Effect of Saccharomyces cerevisiae UFMG A-905 in a murine model of food allergy.

Food allergy is triggered when there is an abnormal activation of the immune system by food allergens. Currently, there is no curative therapy for this pathological condition. Due to the immunomodulatory properties of probiotics they are potential candidates as therapeutic tools for food allergy. Therefore, the aim of this study was to evaluate the probiotic effect of Saccharomyces cerevisiae UFMG A-905 (905) in an in vivo model of food allergy. Probiotic effect was assessed by clinical, histological, immunological and microbiological parameters analysis. Furthermore, we also evaluated if 905 after inactivation has an effect, as well as if such an effect is dose dependent. Our results showed that oral administration of only viable 905 promotes a significant attenuation of tissue injury and myeloperoxidase (MPO) activity levels. Moreover, the treatment reduced interleukin 17 levels, and administration of the supernatant from the yeast culture also promoted a significant decrease in MPO levels. However, considering the systemic parameters, immunoglobulin (Ig)E and IgG anti-ovalbumin, which are essentials for triggering the allergic process, there was no effect, suggesting that the yeast promotes a local but not a systemic effect in the model evaluated. In addition, we found that only high doses of viable 905 were able to attenuate the signs of inflammation. In conclusion, oral administration of 905 led to a local effect that depends on the viability of the yeast.

Acute and long-term effects of trophic exposure to silver nanospheres in the central nervous system of a neotropical fish Hoplias intermedius.

Nanotechnologies are at the center of societal interest, due to their broad spectrum of application in different industrial products. The current concern about nanomaterials (NMs) is the potential risks they carry for human health and the environment. Considering that NMs can reach bodies of water, there is a need for studying the toxic effects of NMs on aquatic organisms. Among the NMs' toxic effects on fish, the interactions between NMs and the nervous system are yet to be understood. For this reason, our goal was to assess the neurotoxicity of polyvinylpyrrolidone coated silver nanospheres [AgNS (PVP coated)] and compare their effects in relation to silver ions (Ag+) in carnivorous Hoplias intermedius fish after acute and subchronic trophic exposure through the analysis of morphological (retina), biochemical (brain) and genetic biomarkers (brain and blood). For morphological biomarkers, damage by AgNS (PVP coated) in retina was found, including morphological changes in rods, cones, hemorrhage and epithelium rupture, and also deposition of AgNS (PVP coated) in retina and sclera. In the brain biomarkers, AgNS (PVP coated) did not disturb acetylcholinesterase activity. However, lowered migration of the DNA tail in the Comet Assay of blood and brain cells was observed for all doses of AgNS (PVP coated), for both acute and subchronic bioassays, and in a dose-dependent manner in acute exposure. Ag+ also reduced the level of DNA damage only under subchronic conditions in the brain cells. In general, the results demonstrated that AgNS (PVP coated) do not cause similar effects in relation to Ag+. Moreover, the lowered level of DNA damage detected by Comet Assay suggests that AgNS (PVP coated) directly interacts with DNA of brain and blood cells, inducing DNA-DNA or DNA-protein crosslinks. Therefore, the AgNS (PVP coated) accumulating, particularly in the retina, can lead to a competitive disadvantage for fish, compromising their survival.

Effects of ecologically relevant concentrations of cadmium in a freshwater fish.

Sub-chronic effects of ecologically relevant concentrations of cadmium (Cd) were evaluated in the catfish Rhamdia quelen. The fish were exposed to Cd (0, 0.1, 1, 10 and 100µgL(-1)) for 15 days. Bioconcentration was observed in the liver of fish exposed to 10 and 100µgL(-1) of cadmium. The liver glutathione S-transferase activity decreased at 0.1 and 1µgL(-1) and increased at 100µgL(-1) and lipoperoxidation increased in all tested concentrations. Fish exposed to 0.1, 1 and 100µgL(-1) Cd presented increase in hepatic lesion index. In the kidney, the catalase activity and LPO reduced in all exposed groups. The gluthatione peroxidase, etoxiresorufin-O-deethylase activities and metallothionein increased at the highest concentration of Cd, but the level of reduced glutathione decreased. The genotoxicity was observed at 0.1 and 100µgL(-1). Neurotoxicity was not observed. The results showed that low concentrations (range of µgL(-1)) of Cd caused hepato-, nephro- and hematological alterations in this freshwater fish species.

Multibiomarker in fish to evaluate a river used to water public supply.

We aimed to evaluate the ecological integrity of a large river, which receives agricultural and urban effluents and is used to water public supply. The fish species Astyanax bifasciatus was used as bioindicator during winter and spring 2012, and summer 2013 at the Middle Iguaçu River basin in Paraná state, Brazil. Water chemical and physical measures and ecotoxicological tests were carried out as well biochemical and genetic biomarkers in sampled fish in each period. The studied area was divided in three sample points: SP1, located where the water is collected to public supply; SP2, located in an urbanized area, and SP3, located at an urbanized area with the discharge of the sewage treatment. Although water chemical and physical analyzes were range of the Brazilian law to hydric bodies, anticholinesterasic effects were found in winter, oxidative stress in summer and spring. The higher genotoxic effect was in winter to all sample points. The temporal variation in biomarkers and the detection of caffeine in the water call attention to the water quality in this river mainly to be used to public supply.

Mesotrione herbicide promotes biochemical changes and DNA damage in two fish species.

Mesotrione is one of the new herbicides that have emerged as an alternative after the ban of atrazine in the European Union. To our knowledge, any work using genetic or biochemical biomarkers was performed in any kind of fish evaluating the toxicity of this compound. The impact of acute (96 h) exposure to environmentally relevant mesotrione concentrations (1.8, 7, 30, 115 e 460 µg L-1) were evaluated on the liver of Oreochorimis niloticus and Geophagus brasiliensis by assessing the activity of superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione-S- transferase (GST), the levels of reduced glutathione (GSH), carbonyl assays (PCO) and lipid peroxide (LPO) as well as the DNA damage to erithrocytes, liver and gills through the comet assay. We observed an increase in the concentration of GSH and the GPx activity in O. niloticus, and the GST and SOD activity in G. brasiliensis. We found significant increase in DNA damage in all tissues in both species. The results indicated that the acute exposure to mesotrione can induce oxidative stress and DNA damage in both species.

Biochemical changes in the liver and gill of Cathorops spixii collected seasonally in two Brazilian estuaries under varying influences of anthropogenic activities.

In order to understand environmental health by the use of a bioindicator species in estuaries, biochemical responses observed in the catfish Cathorops spixii such as catalase (CAT), glutathione-S-transferase (GST), lipid peroxidation (LPO) and acetylcholinesterase (AChE) were evaluated in liver and muscle. Furthermore, histological changes were also verified in liver and gills preparations. Fish were collected in three sites of the Santos-São Vicente estuary located at São Paulo (Brazil), subjected to varying levels of inputs of pollutants. For a reference site, specimens were sampled at Cananéia estuary at southern coast of São Paulo, a region with low anthropogenic influence. In general, no significant seasonal differences in antioxidant enzymes and lipid peroxidation responses were found in the organisms from the Cananéia estuary. However, in the polluted estuary (Santos-São Vicente), biochemical responses were observed by increases in GST hydroperoxides and decreases in AChE activities in the summer. Inhibition of AChE expression in fish from different areas of the Santos-São Vicente estuary in the summer was also found and can indicate neurotoxic effects in these organisms. Histopathological observation of gill and liver showed severe lesions, such as lamellar fusion and necrosis.

Effects of the herbicide atrazine in neotropical catfish (Rhamdia quelen).

The exposure to a world-wide used herbicide atrazine (ATZ) (96h exposure to 2, 10, and 100µgL(-1)), was investigated on the freshwater fish Rhamdia quelen through a multi biomarker approach. Liver histopathology revealed leukocyte infiltration, hepatocyte vacuolization like steatosis and necrosis areas, leading to raised lesion index levels in all tested concentrations. The increase of free melanomacrophage numbers was observed. Gill filaments revealed considerable loss of the microridges on pavement cells at 10 and 100µgL(-1) of ATZ, and a significantly increased of chloride cell (CC) number and density on apical surface area at 100µgL(-1) of ATZ. CAT, GST, GPx, and GR activities were inhibited by all tested concentrations. GSH levels were reduced in individuals exposed to 100µgL(-1). Osmoregulatory function was also disturbed. We observed an increase of plasma magnesium concentrations at 10µgL(-1). Additionally the inhibition of branchial carbonic anhydrase activity was observed at 100µgL(-1). In the kidney, carbonic anhydrase activity decreased only in the group exposed to 2µgL(-1). These results suggest that ATZ, represents a potential ecotoxicological hazard and can be hepatotoxic and nephrotoxic even low concentrations. The current study was the first to show the nephrotoxic effect of ATZ in fish. Besides, in Brazil, the environmental protection agency (CONAMA) establishes that the maximum allowed level of dissolved ATZ in water is 2µgL(-1), but the present results showed that this concentration may cause histopathological, biochemical and physiological changes in R. quelen.

Risks of waterborne copper exposure to a cultivated freshwater Neotropical catfish (Rhamdia quelen).

As it is the case in all animal food production systems, it is often necessary to treat farmed fish for diseases and parasites. Quite frequently, fish farmers still rely on the aggressive use of copper to control bacterial infections and infestations by ecto-parasites, and to manage the spread of diseases. The susceptibility of the neotropical fish Rhamdia quelen to copper was here evaluated at different waterborne copper concentrations (2, 7 or 11 µg Cu L(-1)) for 96 h, through a multi biomarkers approach. Liver histopathological findings revealed leukocyte infiltration, hepatocyte vacuolization and areas of necrosis, causing raised levels of lesions upon exposure to 7 and 11 µg Cu L(-1). Decreased occurrence of free melano-macrophages and increased densities of melano-macrophage centers were noted upon exposure to 11 µg Cu L(-1). Gills showed damages on their secondary lamellae already at 2 µg Cu L(-1); hypertrophy and loss of the microridges of pavement cells at 7 and 11 µg L(-1), and increased in chloride cell (CC) apical surface area (4.9-fold) and in CC density (1.5-fold) at 11 µg Cu L(-1). In the liver, catalase (CAT), glutathione peroxidase activities (GPx) and glutathione concentration (GSH) remained unchanged, compared to the control group. However, there was inhibition of 7-ethoxyresorufin-O-deethylase (EROD) at all copper concentrations tested. Glutathione reductase activity (GR) was reduced and levels of lipid peroxidation (LPO) were increased at 11 µg Cu L(-1). Glutathione S-transferase activity (GST) at 7 µg Cu L(-1) and superoxide dismutase activity (SOD) at both 7 and 11 µg Cu L(-1) were reduced. However, copper exposure did not alter brain and muscle acetylcholinesterase (AChE) activity. Osmoregulatory function was also disturbed, in agreement with the above-mentioned changes noted in the gills, as detected by plasma osmolality reduction in the group exposed to 11 µg Cu L(-1), and plasma chloride reduction at 2 µg Cu L(-1). These concentrations also, coherently, lead to inhibition of branchial carbonic anhydrase activity. In the kidney, increased carbonic anhydrase activity was measured in the groups exposed to 2 and 7 µg Cu L(-1). When these effects are compared to data available in the literature for other freshwater fish, also for 96 h of exposure, R. quelen appears as a relatively sensitive species. In addition, the concentrations employed here were quite low in comparison to levels used for disease control in real culture practices (ranging from 4 µg Cu L(-1) used against bacteria to 6000 µg Cu L(-1) against fungal infections). We can conclude that the concentrations frequently employed in aquaculture are in fact not safe enough for this species. Such data are essential for the questioning and establishment of new policies to the sector.

Bioaccumulation and related effects of PCBs and organochlorinated pesticides in freshwater fish Hypostomus commersoni.

Few studies have investigated the bioaccumulation of persistent organic pollutants (POPs) in Brazilian native freshwater fish. In order to evaluate the bioavailability, potential risk to human exposure and the effects of POPs in the fish Hypostomus commersoni, muscle and liver samples of thirteen specimens were collected in a lake located in the city of Ponta Grossa (Parana State, Southern Brazil). Also, the liver and gills were considered for histopathological studies, and oxidative stress was investigated in the liver. Expressive concentrations of POPs were observed in the liver and muscle, with a total of 427 ± 78.7 and 69.2 ± 18.1 ng g(-1) dry weights of polychlorinated biphenyls (PCBs), respectively. Negative correlations between the concentration of several POPs and glutathione S-transferase and glucose-6-phosphate dehydrogenase were found. Otherwise, the cholinesterase activity in the muscle and brain presented positive correlations with the concentration of POPs. The hepatic bioaccumulation of some banned pesticides like aldrin, dieldrin and DDT was associated with various histopathological findings in the liver and gills. Necrotic areas, fibrosis, leukocyte infiltration, and the absence of macrophage centers were observed in the liver, indicating both chronic exposure and immunological suppression. Neoplasic changes were observed in the gills, confirming the carcinogenic potential reported for some of the investigated pollutants. The current work was the first to study the bioaccumulation of POPs in H. commersoni, an important species in ecological aspects and as a vehicle to human exposure to PCBs and organochlorine pesticides (OCPs).

Subchronic effects of dipyrone on the fish species Rhamdia quelen.

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.

An assessment of acute biomarker responses in the demersal catfish Cathorops spixii after the Vicuña oil spill in a harbour estuarine area in Southern Brazil.

The Vicuña oil tanker exploded in Paranaguá Bay (South of Brazil), during methanol unloading operations in front of Paranaguá Harbour, on November 15th, 2004, releasing a large amount of bunker oil and methanol. Two weeks after the accident, the acute effects of the Vicuña Oil Spill (VOS) were evaluated in the demersal catfish Cathorops spixii, comparing a contaminated (at the spill site) and a reference site inside the Bay. Data were compared to previous measurements, taken before the accident, in the same species, from the same sites. The physiological biomarkers were the ones that best reflected acute effects of the spill: plasma osmolality, chloride, calcium, magnesium, and potassium. Morphological (liver and gill histopathology) and genetic (piscine micronucleus and DNA strand breaks) biomarkers revealed that damage was already present in fishes from both reference and contaminated sites inside the Bay. Thus, the reference site is not devoid of contamination, as water circulation tends to spread the contaminants released into other areas of the Bay. Acute field surveys of oil spill effects in harbour areas with a long history of contamination should thus be viewed with caution, and whenever possible previous evaluations should be considered for proper appraisal of biomarker sensitivity, especially in mobile bioindicators such as fish.

Evaluation of waterborne exposure to oil spill 5 years after an accident in Southern Brazil.

Fish (Astyanax sp.) were exposed to water samples from the Arroio Saldanha, locality of an oil spill, and the Barigui River (upstream and downstream of the confluence of the river and Arroio Saldanha), Brazil. Histopathological and biochemical biomarkers, as acetylcholinesterase (AchE), ethoxyresorufin-O-deethylase (EROD), catalase (CAT), glutathione-S-transferase (GST) activities, lipid peroxidation (LPO) levels, and amounts of polynuclear aromatic hydrocarbon residues (PAHs) in bile were evaluated. PAHs, metals, and surfactants were investigated in the water samples. CAT and GST activities increased in the Arroio Saldanha sample, whereas CAT activity and LPO levels were increased in the upstream and downstream sites. The results suggested a toxic action of the free radicals and disturbance of the antioxidant defense mechanisms. However, the EROD and AchE activities were not affected. Histopathology studies showed severe lesions. The oil is still bioavailable to biota, moreover, other pollution sources continue to affect the water in the Barigui River.

Toxic effects of DDT and methyl mercury on the hepatocytes from Hoplias malabaricus.

Here, we examined the impact of dichlorodiphenyltrichloroethane (DDT) and monomethyl mercury (MeHg) on the redox milieu and survival of hepatocytes from Hoplias malabaricus (traíra). After isolation and attachment of cells, we established one control and four treatments: DDT (50nM of DDT), MeHg I (0.25microM of MeHg), MeHg II (2.5microM of MeHg) and DDT * MeHg I (combination of 50nM of DDT and 0.25microM of MeHg). After four days the exposed hepatocytes presented significantly increased damage in lipids (all treatments), proteins (DDT * MeHg I and MeHg II) and reduced cell viability (all treatments). Also the antioxidant enzymes catalase, glucose-6-phosphate dehydrogenase (G6PDH), glutathione reductase and superoxide dismutase were affected. The current data showed that despite of some protective responses, the increased disturbs on membrane lipids and proteins, increased hydrogen peroxide levels, and decreased glutathione concentration and cell viability strongly indicate oxidative stress as the reason of hepatotoxicity due to DDT and MeHg exposure. In addition, DDT and MeHg together had greater effect than alone when G6PDH and glutathione-S-transferase activities and lipids damage were considered. These findings are indicative of hepatotoxicity occurring at realistic concentrations of DDT and MeHg found in Amazonian fish tissues.

Multibiomarker assessment of three Brazilian estuaries using oysters as bioindicators.

Oysters have been largely employed as bioindicators of environmental quality in biomonitoring studies. Crassostrea rhizophorae was selected to evaluate the health status of three estuarine areas impacted by anthropogenic activities along the Brazilian coast, in three estuarine complexes, ranging in latitude from 7 to 25 degrees S. In each estuary three sites were sampled in Winter and in Summer: a site considered as reference, and two sites next to contamination sources. Condition index was similar at all sites and estuaries, with the highest values found for Itamaracá oysters in Summer. Necrosis, hyperplasia, mucocyte hypertrophy and fusion of ordinary filaments were the main histopathological lesions observed. Muscle cholinesterase activity was overall similar, but with a strong seasonal effect. Inhibition or activation of branchial total ATPase and Na,K-ATPase activities at the contaminated sites was observed. The health status of these estuarine areas is quite similar, and the combined use of biomarkers is recommended.

Use of hepatocytes from Hoplias malabaricus to characterize the toxicity of a complex mixture of lipophilic halogenated compounds.

Organisms are continuously exposed to a plethora of anthropogenic toxicants daily released to the environment. In the present study, the effects of a mixture of halogenated organic compounds (HOCs) extracted from hepatic lipids were evaluated on the primary hepatocyte culture from fish Hoplias malabaricus. Cells were isolated through non-enzymatic perfusion protocol and cultured during 3 days to allow attachment. Two concentrations of the mixture of HOCs (10 ng ml(-1) [Mix10] and 50 ng ml(-1) [Mix50]) were tested in cells for 2 days by medium replacement. The control groups, with and without solvent (DMSO) were run in the same conditions. Both tested concentrations of HOCs increased the catalase and GST activities, but only the Mix50 increase the DNA damage and decreased the GSH concentration and cell viability. Lipid peroxidation increased in the Mix10 group, but it seems to be more a consequence of DMSO presence than the HOCs themselves. The DMSO at 0.1% increased the lipid peroxidation, GSH concentration, apoptosis and DNA damage. The present data suggest that DMSO interferes with the hepatocytes of H. malabaricus in culture and that the mixture of HOCs tested alters the redox state of the hepatocytes.

The effect of trichlorfon on acetylcholinesterase activity and histopathology of cultivated fish Oreochromis niloticus.

Among the most extensively used compounds in aquaculture for the treatment of parasitic organisms are the organophosphates (e.g., trichlorfon). The influence of trichlorfon, applied to the treatment dose, on the acetylcholinesterase (AChE) activity and gill histopathology of the Nile tilapia (Oreochromis niloticus L.) was evaluated. Cultivated fish were exposed to a single application of 0.25 ppm of trichlorfon. The results indicated a significant decrease in the muscular AChE activity in the treated individuals. After 4, 8, and 24h of exposure to trichlorfon, several histopathological alterations on gill of O. niloticus were observed but only edema and blood congestion were presented up to 72 h. No inflammatory processes were observed after 96 h of exposure. The treatment dose of trichlorfon causes biochemical effects to the fish. An indiscriminated use of this pesticide will probably decrease the production parameters of a cultivated fish such as in growth rates and final weight.

Effects of dietary Pb(II) and tributyltin on neotropical fish, Hoplias malabaricus: histopathological and biochemical findings.

Trahira (Hoplias malabaricus) used to investigate the effects of successive Pb(II) or tributyltin (TBT) dietary doses. After 70 days of acclimation, individuals were exposed to 21 microg Pbg(-1) or 0.3 microg TBTg(-1) (5-day intervals, 14 doses). Two experiments were conducted to investigate the histopathological effects (liver and kidney) and measure the cholinesterase activity (muscle and brain) after Pb(II) or TBT dietary doses. A number of morphological effects were observed in liver, including cytoskeleton disturbance, microautophagy of mitochondria, nuclear damage, and cell death. In kidney, necrosis area, increasing of the neutrophils cell number, changes in melano-macrophage centers, and free macrophages were frequently registered after both Pb(II) and TBT exposures. The cholinesterase activity was inhibited in muscle after 14 doses of Pb(II), but no effects were found in individuals exposed to TBT. In summary, this work is the first to report detailed in vivo toxic effects in tropical fish, H. malabaricus, after dietary sublethal exposure to Pb(II) and TBT.

Morphological and neurotoxicological findings in tropical freshwater fish (Astyanax sp.) after waterborne and acute exposure to water soluble fraction (WSF) of crude oil.

The water-soluble fraction (WSF) of crude oil is a complex highly volatile and toxic mixture of hydrocarbon chains (polyaromatics, heterocyclics), phenols, and heterocyclic compounds containing nitrogen and sulfur. To evaluate the toxic effects of WSF in tropical freshwater teleosts and to develop methodologies that could investigate the toxic mechanisms of WSF in tropical organisms, an acute toxicity experiment was conducted with Astyanax sp. Three dilutions (15%, 33%, and 50%) of WSF obtained from Campos Bay's crude oil (Brazil) were used to study morphological and biochemical responses of the fish. Prior to exposure, the distribution and rate of volatilization of the WSF into each aquarium for the same exposure period was quantified by spectrofluorimetry. Five individuals of Astyvanax sp. were exposed to duplicate WSF of 0, 15, 33, and 50% for each of 12-, 24-, and 96-h exposures for a total of 120 individuals. Liver and gills were sampled from five fish from each treatment and were analyzed by histology, scanning and transmission electron microscopy. A fragment of muscle was also collected from each fish to measure acetylcholinesterase activity. Water analysis showed that only 4 h after dilution, an important loss of hydrocarbons in 33% and 50% of WSF was observed. In addition, 50% of hydrocarbon mass was lost in all tested dilutions after 24 h with significant difference for the 50% WSF at all measured times, demonstrating the high volatility of WSF in freshwater. Damage in the liver and the gills included the presence of necrosis, loss of hepatocytes limit, inflammation areas, cellular proliferation, aneurysms, and disorganization of the second lamellae. The 33% WSF significantly reduced acetylcholinesterase activity in fish. Our study demonstrated that the WSF of crude oil caused damage in organs and tissues of tropical freshwater Astyanax sp. and provided also the basis for a better understanding of the toxic mechanisms of WSF in freshwater fishes.

Evaluation of tributyltin subchronic effects in tropical freshwater fish (Astyanax bimaculatus, Linnaeus, 1758).

Tributyltin (TBT) is widely used as a biocide in antifouling paints. Astyanax bimaculatus adult fish were acclimatized in a laboratory and isolated in groups of eight individuals. Two groups were used as a control (similar handling and corn oil control) and one group was exposed to TBTCI dissolved in corn oil (0.0688 +/- 0.0031 microg TBT.g(-1)) every 6 days for 32 days. Light and transmission electron microscopy were used to investigate the morphological effects on liver. A muscle fragment was excised for the determination of the acetylcholinesterase activity and blood smears were obtained for differential white cell counts. The results indicated nuclear irregular shapes, chromatin condensation, presence of intranuclear lipid bodies, and degenerative nuclei. Acetylcholinesterase activity was not affected by TBT exposure. The increasing number of neutrophils may represent cytotoxic and stress conditions facilitating the invasion of opportunist organisms. According to the authors' data, TBT was confirmed as a powerful aquatic contaminant to fish in tropical ecosystem.