Epilithic biofilms, POCIS, and water samples as complementary sources of information for a more comprehensive view of aquatic contamination by pesticides and pharmaceuticals in southern Brazil.

Spatial-temporal monitoring of the presence of pesticides and pharmaceuticals in water requires rigor in the choice of matrix to be analyzed. The use of matrices, isolated or combined, may better represent the real state of contamination. In this sense, the present work contrasted the effectiveness of using epilithic biofilms with active water sampling and with a passive sampler-POCIS. A watershed representative of South American agriculture was monitored. Nine sites with different rural anthropic pressures (natural forest, intensive use of pesticides, and animal waste), and urban areas without sewage treatment, were monitored. Water and epilithic biofilms were collected during periods of intensive pesticide and animal waste application. After the harvest of the spring/summer crop, a period of low agrochemical input, the presence of pesticides and pharmaceuticals was monitored using the POCIS and epilithic biofilms. The spot water sampling leads to underestimation of the level of contamination of water resources as it does not allow discrimination of different anthropic pressures in rural areas. The use of endogenous epilithic biofilms as a matrix for the analysis of pesticides and pharmaceuticals is a viable and highly recommended alternative to diagnose the health of water sources, especially if associated with the use of POCIS.

A mixture of pesticides at environmental concentrations induces oxidative stress and cholinergic effects in the neotropical fish Rhamdia quelen.

The insecticides imidacloprid (IMI), a neonicotinoid, and propoxur (PRO), an N-methylcarbamate compound, are pesticides widely used throughout the world. Although they are not used together to combat pests, both are often found in freshwater near agricultural areas. Thereby, the goal of this study was to evaluate the additive effects of IMI and PRO mixtures at environmental concentrations in relation to isolated compounds on Rhamdia quelen, a neotropical fish. The fish was exposed to IMI (0.11 µg/L), PRO (0.039 µg/L), or Mix (0.11 µg/L IMI plus 0.039 µg/L PRO) during 96 h. Glutathione S-transferase (GST), superoxide dismutase (SOD), catalase (CAT), acetylcholinesterase (AChE) activities were determined. To verify oxidative damage thiobarbituric acid reactive substances (TBARS), protein carbonyl (PC), reactive oxygen species contents (ROS), antioxidant capacity against peroxides (ACAP) were determined in gills, liver, brain and muscle. The results shows that a mixture of these pesticides at environmental concentrations inhibited acetylcholinesterase activity in the brain and induced oxidative damage in all analyzed tissues. These results reinforce the hypothesis that mixture of contaminants present in environment could induce additive or synergistic effects on fish species.

Environmentally relevant pesticides induce biochemical changes in Nile tilapia (Oreochromis niloticus).

The high demand for food consequently increases the entry of agricultural residues into water resources, and this phenomenon can affect non-target organisms in different ways. Environmentally relevant pesticide effects (per se or in combinations) are scarce in the scientific literature. Therefore, the aim of this study was to investigate: (1) the presence of pesticide residues in an important Brazilian source of water supply and power generation (Jacuí river), during 1 year of monitoring. (2) in a laboratory study verify the effects of the most frequently, herbicide, fungicide, and insecticide found in Jacuí river (individualized or in a mixture) on biochemical parameters in different tissues of Oreochromis niloticus. Twenty pesticide residues were detected in superficial water samples, and two of them are banned in Brazilian territory. Atrazine (0.56 µg L-1), azoxystrobin (0.024 µg L-1), and imidacloprid (0.11 µg L-1) were the most frequently herbicide, fungicide, and insecticide, respectively, found in the river and were used in the laboratory assay. O. niloticus exposed to the pesticide mixture exhibited more biochemical effects than individualized exposure groups. This response can be a result of the combined pesticide effects, culminating in an additive or synergistic effect, depending on the biomarker. In individual exposure groups, atrazine presented the most pronounced alterations, followed by azoxystrobin and imidacloprid. Overall, pesticide exposure increased levels of oxidative stress parameters, reduced antioxidant enzyme activities, and induced acetylcholinesterase activity. These findings highlight the threat to aquatic organisms which may be exposed to a miscellaneous of toxic compounds in the environment.

Biochemical and Behavioral Responses in Zebrafish Exposed to Imidacloprid Oxidative Damage and Antioxidant Responses.

Imidacloprid (IMI) is an insecticide used worldwide, a neonicotinoid that could cause toxicity in non-target organisms. Zebrafish (Danio rerio) is a model organism widely used in different fields of research such as behavioral studies, biochemical parameters as well as neurotoxicity research. Here, we investigate whether the exposure to three concentrations (0.15, 15, and 45 µg/L) of IMI for 96 h alters responses in zebrafish. Oxidative stress parameters and acetylcholinesterase activity (AChE) as well as the behavioral responses of locomotion were measured. IMI exposure decreased distance traveled in fish exposed to the 45 µg/L. In the exploratory activity, time spent and transitions to the top area of the water column decreased in fish exposed to all concentrations of IMI. In addition, exposures to 45 and 15 µg/L of IMI decreased episodes of erratic movement in the zebrafish. Exposures to IMI at a concentration of 45 µg/L decreased the time spent in erratic movements and increased the time spent with no movement (i.e., "freezing"). Glutathione S-transferase (GST) activity was increased in the brain of zebrafish exposed for 96 h to concentrations of 0.15 and 45 µg/L. Brain AChE activity was reduced and the levels of carbonyl protein (CP) increased in brain of zebrafish at concentrations of 15 and 45 µg/L. Lipid peroxidation measured by TBARS and, also non-protein thiols (NPSH) did not show any variation in the brain of zebrafish exposed to IMI. Changes in the activity of cholinergic neurotransmitters in the brain tissues of zebrafish indicate IMI toxicity. Exposures of fish over 96 h to IMI at a nominal concentration of 45 µg/L caused more extensive sublethal responses in zebrafish, but this concentration is well above those expected in the aquatic environment. Studies are warranted to evaluate the effects on behavior and biomarker responses in fish exposed over longer periods to IMI at environmentally relevant concentrations.

Acute Silver Catfish (Rhamdia quelen) Exposure to Chlorantraniliprole Insecticide.

The aim of the current study is to investigate whether silver catfish (Rhamdia quelen) individuals exposed to commercial formulation of the chlorantraniliprole insecticide used in rice crops present changes in biochemical parameters. Fifty-four (54) silver catfish individuals were distributed in six units per tank (n = 6/repetition; triplicate/treatment) and subjected to the following treatments: T1-control, without insecticide; T2 (0.02 µg/L of insecticide) and T3 (0.20 µg/L of insecticide). Exposure time lasted 24 or 96 h, and it was followed by 96 h recovery in pesticide-free water. Results have indicated biochemical changes in cortisol, glucose, lactate and plasma protein levels, as well as few ionic changes in animals' gills during the exposure and recovery periods. Chlorantraniliprole incidence in water resulted in some biochemical changes in silver catfish specimens' plasma and gills throughout the acute exposure protocol (sub-lethal dose). Thus, chlorantraniliprole insecticide has caused osmoregulatory and/or biochemical imbalance in the investigated species under the herein adopted laboratory conditions; these changes did not get back to normal levels even after specimens were left to recover for 96 h in clean water.

Effects of chlorantraniliprole insecticide on innate immune response of silver catfish (Rhamdia quelen) naturally infected with Aeromonas hydrophila.

It is well documented today that pesticides, used in crop production, may modulate the immune system of healthy fish. However, there is still only limited information regarding the effects of these anthropogenic stressors in conjunction with natural stressors (pathogens), on the innate immune responses of freshwater fish. Thus, the aim of this investigation was to compare the innate immune response of two groups of fish (Rhamdia quelen), naturally infected with Aeromonas hydrophila, exposed and unexposed to a non-lethal concentration of chlorantraniliprole (CAP) insecticide (0.0 and 1.3 µg/L/24 h). Unhealthy fish exposed to CAP showed significant higher total leukocyte counts and neutrophils percentage compared to non-exposed infected fish). However, the monocytes and eosinophils percentage significantly decreased in fish exposed to CAP. Furthermore, lysozyme activity values measured in plasma, skin mucus, gill and intestine significantly reduced in fish exposed to CAP. The CAP-induced immunomodulation may interfere on the ability of the animal to heal or fight the infection, and possible contribute to the spread of bacterial infection in fish production or environment.

Ecological risk of pesticide contamination in a Brazilian river located near a rural area: A study of biomarkers using zebrafish embryos.

Aquatic environments are affected by the use of pesticides in agricultural areas near rivers. To assess the impact of pesticide residues on affected environments Danio rerio (zebrafish) embryos have become an alternative model for biomonitoring studies. In the present study, zebrafish embryos were used as bioindicator of water quality in the Vacacaí river, located in the city of Santa Maria, southern Brazil. We hypothesized that it would be possible to observe changes in the biomarkers tested in the embryos. Exposures were performed over a total of eight months during the year 2018 using water collected in a river located near agricultural areas. Twenty-four pesticides were found in river water samples. The most frequently found were atrazine, quinclorac and clomazone. During exposure (96 h) spontaneous movement, the heart rate and hatching rate were evaluated. After the exposure time the embryos were euthanized for biochemical assays. We analyzed biomarkers such as thiobarbituric acid reactive substance (TBARS), acetylcholinesterase (AChE), glutathione S-transferase (GST) and catalase (CAT). We observed increases in GST and TBARS, especially during periods of major water contamination such as January, February, October, and November. Pesticides can affect the development of native species that reproduce during periods of high agricultural production. These results demonstrate the potential use of biochemical parameters combined with developmental and behavioral analyses in zebrafish embryos for biomonitoring studies.

Influence of pesticides and abiotic conditions on biochemical biomarkers in Aegla aff. longirostri (crustacea, anomura): Implications for conservation.

Freshwater ecosystems are constantly threatened by the advance of agricultural activities. Abiotic variables (such as temperature, ammonia, and nitrite) and contaminants (e.g. pesticides) can potentially interact, increasing metabolism and the absorption of toxic substances, which can alter the ability of organisms to establish adequate stress responses. This study aimed to verify which pesticides were most frequently found and in the greatest quantities in low-order streams, and whether the combination of these pesticides with the abiotic variables altered the biological metabolism of aeglids. These freshwater crustaceans are important shredders that inhabit low-order streams and are sensitive to disturbances and/or abrupt environmental variations. The animals were exposed in situ in four streams (reference site and sites 1, 2, and 3). The reference site is a preserved stream with no apparent anthropogenic interference where aeglids still occur, while the other sites no longer exhibit populations of these animals and are influenced by agricultural activities. The exposure was performed bimonthly from November 2017 to September 2018 and lasted 96 h. Measured abiotic data and water samples were collected through all days of exposure. The analyzed biochemical parameters were acetylcholinesterase activity in muscle; and glutathione S-transferase, lipid peroxidation, protein carbonylation, non-protein thiols, antioxidant capacity against peroxides, and reactive oxygen species (ROS) in muscle, gills, and hepatopancreas. We found 24 active principles of pesticides, the most frequently being clomazone, atrazine, and propoxur. Bentazone was present at the highest amounts. The parameters evaluated in this study, including biochemical biomarkers and abiotic factors measured from the water, provided a separation of the months as a function of environmental conditions. There was a difference in activity and biomarker levels throughout the year within the same site and in some months between sites. The greater concentration or variety of pesticides associated with extreme abiotic (very high temperatures) data generated increased oxidative stress, with high levels of protein damage and considerable lipid damage in all tissues, as well as elevation in ROS, even with high levels of antioxidant capacity and non-protein thiols. With these data, we intend to warn about the risks of exposure to these environmental conditions by trying to contribute to the preservation of limnic fauna, especially aeglid crabs, because most species are under some degree of threat.

Protective effects of diet containing rutin against trichlorfon-induced muscle bioenergetics disruption and impairment on fatty acid profile of silver catfish Rhamdia quelen.

Trichlorfon is an organophosphate insecticide that is widely used on fish farms to control parasitic infections. It has been detected in freshwater ecosystems as well as in fishery products. There is a growing body of evidence to suggest that certain feed additives may reduce or prevent pesticide-induced toxicity in fish. The aim of the present study was to determine whether acute exposure to trichlorfon would alter bioenergetic homeostasis and alter fatty acid profiles in muscles of silver catfish (Rhamdia quelen). We also sought to determine whether rutin prevents or reduces these effects. Cytosolic and mitochondrial creatine kinase (CK) and activities of complexes II-III and IV in muscle were significantly inhibited by exposure to 11 mg/L trichlorfon for 48 h compared to effects in the unexposed group. Total content of polyunsaturated fatty acids (omega-3 and omega-6) were significantly lower in muscle of silver catfish exposed to 11 mg/L trichlorfon for 48 h than in the unexposed group. Addition of 3 mg rutin/kg feed increased CK activity and prevented inhibition of complex IV activity, as well as preventing all alterations of muscle fatty acid profiles elicited by exposure to trichlorfon. No significant differences were observed between groups with respect to muscle adenylate kinase or pyruvate kinase activities, as well as total content of saturated and monounsaturated fatty acids. Our findings suggest that exposure (48 h) to 11 mg trichlorfon/L water inhibits cytosolic and mitochondrial CK activity in muscle. Trichlorfon also affects activities of complexes II-III and IV in respiratory chain, with important consequences for adenosine triphosphate production. The pesticide alters fatty acid profiles in the fish and endangers human consumers of the product. The most important finding of the present study is that inclusion of rutin improves bioenergetic homeostasis and muscle fatty acid profiles, suggesting that it reduces trichlorfon-induced muscle damage.

Ecological impacts of pesticides on Astyanax jacuhiensis (Characiformes: Characidae) from the Uruguay river, Brazil.

Brazilian freshwater ecosystems are continuously exposed to pesticides and domestic sewage. The Uruguay River was chosen for this study because of its international importance, as it flows through Brazil, Argentina, and Uruguay. It receives contaminants such as pesticides and domestic residues. Thus, the aim of this study to assess the accumulation of pesticides in muscle of the fish Astyanax jacuhiensis, its biochemical responses, and the presence of pesticides in water. In total, seven pesticides were registered in water from both river sites. Eight pesticides were detected in fish muscle. The biochemical responses showed that brain lipid peroxidation (LPO) and protein carbonyl (PC) in A. jacuhiensis were higher in the summer. Muscle showed the highest LPO levels in the spring and the highest PC in the summer. Liver LPO and PC levels were higher in the spring and summer. In the gills, the PC was higher in the spring and the LPO in the spring and winter. In the brain and in the gills, glutathione-S-transferase activity was high in the summer and autumn. Catalase activity was lower during the winter and spring. Non-protein thiol (NPSH) levels were lower in the brain in the winter and spring. Muscle tissue showed lower NPSH in the winter (site 1). Liver NPSH showed increased levels in liver in the spring and winter (site 2). The biochemical results clearly is related to pesticides and/or to the presence of other contaminants in the water such as metals or domestic sewage. The accumulation of pesticides in fish muscle added evidence that pesticides have been used in the area surrounding the Uruguay River. In conclusion, the biomarkers assayed in the present study could be used in future investigations considering other sampling sites along Uruguay River.

Potential environmental toxicity of sewage effluent with pharmaceuticals.

Sewage effluent effects on the biochemical parameters of Astyanax bimaculatus organs were investigateted. Treated sewage was collected in a treatment plant; 43 compounds, among them, pharmaceuticals and hormones, were investigated. Caffeine, ciprofloxacin, clindamycin, ofloxacin, oxytetracycline, paracetamol, sulfadiazine, sulfamethoxazole, sulfathiazole and tylosin waste was detected in the collected material. Fish were divided into four groups: control, TSE (treated sewage effluent), TSE + P (TSE with increased concentration of five pharmaceuticals) and PTSE (TSE + P post-treated with O3/H2O2/UV). Biochemical parameters were evaluated in different organs after 14-day exposure. TBARS levels increased significantly in the brain of animals in the TSE and TSE + P groups in comparison to the control. There was significant reduction in TBARS levels recorded for the liver, muscle and gills of animals in the PTSE group in comparison to those of animals in the other groups. AChE activity reduced in the muscle of animals in the groups showing the highest pharmaceutical concentrations. CAT activity in the liver of animals in groups exposed to pharmaceutical effluent was inhibited. GST activity increased in brain of animals in the TSE + P and PTSE groups, whereas reduced levels of this activity were observed in liver of animals in the TSE group. Increased GST activity was observed in the brain of animals in TSE + P and PTSE groups. Based on integrated biomarker response values, the TSE + P group presented greater changes in the analyzed parameters. Results point out that pharmaceutical waste can cause oxidative stress, as well as affect biochemical and enzymatic parameters in Astyanax sp. Post-treatment can also reduce damages caused to fish, even in case of the likely formation of metabolites. Based on these results, these metabolites can be less toxic than the original compounds; however, they were not able to fully degrade the pharmaceutical waste found in the sewage, which can interfere in fish metabolism.

Phytostimulation of lowland soil contaminated with imidazolinone herbicides.

The phytostimulation is a phytoremediation technique that can be used to remediate area contaminated with herbicides. It is necessary to select plants with high capacity to stimulate soil microbial activity. The present work aimed at evaluating seven plant species regarding their ability to phytostimulate soil and enhance the degradation of the herbicides imazethapyr, imazapic and imazapyr in a lowland soil. An Alfisol Albaqualf was cultivated with the following species, Canavalia ensiformis, Glycine max, Oryza sativa cultivar PUITÁ INTA CL, Lolium multiflorum, Vicia sativa and consortium Lotus corniculatus + Trifolium repens. The rhizosphere of these plants and non-rhizospheric (uncultivated soil) as a control were contaminated in laboratory with analytical standart of the three herbicides, at rates of 0, 150, 300 and 750 g a.i. ha-1, in separate assays. Biodegradation was estimated by quantifying C-CO2 production and through analysis of herbicides residues in soil using liquid chromatography. Results show that biodegradation of herbicides imazethapyr, imazapic and imazapyr was higher in vegetated soil than in unvegetated soil. The leguminous species Canavalia ensiformis, Glycine max, Vicia sativa and consortium of Lotus corniculatus + Trifolium repens showed a great capacity to promote soil microbial, resulting in average biodegradation rates of 91, 92 and 93% for herbicides imazethapyr, imazapic and imazapyr in soil, respectively.

Toxicological response of silver catfish (Rhamdia quelen) after acute exposure to a commercial insecticide containing thiamethoxam.

This study assessed the hematological, enzymatic and osmoregulatory responses of silver catfish (Rhamdia quelen) exposed to sublethal concentrations (1.125 and 3.750 µg/L) of a commercial thiamethoxam-containing insecticide used on rice crops. Groups of 6 fish per tank (in triplicate, n = 3, total 54 fish) were exposed for up to 96 h to different concentrations of the compound. After this period, fish were placed in clean water for 48 h. Two fish from each tank (6 per treatment) that had been exposed to the insecticide for 24 h were anesthetized with eugenol and blood was collected to evaluate hematological and biochemical parameters. Blood, liver and muscle were collected for determination of metabolic parameters, plasma cortisol, Cl-, Na+ and K+ levels and H+-ATPase and Na+/K+-ATPase activity in the gill. H+-ATPase activity was higher in fish exposed to 1.125 µg/L insecticide at 24 h compared to control (0.0 µg/L). Differences in cortisol levels were evidenced throughout the experimental period. These results indicated that exposure to the insecticide changed the hematological, biochemical and metabolic profile of the animals, suggesting concern about environmental safety. Therefore, we discourage the use of this pesticide in areas that come into contact with water bodies inhabited by fish.

Assessment of River Water Quality in an Agricultural Region of Brazil Using Biomarkers in a Native Neotropical Fish, Astyanax spp. (Characidae).

Intensive agricultural and livestock activities demand high pesticide use and, consequently, contaminants reach aquatic ecosystems. In the lower Jacuí River, southern Brazil, there is a lack of knowledge about pesticide residues in water samples and the biochemical responses in native fish species. Thus, this study aimed to estimate the influence of pesticide residues and water parameters to biomarker responses in the native fish Astyanax spp. We performed seasonal biomonitoring in 2017 with water samples and fish collections. Biomarkers of oxidative stress, antioxidants, biotransformation, and neurotoxicity were analyzed in fish tissues. Fourteen pesticide residues were detected; they presented correlations with detoxification enzyme and oxidative stress biomarkers. These data indicate that most of variations can be related to the pesticide presence in water indicating high aquatic pollution in this place.

Purinergic signaling as potential target of thiamethoxam-induced neurotoxicity using silver catfish (Rhamdia quelen) as experimental model.

Thiamethoxam is a broad-spectrum pesticide widely used in agricultural practice throughout the world. Worryingly, this pesticide is considered a potential contaminant on the surface and underground water, being a significant risk to aquatic ecosystems and humans. In this sense, we decided to evaluate the activity of enzymes belonging to purinergic system, which is linked with regulation of extracellular nucleotides and nucleosides, as adenosine triphosphate (ATP) and adenosine (Ado) molecules involved in the regulation of immune and inflammatory responses. Such as the neurotoxic effects of thiamethoxam remain poorly understood, the aim of this study was to evaluate whether purinergic signaling may be considered a potential target of thiamethoxam-induced neurotoxicity in silver catfish (Rhamdia quelen). Brain ectonucleoside triphosphate diphosphohydrolase (ATP as substrate) and 5'-nucleotidases activities were inhibited at 3.75 µg L-1 after 24 h of exposure and at 1.125 and 3.75 µg L-1 after 96 h of exposure compared with the control group. On the other hand, brain adenosine deaminase activity was stimulated at 3.75 µg L-1 after 24 h of exposure and at 1.125 and 3.75 µg L-1 after 96 h of exposure compared with the control group. Brain ATP levels increased at 3.75 µg L-1 after 24 h of exposure and at 1.125 and 3.75 µg L-1 after 96 h of exposure compared with the control group, while the Ado levels decreased. The enzymatic activity of the purinergic signaling did not return to control levels after a 48-h recovery period, revealing the potential neurotoxic effects of thiamethoxam. In summary, the brain purinergic signaling may be considered a potential target for thiamethoxam-induced neurotoxicity in silver catfish.

Integrated biomarkers response confirm the antioxidant role of diphenyl diselenide against atrazine.

Atrazine (ATZ) is a herbicide worldwide used. That can cause oxidative damage in non-target organisms, such as fish. Furthermore, the threat of exposure to pesticides together with poor nutrition is hazardous to the normal development of fish, and supplementation of the fish diet with antioxidants compounds is an alternative approach to prevent the hazardous effects of pesticide exposure. Here we aimed to investigate the capacity of diphenyl diselenide (PhSe)2 diet supplementation to improve the antioxidant defense of Cyprinus carpio (carp) exposed to environmental concentrations of ATZ. To prove the efficiency of (PhSe)2, we used the Integrated Biomarkers Response (IBR) methodology. Therefore, carp were fed for 8 weeks diets either with or without (PhSe)2 and exposed to 2 or 10µg/L of ATZ for 96h, euthanized, and their liver, gills, and muscle tissues were removed for biochemical assays. ATZ was able to cause oxidative damage from reactive species production in all tissues of carp, as observed by the increase of lipid peroxidation and protein damage. The activity of some antioxidant enzymes was inhibited in carp exposed to ATZ. However, (PhSe)2 supplementation was able to prevent this ATZ-induced damage by improving the activities of antioxidant enzymes and through antioxidant competence of (PhSe)2per se. Furthermore, IBR was shown to be a useful tool to compare treatments, even at different concentrations, and identify the efficiently antioxidant behavior of the organoselenium compound.

Biochemical Responses in Freshwater Fish Exposed to Insecticide Propoxur.

Although designed to control pests selectively, there is some evidence that environmental contamination by pesticides increases risks for humans and wildlife. In the present study, we evaluated biomarkers of oxidative stress in Astyanax jacuhiensis exposed to (5, 15 and 30 µg L-1) of carbamate Propoxur (PPX) for 96 h. Glutathione S-transferase (GST) in liver and gills showed reduced activity in all PPX concentrations tested. Acetylcholinesterase (AChE) activities reduced in brain and muscle at concentrations 15 and 30 µg L-1 of PPX. Lipid peroxidation (LPO) and hydrogen peroxide (HP) had no significant differences. In the brain, protein carbonyl (PC) increased in all groups treated with PPX. Although PPX is a selective pesticide, it causes oxidative damage and enzyme alteration in fish. This study pointed out some biomarkers that could be used to assess effects of environmentally relevant concentrations of pesticides, and infer about studies using fish as bioindicator.

Richness and density of aquatic benthic macroinvertebrates after exposure to fungicides and insecticides in rice paddy fields.

The objective of this study was to verify the richness and density of aquatic benthic macroinvertebrates after exposure to fungicides and insecticides of the rice paddy fields. In the crop seasons of 2012/13 and 2013/14, field experiments were performed, which consisted of single-dose applications of the fungicides trifloxystrobin + tebuconazole and tricyclazole, and the insecticides lambda-cyhalothrin + thiamethoxam and diflubenzuron, in 10 m2 experimental plots, over rice plants in the R3 stage. Control plots with and without rice plants were maintained in order to simulate a natural environment. Soil samples were collected during rice cultivation for assessment of the macroinvertebrate fauna. Chemical-physical parameters assessed in the experiments included temperature, pH and oxygen dissolved in the water and pesticide persistence in the water and in the soil. The application of a single dose of the pesticides and fungicides in the recommended period does not cause significant negative effects over the richness and density of the macroinvertebrates. Tebuconazole, tricyclazole and thiamethoxam showed high persistence in the irrigation water of rice paddy fields. Thus, the doses and number of applications of these products in crops should be carefully handled in order to avoid contamination of the environment.

Evaluation of alternative sorbents for dispersive solid-phase extraction clean-up in the QuEChERS method for the determination of pesticide residues in rice by liquid chromatography with tandem mass spectrometry.

Many compounds are used for pest control during the production and storage of rice, making it necessary to employ multiclass methods for pesticide residues determination. For this purpose, QuEChERS-based methods are very efficient, fast and accurate, and improvements in the clean-up step are important, especially for complex matrices, like cereals. In this work, different sorbents such as chitosan, florisil(®) , alumina, diatomaceous earth, graphitized carbon black, besides the commonly used primary secondary amine and octadecylsilane, were evaluated for dispersive solid-phase extraction clean-up in acetate-buffered QuEChERS method for the determination of residues of 20 representative pesticides and one metabolite in rice by liquid chromatography coupled to tandem mass spectrometry. The sorbent C18 presented the best results, however, chitosan showed similar results, and the best performance among the unconventional sorbents evaluated. The method limit of quantification, attending accuracy (70-120% recovery) and precision (RSD ≤20%) criteria, ranged from 5 to 20 µg/kg. Results showed that chitosan is an effective alternative to reduce analysis costs, maintaining the method reliability and accuracy.

Imazapyr+imazapic herbicide determines acute toxicity in silver catfish Rhamdia quelen.

Imazapyr (IMY) and imazapic (IMI) are imidazolinone herbicides which have been associated in a commercial formulation (Kifix(®)). To date, there are no studies on the toxicity of an IMY+IMI herbicide in fish. This work aimed to assess the acute toxicity (24 and 96 h) of IMY+IMI (0, 0.488 and 4.88 µg/L) towards Rhamdia quelen through hematological, biochemical, immunological, ionoregulatory and enzymatic indexes. Red blood cell count was lower at 4.88 than at 0.488 µg/L (24 and 96 h); mean corpuscular volume was lower than control at both concentrations (24 h) and at 0.488 µg/L (96 h); lymphocytes declined at 4.88 µg/L comparing to control (96 h); and monocytes increased at 4.88 µg/L (96 h) in comparison with the respective control and with 4.88 µg/L at 24h. Aspartate aminotransferase was higher at 0.488 µg/L (96 h) than the respective control and the respective concentration at 24 h; uric acid reduced at 4.88 µg/L comparing with 0.488 µg/L (96 h); and cortisol was lower at 4.88 µg/L compared to 0.488 µg/L and control (96 h). Herbicide exposure lowered plasma bactericidal activity at both concentrations (24 h) and at 0.488 µg/L (96 h); and plasma complement activity declined at 4.88 µg/L comparing with 0.488 µg/L and control (96 h), and was lower at all concentrations at 96 h than at 24 h. Plasma K(+) levels were higher at 4.88µg/L than in the remaining groups (24 and 96h); and Na(+) levels decreased at 4.88 µg/L compared to control (96 h). Na(+)/K(+)-ATPase and H(+)-ATPase activities in gills were lower at 4.88 µg/L comparing with control (24 h) and with the respective concentration at 96 h; and AChE activity in brain was higher at 0.488 and 4.88 µg/L than control (24 h) and the respective concentrations at 96 h, while in muscle it was higher at 0.488 and 4.88 µg/L than control (96 h) and the respective concentrations at 24 h. The present findings demonstrate that, despite IMY+IMI targets the animal-absent AHAS enzyme, such formulation displayed an acute toxic effect upon R. quelen homeostasis by impacting on vital functions such as immune defense, metabolism, ionoregulation and neurotransmission.