Dysregulation of Intestinal Physiology by Aflatoxicosis in the Gilthead Seabream (Sparus aurata).

Aflatoxin B1 (AFB1) is a mycotoxin often present in food. This study aimed to understand the physiological effects of AFB1 on the seabream (Sparus aurata) gastrointestinal system. In a first in vitro approach, we investigated ion transport using the short-circuit current (Isc) technique in Ussing chambers in the anterior intestine (AI). Application of apical/luminal AFB1 concentrations of 8 and 16 µM to healthy tissues was without effect on tissue transepithelial electrical resistance (TER), and apparent tissue permeability (Papp) was measured using fluorescein FITC (4 kD). However, it resulted in dose-related effects on Isc. In a second approach, seabream juveniles fed with different AFB1 concentrations (1 and 2 mg AFB1 kg-1 fish feed) for 85 days showed significantly reduced gill Na+/K+-ATPase (NKA) and H+-ATPase (HA) activities in the posterior intestine (PI). Moreover, dietary AFB1 modified Isc in the AI and PI, significantly affecting TER in the AI. To understand this effect on TER, we analyzed the expression of nine claudins and three occludins as markers of intestinal architecture and permeability using qPCR. Around 80% of the genes presented significantly different relative mRNA expression between AI and PI and had concomitant sensitivity to dietary AFB1. Based on the results of our in vitro, in vivo, and molecular approaches, we conclude that the effects of dietary AFB1 in the gastrointestinal system are at the base of the previously reported growth impairment caused by AFB1 in fish.

EROD activity and cytochrome P4501A induction in liver and gills of Senegal sole Solea senegalensis from a polluted Huelva estuary (SW Spain).

EROD activity and induction cytochrome P4501A in liver and gills of Senegal sole, Solea senegalensis, from a heavy metal and PAH polluted estuary, was studied. Liver and gill CYP1A catalytic activity was assessed at the enzyme activity level-measured as 7-ethoxyresorufin-O-deethylase and cellular localization of CYP1A in the liver was studied using immunohistochemistry. Liver EROD was correlated with phenanthrene-type metabolites in liver and copper concentrations in water. Strong CYP1A occurrence was observed in acinar pancreatic cells, pancreatic duct epithelium and vascular system endothelium and negative/rare induction were observed in hepatocytes and sinusoidal endothelium. In gills, EROD activity showed a significant correlation with different fractions of heavy metals in sediment but no correlation was observed between EROD activity and PAHs. Strongly positive CYP1A associated staining of the vascular system endothelia and primary filament cells and a moderate staining of pillar cells in gills were observed. The results substantiated the utility of EROD activity and CYP1A induction measurement as biomarkers for use by aquatic toxicologists and indicate that catalytic assays and immunohistochemical assays appear to be sensitive to different kinds of pollutants being the use of both methods recommended for monitoring programs.

Biomarkers responses in muscle of Senegal sole (Solea senegalensis) from a heavy metals and PAHs polluted estuary.

The biochemical responses in muscle, such as the enzymatic activities of acetylcholinesterase, lactate dehydrogenase and isocitrate dehydrogenase, were studied in sole (Solea senegalensis) collected in Huelva estuary (SW Spain), in the vicinity of a petrochemical and mining industry. The sampling sites showed different type and degree of pollution. The results demonstrated significant differences in muscle activities of AChE and IDH in Odiel and Tinto Rivers compared to control fish. LDH activity did not show any difference between sampling sites. Significant correlations were established between some biomarkers and heavy metals: AChE was correlated with Pb, Cd and Cu concentrations in water; IDH activity was correlated with Cd and Cu concentrations in water and As, Pb and Cd concentrations in sediments; LDH activity was correlated with As and Zn concentration in water and Cd concentration in sediment. Only one correlation was established between the biomarkers analysed and the concentrations of PAHs: benzo(b)fluoranthene concentration in sediment and IDH.

Oxidative stress biomarkers in Senegal sole, Solea senegalensis, to assess the impact of heavy metal pollution in a Huelva estuary (SW Spain): seasonal and spatial variation.

The response of wild fish to heavy metals was studied in sole (Solea senegalensis) collected in 2004, 2005 and 2006 at three sampling sites from Huelva estuary (SW Spain), in the vicinity of a petrochemical and mining industry. Heavy metals As, Cd, Cu, Fe, Pb and Zn were analyzed in samples collected from sediment, water and tissue (liver) to examine their bioconcentration and effects in fish such as lipid peroxidation (LPO), catalase (CAT; EC 1.11.1.6), glutathione peroxidase (GPx; EC 1.8.1.7), glutathione S-transferase (GST; EC 2.5.1.18) and glutathione reductase (GR; EC 1.11.1.6) were also analyzed in the fish liver. The results showed different effects in sole from diverse locations with varying degrees of pollution. Significant differences in LPO, CAT and GR activities between control fish and fish from sampling sites were observed as well as seasonal differences for biomarkers. Significant correlations were established between some biomarkers and heavy metals concentrations in liver, sediment and water. This study indicates the usefulness of integrating a set of biomarkers to assess the effects of pollutants in aquatic environments under complex mix of pollutants and chronic pollution situation.

Toxicity of lindane (gamma-hexachloroxiclohexane) in Sparus aurata, Crassostrea angulata and Scrobicularia plana.

The purpose of the present study was to research the sublethal and/or lethal effects produced by the exposure of fish and shellfish to the gamma isomer of lindane, gamma-hexachlorocyclohexane (gamma-HCH). The teleostean fish Sparus aurata and the shellfish Crassostrea angulata and Scrobicularia plana, were exposed to 16 micro g/L of lindane for 15 days. Samples of different fish (liver, kidney and gills) and shellfish (gills, gut, digestive gland and mantle) tissues were extracted and processed for histopathological observations. Although mortality was not detected during the bioassay, sublethal effects (histopathological alterations) were observed. Vacuolization in the liver cells and lamellar fusion in gills from exposed fish were observed. Disorganization of normal gill structure, epithelial desquamation with the disappearance of apical ciliature in intestine, and inflammatory response in mantle from exposed shellfish were also observed. Thus, it can be concluded that the lindane concentration employed in the present research did not produce lethal effects in the exposed organisms but it caused sublethal effects. Lindane has time-dependent multiple toxic effects in S. aurata, C. angulata and S. plana, which were more severe at the end of the experimental time. The toxicological implications arising from these results are subjects for further multiconcentration tests dealing with lethal responses (mortality) or with sublethal responses (cellular/molecular biomarkers) of the aforementioned species.

Lindane toxicity on early life stages of gilthead seabream (Sparus aurata) with a note on its histopathological manifestations.

Eggs/embryos and larvae were exposed to nominal concentrations ranging from 0.1 to 10mg/L lindane. High percentage of mortality was observed in larvae exposed to 1mg/L (76.38%) and in embryos exposed to 10mg/L (81.98%) of lindane at 24h exposure. The acute toxicity expressed as LC(50) 48-h was 0.122mg/L for embryos and 0.318mg/L for larvae. Larvae alterations included weak swimming, incapacity to respond to external stimuli, uncoordinated movements, trembling, myoskeletal defects, opaque skin and exophthalmia. Mucous epithelium of the digestive tissue showed a severe alteration with hypertrophy and desquamation of mucous cells. A high cellular disorganization in the renal and hepatic tissue is observed. Results obtained showed the sensitivity of Sparus aurata early life stages to lindane and the presence of sublethal effects like histopathological alterations; therefore, the relevance of pesticides substances control in the aquatic environment.

Evaluation of acute copper toxicity during early life stages of gilthead seabream, Sparus aurata.

In this study, the effects of exposure to copper (mortality and morphological alterations) on the early life stages of the gilthead seabream, Sparus aurata, were examined. Eggs/embryos and larvae were exposed to nominal concentrations of copper ranging from 0.0001 to 10 mg/L Cu (II) in the tests with eggs/embryos and 0.025 to 0.5 mg/L Cu (II) in the test with larvae. Duration of the assays was 48 hours for embryos and 96 hours for larvae. A high percentage of mortality was observed in embryos exposed to 0.1 mg/L (97.2%) and in larvae exposed to 0.5 mg/L (100%). The embryos proved the most sensitive to copper for the same duration of exposure. The acute toxicity expressed as LC(50) 48 hours was 0.054 (0.048-0.058) mg/L for embryos and 0.261 (0.182- 0.375) mg/L for larvae. Morphological alterations or abnormalities in embryos included irregular shapes of chorion, opacity and vitellus retraction/degeneration. In larvae we observed poor capacity to swim, trembling, myoskeletal defects, opacity and exophthalmia. Histopathological alterations are observed in S. aurata larvae. Mucous cells of the digestive tissue present a severe alteration with an increment of exudates. A great cellular disorganization in the renal tissue is observed. Results from this work indicate the high sensitivity of early life stages of Sparus aurata to copper (II) and the persistence of sublethal effects.