Dietary probiotic supplementation in juvenile rainbow trout (Oncorhynchus mykiss) reared under cage culture production: effects on growth,fish welfare,flesh quality and intestinal microbiota

This study was conducted to evaluate the effects of dietary probiotic supplementation on growth performance, fish welfare, sensory evaluation, freshness during ice storage and changes in intestinal microbiota of rainbow trout (Oncorhynchus mykiss). The 9‐week feeding trial was conducted in cages under commercial conditions during the winter season, when water temperature decreased from 14°C to 7°C. Fish were fed either a diet without probiotic supplementation (control) or with 0.2% (2 × 10⁹ CFU kg^?1 diet) of multi‐strain probiotic bacteria (Bacillus sp., Pediococcus sp., Enterococcus sp., Lactobacillus sp.). Fish weight was recorded at 0, 2, 4, 6 and 9 weeks. Growth performance was not affected by dietary probiotic supplementation. Alternative complement activity (ACH50) was significantly higher in fish fed the diet with probiotic (58.4 ± 3.7 units mL^?1) than in the control group (46.9 ± 3.1 units mL^?1) (P = 0.03). Several biochemical markers were altered by the dietary treatment. 7‐ethoxyresorufin‐O‐deethylase (EROD) (P < 0.01), glutathione S‐transferase (GST) (P < 0.001), glutathione peroxidase (GPx) (P < 0.05) increased and lipid peroxidation (LP) (P < 0.01) decreased with dietary probiotic supplementation. Flesh quality and shelf‐life of fish stored on ice was not affected by dietary probiotic supplementation. Probiotic supplementation had a limited effect on the intestinal microbiota. Signal strength of the observed operational taxonomic units (OTUs) differed between the control and treatment samples and could indicate variations in the abundance of certain OTUs. Microbiota adhering to the intestinal mucus varied between sampling points, indicating that changes were associated with water temperature.     

Effect of different plant oils on growth performance,fatty acid composition and flesh quality of rainbow trout (Oncorhynchus mykiss)

This experiment intended to assess the effect of sesame (SO), sunflower (SFO) and linseed (LO) oils on growth performance, fatty acid composition of fillet and liver or flesh quality traits of rainbow trout. Fish fed different four iso‐nitrogenous and iso‐lipidic experimental feeds. The control feed contained only fish oil as the primary lipid source. The fillet eicosapentaenoic acid and docosahexaenoic acid levels were the highest in fish fed control feed. In contrast, the liver eicosapentaenoic acid level was the highest in fish fed LO feed. Fish fed SFO feed had the highest level of total n?6 fatty acids in fillet and liver. Fish fed SO feed had the highest level of 18:1 n?9 fatty acid in fillet and liver. During the 12 days refrigerated storage period at 1°C, thiobarbituric acid (TBA) and pH values gradually increased in all dietary groups. However, trimethylamin nitrogen (TMA‐N) values increased in all dietary groups between days 0 and 9 during the storage period. Generally, pH value in fillets of control group was slightly higher than the other fish groups during 12 days refrigerated storage. Nevertheless, the chemical indicators of spoilage, TBA, TMA‐N and pH values were in the limit of acceptability for human consumption. Results of growth performance and chemical tests in the present study showed that sesame, linseed and sunflower oils could be used in feeds for rainbow trout. Moreover, we concluded that further researches should be carried out on the partial replacement of fish oil by sesame oil in rainbow trout nutrition.     

Improvement of flesh quality in rainbow trout (Oncorhynchus mykiss) fed supranutritional dietary selenium yeast is associated with the inhibited muscle protein degradation

Supranutritional dietary selenium (Se) has been demonstrated beneficial for fish health, however, its effects on fish flesh quality remain unclear. This study investigated the effects of supranutritional dietary Se on the flesh quality of rainbow trout (Oncorhynchus mykiss) and their potential mechanism. Fish were fed a basal diet supplemented with or without graded Se yeast for 10 weeks. Results showed that Se supplementation significantly increased fillet crude protein and enhanced fillet water‐holding capacity as well as fillet firmness. Supplementing with both 2 and 4 mg/kg Se significantly downregulated the expressions of two autophagy–lysosome‐related genes (autophagy‐related 12‐like and gamma‐aminobutyric acid type A receptor‐associated protein‐like 1) in fish muscle, while supplementing with 4 mg/kg Se also significantly downregulated the expression of two ubiquitin–proteasome‐related genes (muscle RING finger 2 and F‐box protein 25). Correlation analysis indicated that the improved fillet quality parameters were closely correlated with the expressions of these differentially expressed genes. This study revealed that dietary Se was effective for the improvement of rainbow trout flesh quality, and the improved fish flesh quality was associated with the inhibited protein degradation in fish muscle.     

Immunohistochemical evaluation of experimental Vagococcus salmoninarum infection in rainbow trout (Oncorhynchus mykiss,Walbaum 1792)

The aim of this study was to investigate the pathogenesis and histopathological and immunohistochemical findings in rainbow trout (Oncorhynchus mykiss) following experimental vagococcosis. For this purpose, 60 rainbow trout were used. The experimental study used the pathogen Vagococcus salmoninarum. The fish were intraperitoneally (IP) administered with an inoculate containing 0.1 mL of the bacteria, resulting in a dose of 1.2 × 10⁹ cfu mL^?1 per fish. For histopathological observations, tissue samples were taken from fish that died during the experiment and fish that survived until the end of the trial (60th day). All the tissue samples were immunohistochemically stained by the avidin–biotin–peroxidase complex and immunofluorescence methods using polyclonal antibody to detect V. salmoninarum antigens. In immunoperoxidase staining, positive reactions to bacterial antigens were most commonly seen in the kidney, heart and liver. In the immunofluorescence analysis, the distribution of antigens in the tissue and organs was similar to that observed with the immunoperoxidase staining. The results reveal an important correlation between histochemical and immunohistochemical staining in demonstrating the distribution of V. salmoninarum antigens in the affected tissues.     

Effects of ascorbic acid enrichment by immersion of rainbow trout (Oncorhynchus mykiss, Walbaum 1792) eggs and embryos

This study was conducted to examine the effects of different forms and concentrations of ascorbic acid (vitamin C), and different enrichment times (24 and 48 h post ovulation) on egg, embryo and alevin ascorbate concentrations and survival of rainbow trout (enrichment was at the ova stage). In experiments 1 and 2, fertilized eggs were immersed in water containing ascorbate at 0 (control), 100, 1000 mg L^?1 l ‐ascorbic acid (AA) and 2000 mg L^?1 l ‐ascorbyl monophosphate (AP). In experiment 3, 0 (control), 500 and 1000 mg L^?1 AA neutralized (N) with NaOH, 1000 mg L^?1 AA non‐neutralized (NN), 1000 and 2000 mg L^?1 AP immersions were used. The mean total ascorbic acid (TAA) and dehydroascorbic acid (DHA) concentrations were measured before fertilization, at 3 and 24 h after fertilization, at the eyed stage, and in hatched alevins. We observed significant differences in TAA concentration at different immersion levels at 3 and 24 h after fertilization. Survival decreased significantly depending on the level of vitamin C, pH of the solutions and immersion time. We suggest that when broodstock rainbow trout do not have enough vitamin C in their ovaries, immersion of eggs in 1000 mg L^?1 of neutralized AA may be useful.     

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.     

Use of probiotics to control furunculosis in rainbow trout, Oncorhynchus mykiss (Walbaum)

Aerobic heterotrophic bacteria were isolated from the intestinal contents of Atlantic salmon, Salmo salar , rainbow trout, Oncorhynchus mykiss , and turbot, Scophthalmus maximus , on tryptone soya agar and De Man Rogosa and Sharpe agar, of which 11 of 177 (6% of the total) of the isolates were antagonistic to Aeromonas salmonicida . Four of these cultures, which were identified tentatively as A. hydrophila , Vibrio fluvialis , Carnobacterium sp. and an unidentified Gram-positive coccus, were beneficial to fish when fed singly or as an equi-mixture. Feed supplemented with the putative probiotics indicated survival of the organisms in the gastrointestinal tract for 7 days. Feeding with the probiotics for 7 and 14 days led to better survival following challenge with A. salmonicida . There was no indication of serum or mucus antibodies to A. salmonicida , but there was an increased number of erythrocytes, macrophages, lymphocytes and leucocytes, and enhanced lysozyme activity in the fish.     

Relationships between growth and disease resistance in rainbow trout,Oncorhynchus mykiss (Walbaum)

Rainbow trout from 23 families were evaluated for growth and resistance to the bacterial coldwater disease (BCWD) caused by Flavobacterium psychrophilum and infectious haematopoietic necrosis (IHN) caused by IHN virus. Average family weights were between 161 and 263 g with an average of 225 g at 213 days post‐fertilization with specific growth rates ranging from 2.37 to 2.88. Per cent survival of fish challenged with F. psychrophilum was between 18% and 100%, while for those challenged with IHNV, the range was between 12% and 93%. Significant positive correlations were found for end body weight and resistance to IHN (P < 0.05) and for early body weight and resistance to BCWD (P < 0.1). However, no significant correlations were detected between resistance to both pathogens or disease resistance and overall genetic diversity or diversity within the major histocompatibility locus.     

An experimental vaccine against Aeromonas hydrophila can induce protection in rainbow trout,Oncorhynchus mykiss (Walbaum)

A candidate vaccine against Aeromonas hydrophila in rainbow trout, Oncorhynchus mykiss, was developed using a bacterial lysate. To test the strength of protection, A. hydrophila challenge models were compared using injection into both the intraperitoneal (IP) cavity and the dorsal sinus (DS) with selected doses of live bacteria washed in saline or left untreated. Unlike the IP route, injection into the DS with either saline washed or unwashed cells resulted in consistent cumulative mortality and a dose response that could be used to establish a standard challenge having an LD₅₀ of approximately 3 × 10⁷ colony forming units per fish. Survivors of the challenge suffered significantly lower mortality upon re‐challenge than naïve fish, suggesting a high level of acquired resistance was elicited by infection. Passive immunization using serum from hyper‐immunized fish also resulted in significantly reduced mortality indicating protection can be transferred and that some portion of resistance may be antibody mediated. Vaccination of groups of rainbow trout with A. hydrophila lysate resulted in significant protection against a high challenge dose but only when injected along with Freund’s complete adjuvant. At a low challenge dose, mortality in all groups was low, but the bacterial lysate alone appeared to offer some protection.     

Continuous exposure to infectious pancreatic necrosis virus during early life stages of rainbow trout, Oncorhynchus mykiss (Walbaum)

Rainbow trout, Oncorhynchus mykiss (Walbaum), were exposed continuously to infectious pancreatic necrosis virus (IPNV) at 0, 10¹, 10³ or 10⁵ plaque forming units (pfu) L⁻¹ of water to estimate the effects of chronic IPNV exposure on early life stages. Fish density averaged 35 fish L⁻¹ (low density) or 140 fish L⁻¹ (high density), and the tank flow rate was 250 mL⁻¹ min. Virus exposure began at 6 days before hatch and continued until fish were 44 days old. Cumulative per cent mortality, analysis of survival and hazard functions, and discrete-time event analysis were used to explore the patterns of survival and mortality. In eggs and fish exposed to IPNV, mortality significantly greater than in the 0 pfu L⁻¹ exposure did not occur until IPNV concentration was 10⁵ pfu L⁻¹ at low fish density and 10³ pfu IPNV L⁻¹ at high fish density. These results suggest that in the natural aquatic environment, where rainbow trout densities are likely to be considerably lower than in this study, mortality resulting from infection with IPNV will very likely not occur when ambient concentrations of virus are ≤10³ pfu IPNV L⁻¹. In aquaculture rearing units, trout density is likely to be as high or higher than the densities used in this study. Therefore, continuous inputs of virus at concentrations greater than 10¹ pfu L⁻¹ may result in IPN epidemics in aquaculture facilities.     

Cellular components of probiotics control Yersinia ruckeri infection in rainbow trout, Oncorhynchus mykiss (Walbaum)

Subcellular components of the probiotics Aeromonas sobria GC2 and Bacillus subtilis JB-1, when administered to rainbow trout, Oncorhynchus mykiss , conferred protection against a new biogroup of Yersinia ruckeri . Thus, intraperitoneal or intramuscular injection of rainbow trout with cell wall proteins (CWPs), outer membrane proteins (OMPs), lipopolysaccharides (LPS), whole cell proteins (WCPs) and live cells followed by challenge on day 8 with Y. ruckeri led to 80–100% survival compared with 10% survival in the controls. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) profiles of WCPs and OMPs from GC2 had 10 and 5 variable protein bands in comparison to 11 and 5 bands in the WCPs and CWPs from JB-1. Proteomic analyses were employed following SDS-PAGE to categorize one dominant protein of 104.7 kDa from the CWPs of JB-1 and equated it with ' Bacillus spp. endoglucanase' with a Mascot score >69. These results point to the potential of using cellular components of probiotics for protection of fish against bacterial diseases.     

Transfer of toxaphene and chlordane into farmed rainbow trout, Oncorhynchus mykiss (Walbaum) via feed

The study was carried out to quantitate the transfer of toxaphene and chlordane compounds from commercial fish feed into the edible part of rainbow trout, Oncorhynchus mykiss (Walbaum) under normal rearing conditions. Trout were fed with unspiked high energy feed for salmon (fat content 26–30%) over a period of 19 months. The average weight of trout increased from 10 g to more than 2092 g, reaching sizes of 51 cm length. Considerable amounts of toxaphene and chlordane residues were transferred from fish feed into trout muscle. Toxaphene concentrations increased up to 8.6 µg (Σ toxaphene indicator compounds 1–3) kg^?1 wet weight (w.w.) and chlordane reached 5.3 µg Σoxy‐, trans‐, cis‐chlordane + t‐nonachlor kg^?1 w.w. Results are also discussed on the contaminant levels based on the fat content and the effect of sexual maturation is considered in this study. The data allow the establishment of transfer rates for toxaphene and chlordane congeners from high energy diet into the edible part of farmed rainbow trout.     

Performance and immunological responses of rainbow trout (Oncorhynchus mykiss) fed bioprocessed plant‐based proteins

Marine‐derived fish meal (FM) is a traditional component of commercial aquaculture feeds for many farmed fish species. Modern bioprocessing technologies have been developed to produce high‐protein plant‐based ingredients for aquafeeds to further reduce FM and other conventional animal protein sources. A 90‐day feeding trial using juvenile rainbow trout (Oncorhynchus mykiss) (27.9 ± 1.4 g; mean ± SE) was completed to evaluate growth, feed efficiency, general health and immunological responses to diets containing experimental plant‐based protein sources and reduced FM. Trout were fed one of four dietary treatments: experimental bioprocessed soy protein concentrate (BSPC), commercially available enzymatically hydrolysed soy protein concentrate (CSPC), experimental bioprocessed barley protein concentrate (BBPC) and a FM control (FMC). At trial termination, there were no significant differences in relative growth (RG) or specific growth rate (SGR) between fish fed either FMC or BSPC (p > .05). Fish fed either CSPC or BBPC resulted in significantly lower RG and SGR than the FMC (p < .05). Significant respiratory burst analysis differences were observed among treatments at day 90 (p < .01), for fish fed either FMC or CSPC diets. This assessment of bioprocessed plant‐based protein ingredients facilitates the characterization and incorporation of bioprocessed soy and barley protein as the industry continues to evaluate FM replacements in rainbow trout feeds.     

Effects of low‐level dietary microalgae supplementation on the distal intestinal microbiome of farmed rainbow trout Oncorhynchus mykiss (Walbaum)

In this study, high throughput 16S rRNA sequencing was used to investigate the effect of a novel whole‐cell dietary microalgae meal (Schizochytrium limacinum), on the distal intestinal microbiome of farmed rainbow trout, Oncorhynchus mykiss. Heterotrophic microalgae are rich in omega 3 polyunsaturated fatty acids, can be produced sustainably and have been shown to have beneficial effects on host health. After a 15‐week trial period, microbial community profiles were compared between the distal intestinal contents of fish fed either a control diet or a treatment diet that partially replaced fish oil with microalgae meal, at a substitution level of 5%. The results of this research showed that the microbial communities of both fish populations were composed of similar microbial taxa, however, the treatment group fed the microalgae supplement possessed a greater level of microbial diversity than those in the control group. A limited number of bacterial taxa were discriminatory between diets and were significantly elevated in the treatment group, notably operational taxonomic units (OTUs) assigned to the genera Streptococcus, Leuconostoc, Lactobacillus, Lactococcus and Weissella. However, the overall structure of the intestinal microbiome between control and treatment groups was not found to be significantly different. The treatment group displayed a heavier mean weight and condition factor at the end of the trial period. The results of this study suggest that the tested microalgae meal can be used as a replacement for a proportion of fish oil in aquafeeds, with minor changes to the intestinal microbiome of farmed rainbow trout, and positive effects on growth.