Using underwater currents as an occupational enrichment method to improve the stress status in rainbow trout.

This study investigated the effects of occupational enrichment, specifically underwater currents, on the stress status of rainbow trout (Oncorhynchus mykiss). A total of 540 fish were divided into three groups: control tanks without artificial currents (CO), tanks with randomly fired underwater currents (RFC), and tanks with continuous current throughout the day (CT). After 30 days, half of the fish in each group underwent a 5-day pre-slaughter fasting (5D), while the others were fed until the day before slaughter (0D). Fish in the RFC group exhibited lower levels of plasma cortisol and acetylcholinesterase enzyme activity in hypothalamus and optic tract than other groups, suggesting an improved stress status. RFC group also showed higher levels of non-esterified fatty acids (NEFA) in 5D fish and higher liver glycogen stores, suggesting improved energy reserves. In comparison, the CT group had higher LDH levels, possibly due to their increased swimming activity. The CO group had significantly lower NEFA levels at 5D compared to the RFC group, suggesting lower energy reserves. The RFC fish had darker and yellow-reddish skin and liver color, suggesting an improved stress status and lower lipid reserves, respectively. Overall, although a significant stress response was not observed in fasted individuals, possibly due to the relatively short fasting period, the study suggests that providing occupational enrichment using randomly fired underwater currents for 1 month helped to improve stress status in rainbow trout, indicating that occupational enrichment during the grow-out phase can positively impact the welfare of rainbow trout during routine handling procedures.

Cytotoxicity of three graphene-related materials in rainbow trout primary hepatocytes is not associated to cellular internalization.

As a consequence of increasing production and use of graphene-related materials (GRM), their release into the aquatic environment is likely to be expected. Development of appropriate model systems to assess their potential toxicity toward aquatic organisms is undoubtedly needed. Of particular relevance are primary cultures of fish hepatocytes, since they maintain similar functionalities as those of the original tissue. Isolated hepatocytes from rainbow trout (Oncorhynchus mykiss) were exposed to ranges of concentrations of different forms of GRM, two graphene oxides (GO) of sheet-like structure and one tubular-shaped carbon nanofiber (CNF) in the presence or absence of fetal bovine serum (FBS) for 24 and 72 h. Metabolic activity, cell membrane integrity, lysosomal function, reactive oxygen species (ROS) formation and interaction with cytochrome P450 1 A enzyme were assessed by using AlamarBlue, 5-carboxyfluorescein diacetate-acetoxymethyl ester, neutral red uptake, dichlorofluorescein and 7-ethoxyresorufin-O-deethylase (EROD) assays, respectively. In the presence of FBS, GO affected metabolic activity and cell membrane integrity more than CNF, whilst absence of serum further reduced cell viability in GRM-exposed cells. GRM did not alter lysosomal function nor did it induce ROS formation or EROD activity. Intracellular uptake was observed only in the case of CNF when incubated without FBS. Primary hepatocytes from rainbow trout appear to be a suitable model to screen for cytotoxicity and to reveal any interaction with GRM. Results emphasize the role of serum proteins in the toxicological responses following exposure to GRM with important implications for the environmental risk assessment of these nanomaterials.

Physio-metabolic response of rainbow trout during prolonged food deprivation before slaughter.

Fish normally undergo periods of food deprivation that are longer than non-hibernating mammals. In aquacultured rainbow trout (Oncorhynchus mykiss), it is unclear how fasting may affect their physiological adaptative response, especially when they are normally fed daily. In addition, that response may vary with temperature, making it necessary to express fasting duration in terms of degree days. In the current study, trout were fasted for 5, 10, and 20 days (55, 107, and 200 degree days (°C d), respectively). To assess the physiological response of fish to fasting, different biometric, blood, plasma, and metabolic parameters were measured, as well as liver fatty acid composition. The fish weight, condition factor, and the hepato-somatic index of 5-day fasted trout were not significantly different from those of control fish. Gastric pH increased as fasting progressed while plasma concentrations of glucose, triglycerides, and total proteins decreased significantly after 10 days of fasting, while the percentage of non-esterified fatty acids increased. There were no significant differences in plasma ions (sodium, potassium, and calcium), except for chloride ion which decreased after 5 days of fasting. Liver glycogen decreased after 5 days of fasting while glycogen concentration in muscle did not decrease until 20 days of fasting. Liver color presented a higher chroma after 5 days of fasting, suggesting a mobilization of reserves. Finally, acetylcholinesterase activity in the brain was not affected by food deprivation but increased after 10 days of fasting in liver and muscle, suggesting the mobilization of body reserves, but without severely affecting basal metabolism.

Tissue distribution of zinc and subtle oxidative stress effects after dietary administration of ZnO nanoparticles to rainbow trout.

The increasing use of ZnO nanoparticles (ZnO NPs) in different fields has raised concerns about the possible environmental risks associated with these NPs entering aquatic systems. In this study, using a dietary exposure route, we have analysed the tissue distribution and depuration pattern of Zn as well as any associated redox balance disturbances in rainbow trout (Oncorhynchus mykiss) following exposure to ZnO NPs (20-30nm). Fish were fed a diet spiked with ZnO NPs prepared from a dispersion in sunflower oil at doses of 300 or 1000mg ZnO NPs/kg feed for 10days. This uptake phase was followed by a 28days depuration phase in which fish from all groups received untreated feed. While no overt signs of toxicity were observed and no important effects in fish growth (weight and length) or in the hepatosomatic index among groups were recorded, we observed high levels of Zn bioaccumulation in the gills and intestine of exposed fish following exposure to both dose levels. Zn levels were not eliminated during the depuration phase and we have evidenced oxidative stress responses in gills associated with such long term ZnO NPs bioaccumulation and lack of elimination. Furthermore, exposures to higher doses of ZnO NPs (1000mg/kg feed) resulted in Zn distribution to the liver of fish following 10days of exposure. Fish from this exposure group experienced biochemical disturbances associated with oxidative stress in the liver and ethoxy-resorufin-O-deethylase (EROD) activity which may point to the ability of ZnO NPs or its ions to interfere with cytochrome P450 metabolic processes.

Use of fish farms to assess river contamination: combining biomarker responses, active biomonitoring, and chemical analysis.

Here we addressed the possible effects of trace levels of contaminants on fish by means of a combination of biomarker responses, active biomonitoring (ABM), and chemical analysis. In environmental studies, cytochromes P4501A (Cyp1A) and Cyp3A and related enzyme activities (7-ethoxyresorufin-O-deethylase, EROD, and benzyloxy-4-[trifluoromethyl]-coumarin-O-debenzyloxylase, BFCOD, respectively) are commonly used as biomarkers for evidencing exposure to a variety of contaminants. In a rainbow trout (Oncorhynchus mykiss) fish farm that is routinely sampled to obtain references regarding normal levels of such enzyme activities in freshwater fish, we observed a strong and punctual increase in these activities at the end of 2011. In order to shed light on the causes of this induction, we transferred some fish to a fish farm with controlled conditions and examined them using an active biomonitoring (ABM) approach. EROD activity showed a decrease of 80% from the original values after 7 days in the control farm, while BFCOD activity was also reduced after 15 days. Although not significant, a decrease in cyp1A and cyp3A mRNA levels was also observed. To determine the presence of pollutants, water and sediment samples from the river feeding the fish farm were analyzed by two-dimensional gas chromatography-time-of-flight mass spectrometry (GC×GC-TOF-MS). The screening study reflected a weak inflow of pollutants in the monitored area, which is located far from any industrial activity or densely populated cities. Trace levels of polyaromatic hydrocarbons (PAHs) and personal care products (the polycyclic musk fragrance HHCB, and triclosan) were detected in sediments, at concentrations ranging from 0.01 to 38 ng/g dry weight, and in water from 4 to 441 ng/L. The approach followed in this study proved useful as a biomonitoring technique for the early detection of trace contaminants.