of the Amazon due to the mining process, was evaluated using the piscine micronucleus test and comet assay in fish. Specimens of Hoplias malabaricus (popularly known as traira), a neotropical specie, was exposed to methylmercury via food web, over 70 days, in two different concentrations: two groups of fifteen fish were tested with 0.075 μg CH3Hg+/g and 0.75 μg CH3Hg+/g of methylmercury per gram of fish, at 5-day intervals and over 14 successive intervals whereas control group, composed by nine fish, was fed by uncontaminated prey fish (Astyanax sp). One of the aims of this study was to reproduce the contamination found in nature in an attempt to increase our biological knowledge on the neotropical species. Hoplias malabaricus specimens were then anesthetized for removal of blood samples and dissected, for cephalic kidney removal. As a result, it was observed that the piscine micronucleus test showed no significant differences between exposed groups and control group. The comet assay performed on erythrocyte system cells, showed a significant difference between controls and contaminated, but there was no difference between doses. In contrast, the kidney cells comet assay showed no significant difference between groups, probably due to the type of damage caused by xenobiotics to be related to the tissues of most contact with it, as well as its mode of action and the chain of bioaccumulation within bodies.

The use of the non-steroidal anti-inflammatory drugs such as dipyrone is so widespread that this drug and its metabolites have been detected in effluents and surface water. This study aimed to evaluate the potential toxic effects of dipyrone on the aquatic environment, using a native fish species, Rhamdia quelen. Fish were exposed to three concentrations of dipyrone, 0.5, 5 and 50 μg/L, in the water for 15 days, and hematological, biochemical, genetic and morphological biomarkers were evaluated. The glutathione S-transferase activity decreased in the highest concentration in relation to the control group. In addition, hematocrit, red blood cells and thrombocyte counts were decreased in all three exposed groups in relation to the control group. The comet assay showed DNA damage at the lowest concentration of dipyrone and significant kidney damage. Those results suggest that a constant exposure of aquatic organisms to dipyrone presents potential toxic effects.

Diverse genetic biomarkers have been used to evaluate the effects of pollution by mutagenic agents such as metals and pesticides, as well as a large variety of chemical substances derived from human activities. This work researched the effects that an exposure of 60 days to the insecticide Fipronil (concentrations of 0.05, 0.10 and 0.23 μg/L) can cause in the fish Rhamdia quelen using Comet assay with gills, histopathological analysis of gills and the Piscine Micronucleus test and Nuclear Morphological Alterations. The results for the Comet assay and for gills histopathological injuries showed no difference between the control group and the contaminated groups. In the Piscine Micronucleus test, the smallest concentration of Fipronil (0.05 μg/L) was similar as the control group, while concentrations of 0.10 and 0.23 μg/L caused more damage to the DNA. These results suggested that only the highest concentrations of Fipronil tested cause damage in erythrocytes, but none of these concentrations was sufficient to alter the DNA in the gill cells. R. quelen may be a less sensitive bioindicator than other fish that have been tested. On the other hand, the concentrations used may not have been sufficient to detect alterations in the DNA of R. quelen with the chosen tests. Works like this take on great importance given the enormous quantity of substances that are thrown daily into the environment in an uncontrolled way, without evaluation of the consequences. The application of these tests with other concentrations, tissues and exposure times is suggested for future works.

The Alagados Reservoir (Brazil) is an important source for the supply of water, recreation and fishery. Since 2002, the occurrence of cyanobacterial blooms (paralytic shellfish toxins - PST producers) have been noted. This study was aimed at the monitoring of PST occurrence in the Reservoir's water and fish. Biomarkers such as ethoxyresorufin-O-deethylase (EROD), glutathione S-transferase (GST), catalase (CAT), and acetylcholinesterase (AchE) activities, lipoperoxidation (LPO), histopathology, and comet assay were analyzed in fish. Water and fish were sampled in spring, summer and autumn. The PST concentrations in water were 5.15, 43.84, and 50.78 ng equiv Saxitoxin/L in the spring, summer and autumn, respectively. The PST muscle concentration was below the limit for shellfish. Gonyautoxins (GTX) were found in water samples and fish muscle, and GTX 5 was the major analogous found in muscle. In the summer samples, the LPO, genetic damage, and the GST and AchE activities increased while in the autumn an increase in EROD activity and genetic damage were observed. In all samplings, histopathological alterations in the fish gills and liver were found. The results showed a seasonal variation in the fishes health, which could be related also to farming activities and to the contaminants bioavailability during the year.

Cylindrospermopsin is a potent toxicant for eukaryotic cells produced by several cyanobacteria. Recently, primary hepatocyte cultures of Neotropical fish have been established, demonstrating to be a quite efficient in vitro model for cellular toxicology studies. In the current study, a protocol for culture of Prochilodus lineatus hepatocytes was established and utilized to investigate the cellular responses to purified cylindrospermopsin exposure. Hepatocytes were successfully dissociated with dispase, resulting in a cell yield of 6.36 × 10 7 cells g −1 of liver, viability of 97% and attachment on uncoated culture flasks. For investigation of cylindrospermopsin effects, hepatocytes were dissociated, cultured during 96 h and exposed to three concentrations of the toxin (0.1, 1.0 or 10 μg l −1 ) for 72 h. Cylindrospermopsin exposure significantly decreased cell viability (0.1 and 1 μg l −1 ) and multixenobiotic resistance mechanism, MXR (all exposed groups), but increased reactive oxygen/nitrogen species levels (all exposed groups) and lipid peroxidation (10 μg l −1 ). On the other hand no significant alterations were observed for other biochemical biomarkers as 2GSH/GSSG ratio, protein carbonyl levels and DNA strand breaks or glutathione S -transferase and glucose 6-phosphate dehydrogenase activities. In conclusion, hepatocytes might be made sensitive to cylindrospermopsin, at least in part, due to reduction of xenobiotics and endobiotics efflux capacity by MXR. Additionally, the toxin exposure suggests important issues regarding hepatocytes survival at the lowest cylindrospermopsin concentrations.