Beneficial effects of thyme (Thymus vulgaris) extract on oxytetracycline‐induced stress response,immunosuppression, oxidative stress and enzymatic changes in rainbow trout (Oncorhynchus mykiss)

The aim of the present study was to investigate the effects of thyme extract (TE) and oxytetracycline (OTC) on rainbow trout (Oncorhynchus mykiss) stress response, immune parameters, oxidative stress and enzymatic changes. The fish were assigned into eight treatments receiving diets with OTC (0 and 2.5 g/kg) and TE (0, 5, 10 and 20 g/kg) for 2 weeks. Thereafter, serum characteristics and gut oxidative status were evaluated. OTC significantly increased serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, and cortisol and glucose levels; however, TE significantly suppressed these changes. OTC significantly decreased, but TE significantly increased gut catalase (CAT) and glutathione peroxidase (GPx) activities. OTC significantly decreased gut superoxide dismutase (SOD) and increased gut glutathione‐s‐transferase (GST) activities, malondialdehyde (MDA) content and blood monocyte percentage; TE significantly mitigated these changes. TE significantly increased serum total protein, total Ig, lysozyme and ACH50, and blood WBC; however, OTC significantly decreased serum lysozyme and ACH50 activities and blood WBC and neutrophil percentage. In conclusion, OTC administration causes immunosuppression, which might be related to deterioration of fish health. TE augments trout innate immune and antioxidant status and is capable to counteract OTC‐induced health and immune deterioration.     

Dietary essential amino acids and heat increment in rainbow trout (Oncorhynchus mykiss)

Oxygen consumption attributable to apparent heat increment (AHI) was measured in relation to varying essential amino acid proportions (EAA) infused into rainbow trout,Oncorhynchus mykiss (250–450 g), induced to swim at 1 BL s^–1. Five diets, mimicking EAA concentrations in trout whole body protein, deficient in the branched chain amino acids (isoleucine, leucine and valine), containing unbalanced proportions of EAAs and supplying lysine in excessive and limiting proportions, were tested. Following infusion of the experimental diets, a significant increase in oxygen consumption was observed. Changes in plasma EAAs following infusion paralleled the time course of AHI (i.e., oxygen consumption). AHI represented the equivalent of 15–32% of the gross energy intake depending on dietary EAA composition. Diets supplying EAAs similar to trout whole body protein and limiting in lysine produced the lowest AHI values, indicating efficient utilization of dietary amino acids. Higher AHI values were associated with diets deficient in the branched chain amino acids and diets supplying lysine in excess. Duration of elevated metabolism was independent of both dietary composition and energy intake. Different proportions of EAAs in the diet can increase the energy expended as AHI. In an attempt to reduce the energy liberated as AHI, attention must be paid to the quality, quantity and balance of dietary EAAs.     

Edwardsiellosis in farmed rainbow trout (Oncorhynchus mykiss)

Edwardsiella tarda was isolated from naturally infected rainbow trout (Oncorhynchus mykiss) in raceway culture on a commercial fish farm where the fish were kept at a density of 110 kg m^?3 and at a water temperature of 14°C and with a photoperiod of 13 h light:11 h dark. The clinical signs of diseased fish (150 ± 20 mm standard length) were anorexia and lethargy. The most striking lesions in the fish were in the liver. There were hyperaemia and haemorrhages; on histopathological examination, the liver displayed inflammatory infiltrate in portal area, focal necrosis, dilatation of blood sinus and activation of sinusoidal cells. Infection experiments, performed 2 years after isolation of the original culture of E. tarda, were carried out under laboratory conditions at water temperatures of 15, 18 or 24°C. All experimental fish (common carp, Prussian carp, tench), intraperitoneally injected with 8 × 10⁶ cells, demonstrated a total resistance to E. tarda.     

Effects of hydroalcoholic extract of honeybee pollen on growth performance,flesh quality,and immune and stress response response of rainbow trout (Oncorhynchus mykiss)

The effects of dietary hydroalcoholic extract of honeybee pollen (HBPE) on growth performance, flesh quality, immunity and stress response were investigated. Two hundred fifty‐five rainbow trout (11.14 ± 1.06 g) were fed five diets containing increasing levels of HBPE: control, 0.25HBPE (0.25 g HBPE/kg), 0.5HBPE (0.5 g HBPE/kg), 1HBPE (1 g HBPE/kg) and 1.5HBPE (1.5 g HBPE/kg). After 56 days, rainbow trout fed the 0.5HBPE and 1HBPE diets displayed significantly higher weight gain (49.58 and 53.25 g) and protein efficiency (2.88 and 2.83) compared to those fed the control diet (4.89 and 2.05), respectively (p < .05). For flesh quality, higher protein content in the whole body of fish fed the 1HBPE (633.3 g/kg) and 1.5HBPE (640.9 g/kg) diets was observed when compared to other groups (p < .05). There were no significant differences in the saturated and monounsaturated fatty acid content. Individuals fed the 1HBPE (234.5 g/kg) and 1.5HBPE (234.7 g/kg) diets exhibited higher levels of n‐3 long‐chain polyunsaturated fatty acids in the muscle than the other groups (p < .05). In terms of the immune system, the serum lysozyme and alternative complement pathway haemolytic activity levels in the 0.5HBPE (47.66 and 148.00 U/ml) and 1HBPE (46.00 and 146.33 U/ml) treatments were higher than the other treatments (p < .05). When Aeromonas hydrophila was exposed to different dosages of HBPE, a higher inhibitory zone resulted from 1,000 and 1,500 mg/ml dosage. The lower levels of plasma cortisol were observed in the 1HBPE and 1.5HBPE groups after the complement stress test. Collectively, the present findings suggest that the 1HBPE diet supported superior growth, flesh quality, immunity and stress response of rainbow trout.     

Oxidative stress parameters during starvation and refeeding periods in Adriatic sturgeon (Acipenser naccarii) and rainbow trout (Oncorhynchus mykiss)

This work analyses the changes in the redox balance in two fish species: Adriatic sturgeon ( Acipenser naccarii ) and rainbow trout ( Oncorhynchus mykiss ) during starvation and refeeding period. The starvation period raised the lipid peroxidation (thiobarbituric-acid-reacting substances) levels in liver and blood, while a decline occurred in the antioxidant enzymatic activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX) and glutathione reductase (GR) in both fish species. In liver, after the refeeding period, SOD activity recovered in both species, whereas CAT activity recovered only in trout. Furthermore, in both tissues of the two species, the lipid peroxidation levels remained high after 2 months of refeeding. In white muscle and heart, the lipid peroxidation levels indicate that these tissues did not undergo oxidative stress during the 72-day period. During starvation, in the muscle of both fish the fall in the lipid peroxidation level coincided with a rise in CAT, GPX and GR. The refeeding period in this tissue raised the lipid peroxidation level, and the enzymatic activities reached the values of the first point of starvation. In heart, no oxidative damage was detected during starvation in either species. The CAT and SOD activities increased during the starvation period only in trout.     

The garlic component,allicin, prevents disease caused by Aeromonas hydrophila in rainbow trout,Oncorhynchus mykiss (Walbaum)

Allicin was fed at 0 (= control), 0.5 and 1.0 mL of Allimed^® liquid 100 g^?1 of feed for 14 days to rainbow trout, Oncorhynchus mykiss (Walbaum), fingerlings before infection with Aeromonas hydrophila with a resultant reduction in mortalities from 80% in the controls to 8% [relative percentage survival (RPS) = 90%] and 0% (RPS = 100%) among the treated fish. Allicin was strongly antibacterial compared to the control, with a minimum inhibitory concentration (MIC) of >400 μL mL^?1 of Allimed^® liquid. Use of allicin led to a lower number of white blood cells (132.0 ± 0.4 × 10³) compared to 175.0 ± 0.1 × 10³ in the controls, but elicited increased phagocytic activity, i.e. a phagocytic value of 39.2% compared to 13.6% in the controls, and serum lysozyme activity, which showed significant (P > 0.05) differences compared to the control at 15 and 30 min after the first reading at 0 min of incubation.     

Elevated concentrations of T‐2 toxin cause oxidative stress in the rainbow trout (Oncorhynchus mykiss)

T‐2 toxin is a mycotoxin produced by several Fusarium fungi that can contaminate plant components used in feed for aquaculture. The aim of this 28‐day study was to investigate the effect of the T‐2 toxin in feed in concentrations 1.0 and 1.8 mg/kg (0.01 mg/kg b.w. and 0.018 mg/kg b.w.) on the oxidative stress markers and on the detoxifying enzymes of the rainbow trout. The results showed that T‐2 toxin in both tested concentrations induced oxidative stress and antioxidant defence in the liver of trout manifesting by the increase in activities of enzymes glutathione‐S‐transferase, glutathione reductase and glutathione peroxidase and the decrease in the catalase activity. The increase in lipid peroxidation was recorded only in the higher concentration of T‐2 toxin. Ceruloplasmin activity in the plasma increased at both tested concentrations, and the ferric reducing ability of the plasma increased at the higher toxin concentration. The T‐2 toxin in feed caused alteration of the total protein, albumin and triacylglycerols as well as the alkaline phosphatase activity in the plasma. Despite no changes in histological examination were found, the influence of T‐2 toxin on scavenger system may result in increased sensitivity to other stress factors.     

Metabolism of exogenous histamine in rainbow trout (Oncorhynchus mykiss)

Information on the metabolism of exogenous histamine in fish is of much concern regarding the effect of dietary histamine on fish. Histamine catabolic enzymes, diamine oxidase and histamine N-methyl transferase were measured in the tissues of rainbow trout. Diamine oxidase was detected in the stomach, pylorus caeca and intestine. Histamine N-methyl transferase was detected only in the liver.A change in the contents of histamine and its metabolites was observed in the tissues of rainbow trout after oral administration of histamine. A large amount of imidazole acetic acid was observed in the serum, kidney, liver and muscle. On the other hand, 1-methyl histamine was detected only in the liver. Histamine and its metabolites, imidazole acetic acid and 1-methyl histamine were metabolized and diminished within 48 hr in all tissues.These results showed that histamine was metabolized by two metabolic routes in rainbow trout. One is the main pathway producing imidazole acetic acid by intestinal diamine oxidase and the other is the complementary pathway producing 1-methyl histamine by liver N-methyl-transferase.     

Dietary modulation of digestive enzymes by the administration of feed additives to rainbow trout,Oncorhynchus mykiss Walbaum

The aim of this study was to assess the metabolic and digestive enzyme activities in rainbow trout, Oncorhynchus mykiss Walbaum fed dietary supplements. The biometric indices and the following enzymes activities were measured: acid protease and pepsin from the stomach homogenates, alkaline phosphatase (AP) from the small intestine and the brush border membrane, total proteolytic enzymes and trypsin from the small intestine and liver/pancreas. Dietary garlic induced a higher pepsin activity in the stomach than lipopolysaccharide (LPS) and ginger. Conversely, dietary ginger and LPS showed a significant difference (P > 0.05) in acid protease activity compared to the controls. There was not any significant difference observed between treated groups and the controls for hepato‐pancreas proteolytic enzyme activity, except for trypsin. AP activity increased significantly with dietary garlic, ginger and LPS compared to the controls.     

Effects of dietary nucleotides on the antioxidant status and serum lipids of rainbow trout (Oncorhynchus mykiss)

An 8‐week feeding experiment was conducted to evaluate the effects of dietary nucleotides (0, 0.5, 1, 1.5 and 2 g kg^?1) on serum antioxidative biomarkers and lipid contents of rainbow trout fingerlings. The serum triglycerides and low‐density lipoprotein (LDL)‐C levels were significantly decreased in fish fed diets containing 1.5 and 2 g kg^?1, whereas serum high‐density lipoprotein (HDL)‐C was higher in trout receiving diets with those levels of nucleotides. A significant decrease in thiobarbituric acid reactive substances concentration was observed in groups receiving 1.5 or 2 g kg^?1 of dietary nucleotides in comparison with control group (?32% and ?27%, respectively). A significant decrease was observed in catalase and serum glutathione peroxidase activity in trout fed 1.5 and 2 g kg^?1 dietary nucleotides compare with other groups. There were no significant changes in serum superoxide dismutase activity among the experimental groups. In conclusion, these results suggest that dietary nucleotides may reduce lipid peroxidation by increasing the levels of antioxidant lipoprotein, HDL‐C, and decreasing the LDL‐C, the main substrate of lipid peroxidation.     

Antioxidant and immune defences of rainbow trout (Oncorhynchus mykiss) offered plant oils differing in fatty acid profiles from early stages

The antioxidant defence and immune response of rainbow trout (Oncorhynchus mykiss) that had received plant oils, rich in either n‐3 polyunsaturated fatty acids (PUFA; linseed oil) or n‐6 PUFA (safflower oil) was evaluated upon antigen exposure. The fish employed in this study had been offered the diets for 18 months from the first feeding. Rainbow trout from each group were injected intraperitoneally with formalin‐killed bacteria (Aeromonas hydrophila) or were sham‐injected and observations were made 24 h later. Though the fish fed safflower oil seemed to be under relatively greater oxidative stress, the antioxidant defences (superoxide dismutase, catalase, glutathione peroxidase) were as effective as in those fed linseed oil. The humoral (alternate complement activity and lysozyme activity) and cellular (phagocytic activity and lymphocyte proliferation) immune responses were not significantly affected by the oil offered. With the exception of reactive oxygen production that was significantly greater in the linseed oil fed fish, both groups did not differ greatly in their immune responses after antigen exposure. Thus, fish fed safflower oil that was deficient in n‐3 PUFA was able to sustain most of the critical responses similar to those fed linseed oil suggesting that plant oils of both fatty acid categories were effective for this fresh water fish.     

Dietary formic acid enhances apparent digestibility of minerals in rainbow trout, Oncorhynchus mykiss (Walbaum)

The effect of dietary formic acid on the availability of phosphorus (P) from a fishmeal-based diet by rainbow trout, Oncorhynchus mykiss (Walbaum), reared in brackish water (5–6 g L⁻¹) was determined. Diets containing a low amount of P (6.0 mg P g⁻¹ dry matter) were acidified with 0, 4 and 10 mL kg⁻¹ formic acid and fed to trout (average weight, 520 g) for 4 weeks. The measured pH of the diets were 6.3, 5.8 and 5.3, respectively. The intestinal solubility of P and the digestibility of P were measured by stripping the faecal matter from the fish. The apparent digestibility coefficient (ADC) of P significantly ( P < 0.05) increased from 69.5% to 75.0% of the basal diet in fish fed diets containing 10 mL kg⁻¹ formic acid. The solubility of P in the intestine was highly variable within each treatment, and the differences were not significant. The pH of intestinal content increased with the increase in dietary formic acid concentration. The ADC of magnesium and calcium also showed a significant ( P < 0.05) increase with the acidification of diet by formic acid.     

Embryonic expression of UCP2 in rainbow trout (Oncorhynchus mykiss)

Uncoupling proteins are mitochondrial anions transporters that dissociate respiration from ATP synthesis through proton leaks. Uncoupling protein 2 reportedly plays a role in several physiological processes such as energy partitioning, nutrition, and fatty acid metabolism. The mRNA expression of rainbow trout UCP2 genes (UCP2A and UCP2B) was monitored during embryogenesis and early larval development. Both genes were recruited early and displayed similar steadily decreasing patterns from fertilization until hatching. The expression of UCP2A and UCP2B appeared significantly differentiated after hatching and during the yolk sac absorption, with UCP2A displaying higher expression. We suggest that UCP2 expression profiles in the rainbow trout embryo could be associated with the utilization of lipids as a source of energy during development.     

Trial of probiotics to prevent the vertebral column compression syndrome in rainbow trout (Oncorhynchus mykiss Walbaum)

Two probiotics were tested as alternative treatment to limit the prevalence of the vertebral column compression syndrome (VCCS) in rainbow trout, compared with a preventive treatment with florfenicol. Either the antibiotic, or a lactic acid bacterium, Pediococcus acidilactici, or a yeast, Saccharomyces cerevisiae var. boulardii, was introduced into experimental diets, which were compared with the control diet without supplementation. The antibiotic caused some mortality during the first week after treatment, likely because of the toxicity of the compound. However, this antibiotic treatment limited the occurrence of VCCS to 3%, vs. 13% of fish affected in the control group. Pediococcus acidilactici provided the same level of protection as the antibiotic, but on condition that the treatment was applied during the 5 months of the experiment, without any adverse effect on survival. A treatment with P. acidilactici only during the first 20 days of feeding was not sufficient to limit VCCS, nor was the treatment with the yeast sufficient. These results were discussed while comparing the microbiota associated with the intestine. The experiment reinforced the hypothesis that pathogenic bacteria are involved in VCCS. The long‐term dietary supplementation with P. acidilactici seemed promising as a preventive treatment against the syndrome, but large‐scale investigation in fish farms will be necessary to confirm its prophylactic relevance.     

A comparative tissue distribution study of oxytetracycline in rainbow trout, Oncorhynchus mykiss (Walbaum), and chinook salmon, Oncorhynchus tshawytscha (Walbaum)

Oxytetracycline (OTC), a broad-spectrum antibiotic, is used widely to treat bacterial diseases in farmed fish. In the present study, the time course of OTC concentrations in freshwater rainbow trout, Oncorhynchus mykiss (Walbaum), and seawater chinook salmon, Oncorhynchus tshawytscha (Walbaum), were compared, tissue by tissue, after receiving a bolus dose of the antibiotic (5 mg kg^–1 or 50 mg kg^–1) intra-arterially (i.a.). The OTC concentration–time profiles of rainbow trout tissues were found to be very similar to those of the corresponding tissues in chinook salmon. Therefore, neither water salinity nor fish species seemed to play an important role in the disposition and elimination of OTC in these salmonids. In a separate experiment, rainbow trout were implanted surgically with a urinary cannula and received a single dose of OTC (50 mg kg^–1) i.a. Urine was collected from the cannula daily for 13 days. The amount of OTC excreted into the bile was found to be larger than that eliminated by the urine. These results show the similarity of OTC pharmacokinetics in freshwater rainbow trout and seawater chinook salmon and render support in using a single fish species to study the pharmacokinetics of a drug for other species in the same taxon.     

Dietary probiotic Pediococcus acidilactici MA18/5M modulates the intestinal microbiota and stimulates intestinal immunity in rainbow trout (Oncorhynchus mykiss)

A study was conducted to evaluate the probiotic effect of Pediococcus acidilactici MA18/5M on rainbow trout, Oncorhynchus mykiss. Fish (310 ± 9 g) were fed a control diet or a P. acidilactici‐supplemented diet (at 2.4 × 10⁶ CFU/g) for 4 weeks. The probiotic was observed to populate the intestine with levels ranging from log 3.7 to 5.4 CFU/g. Furthermore, these populations were able to persist for at least 24 hr after the cessation of probiotic feeding. High‐throughput sequencing analysis of bacterial 16S rRNA libraries demonstrated that P. acidilactici was able to modulate the gut microbiome of rainbow trout and that the probiotic was detected as a common taxon on the mucosa and in the digesta of the probiotic fish (p < .05). Real‐time polymerase chain reaction demonstrated that feeding the probiotic upregulated pro‐inflammatory cytokines, interleukin‐1β, and interleukin‐8 and downregulated anti‐inflammatory interleukin‐10 compared to the control‐fed fish. Furthermore, the mRNA levels for the mucosal antibody immunoglobulin T was also elevated in probiotic‐fed fish. These findings help to explain some of the mechanisms behind the previously reported observed benefits of using this probiotic in the intestinal morphology and immunity of rainbow trout.     

Use of dietary ginger, Zingiber officinale Roscoe, as an immunostimulant to control Aeromonas hydrophila infections in rainbow trout, Oncorhynchus mykiss (Walbaum)

Ginger, Zingiber officinale, which was fed at 0, 0.05, 0.1, 0.5 and 1.0 g per 100 g of feed for 14 days to rainbow trout, Oncorhynchus mykiss (Walbaum), led to control of experimental infection with Aeromonas hydrophila . At 0.5 g ginger per 100 g of feed, there was a reduction in mortalities to 0% compared with the controls (64%). Moreover, there was a significant increase in growth, feed conversion and protein efficiency. There was proliferation in the number of neutrophils, macrophages and lymphocytes, and enhanced phagocytic, respiratory burst, lysozyme, bactericidal and anti-protease activities compared with the controls.