Research Article |
Corresponding author: Olga Revina ( dr.revina@gmail.com ) Academic editor: Predrag Simonović
© 2023 Olga Revina, Vjačeslavs Revins, Dina Cīrule, Anda Valdovska.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Revina O, Revins V, Cīrule D, Valdovska A (2023) TNF-Α, IL-6, HSP-70, fish growth hormone, and growth performance of sea trout, Salmo trutta (Actinopterygii: Salmoniformes: Salmonidae) after long-term dietary administration of β-glucan and BGN-2. Acta Ichthyologica et Piscatoria 53: 253-261. https://doi.org/10.3897/aiep.53.113220
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The presently reported study intended to examine the effect of the oral administration of an immunomodulator β-glucan and β-glucan-containing product (BGN-2) on the levels of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), fish heat shock protein 70 (HSP-70), and fish growth hormone (GH) as well as growth performance of cultured sea trout juveniles. The sea trout is a migratory sea-run brown trout, Salmo trutta Linnaeus, 1758. The fish (total of 15 000) were divided into four experimental and one control group including control (consisting of basal diet) (D1); basal diet + 1 g kg–1 β-glucan (D2); basal diet + 3 g kg–1 β-glucan (D3); basal diet + 6 g kg–1 BGN-2 (D4); basal diet + 14 g kg–1 BGN-2 (D5). The fish fed D4 and D5 diets showed significantly higher IL-6, HSP-70, and GH expression levels than other treatments (P < 0.05). Sea trout fed D4 and D5 diets showed significant improvements in growth performance compared to the fish fed the control diet. In conclusion, our results suggest that dietary supplementation with the product BGN-2 provides great immunostimulation and could be used as an effective measure to improve growth performance in juvenile sea trout.
glucan, immunostimulants, sea trout
The decline of wild salmonid populations is a direct result of extensive human influence and the consequences of climate change on their natural environment (
Therefore, there is a pressing need for well-structured breeding and rearing initiatives that emphasize nutritional approaches. This includes the study of feeding behaviors and the development of initial diets that are crafted from ecologically sustainable components (
Immunological studies prove that the immune system of fish is comparable to those of mammals, characterized by different types of leukocytes, complement factors, immunoglobulins (Ig), toll-like receptors (TLRs), major histocompatibility complex (MHC), T-cell receptors (TCR), and cytokines (
Researchers have demonstrated that β-glucan can modulate essential biochemical markers, such as serum hemoglobin, serum protein, and total hemocyte count, as well as crucial immunological parameters like lysozyme activity, phagocytic activity, oxidative burst activity, and phenoloxidase activity. This leads to a more robust immune profile, making it highly valuable in the treatment of fish and aquatic organisms. Fish supplemented with β-glucan have shown reduced sensitivity in genes associated with acute inflammatory reactions (
TNF-α and interleukin-6 (IL-6) are among the most significant cytokines produced by macrophages and monocytes and are listed as a humoral immune component (
Heat shock proteins (HSPs) are a family of highly conserved stress proteins that are produced by cells in response to stressful conditions and are indicators of cellular stress and animal health status (
Fish growth hormone (GH) has a role in feeding, growth-promoting, immune function, and osmoregulation in teleost fish (
The presently reported study intended to investigate the immunomodulatory effects of two Saccharomyces cerevisiae-derived (yeast) β-glucan products on juvenile sea trout. These products included a commercially available purified β-glucan product (Angel Yeast Co., Ltd., China), as well as a biological product known as BGN-2 (JP Biotechnology, Ltd., Latvia), using a patented technological method. BGN-2 comprises derivatives of the yeast S. cerevisiae obtained during the ethanol production process from grain (
In the presently reported study, the hypothesis suggests that the prolonged administration of various oral doses of β-glucan products exerts an influence on the expression of TNF-α, IL-6, HSP-70, and GH, potentially enhancing both the health and growth performance of sea trout.
Experimental fish and culture condition. The research was developed at the state fish farm Pelci of the Institute of Food Safety, Animal Health and Environment BIOR, Latvia (56°55′16.3″N, 021°58′28.6″E) (Pelci) using riverine water.
At the farming trial, 15 000 sea trout juveniles (2.3 ± 0.30 g, mean weight ± SE) were randomly placed into five 1.8 m3 tanks (n = 3000 in each tank), with a volume of 1.2 m3, in a flow-through rearing system. The quality of water was regularly monitored. The fish were acclimated for 2 weeks.
All of the sea trout used in the trial were planned to be released in natural watercourses in April of the same year, according to the Restocking plan of fish resources 2017–2020, Latvia (
Fish diet and preparation of feed. A basal diet was formulated according to the nutrition requirement of sea trout (
Five experimental diets were designed as follows: control (consisting of basal diet) (D1), basal diet + 1 g kg–1 β-glucan (D2), basal diet + 3 g kg–1 β-glucan (D3), basal diet + 6 g kg–1 BGN-2 (D4), and basal diet + 14 g kg–1 BGN-2 (D5). The dosage of Angel yeast product was 1 g kg–1 and 3 g kg–1; the dosage of JP Biotechnology product (BGN-2) was 6 g kg–1 and 14 g kg–1, equivalent to the final concentration of pure β-glucan in the final feed in a ratio of 1 g kg–1 and 3 g kg–1 (according to JP Biotechnology data).
The feed was delivered by an automatic fish feeder following a feeding regime of 4 times/hour, with 4 s feeding/time, at approximately 2% of body weight per day.
Enzyme-linked immunosorbent assay. Tumor necrosis factor-α (TNF-α) (CSB-E13254Fh), fish growth hormone (GH) (CSB-E12121Fh), fish heat shock protein-70 (HSP-70) (CSB-E16327Fh), and interleukin-6 (IL-6) (CSB-E13258Fh) levels were determined in samples of the muscle tissue homogenates via enzyme-linked immunosorbent assay (ELISA), using commercially available kits for fish from CUSABIO (Wuhan, China, http://www.cusabio.com). Samples were taken five times, from September to January. Each month, three pooled samples (n = 5 fish in one pooled sample) from each experimental group were collected.
Collection, storage, and analysis of samples were carried out according to CUSABIO manufacturer’s instructions (see above). Supernatants were stored at –80°C for further analyses. Concentrations were measured at 450 nm on a Thermo Labsystems Multiskan Ascent 354 Microplate Reader (Thermo Labsystems Inc., USA).
Evaluation of growth performance. The weight and length of 50 randomly selected fish from each group were measured individually every 30 days, throughout the treatment period (
Growth performance indices including weight gain (WG) [g], coefficient of variation (CV) [%], size heterogeneity of the weight (SH), specific growth rate (SGR) [%day–1], and Fulton’s condition factor (K) were calculated using the following formulae inspired by
W G = WF − WI
CV = (SDIW × WIW–1) × 100
SH = CVF × CVI–1
SGR = (Ln WF − Ln WI) × t–1 × 100
K = (W × 105) × L–3
where WF is the mean final weight of fish [g], WI is the mean initial weight [g], SDIW is the standard deviation of individual weight, WIW is the mean individual weight [g], CVF is the final coefficient of variation, CVI is the initial coefficient of variation, t is the time of rearing [day], W is the weight, and L is the mean total length.
Statistical analysis. The results were analyzed statistically by the R (version 3.6.2) environment with RStudio software (
Mean TNF-α, IL-6, GH, and HSP-70 concentrations are presented in Table
Expression of TNF-α, IL-6, GH, and HSP-70 in juveniles of sea trout, Salmo trutta, subjected to long-term dietary administration of β-glucan and BGN-2 in experimental groups (treatments).
Group | Parameter | |||
---|---|---|---|---|
TNF-α [(pg/mL)/10] | IL-6 [pg/mL] | GH [pg/mL] | HSP-70 [pg/mL] | |
D1 | 740.67 ± 44.37a | 172.83 ± 0.91b | 3371.42 ± 16.88c | 239.01 ± 1.17c |
D2 | 959.83 ± 70.68a | 176.36 ± 1.16ab | 3436.01 ± 36.50bc | 267.84 ± 5.82ab |
D3 | 872.75 ± 42.28a | 178.28 ± 0.89ab | 3479.20 ± 14.72abc | 263.96 ± 4.63abc |
D4 | 870.69 ± 35.00a | 179.07 ± 0.77a | 3674.71 ± 60.07ab | 291.62 ± 6.06a |
D5 | 871.43 ± 33.59a | 180.81 ± 0.71a | 3722.67 ± 62.40a | 257.82 ± 2.53bc |
Our findings indicate that the mean IL-6 concentration was significantly higher in the D4 and D5 groups compared to D1, with no significant difference observed when compared to D2 and D3. The lowest mean IL-6 levels were observed in D1 (Table
A significantly higher GH concentration was observed in D5 (Fig.
TNF-α, IL-6, and GH levels did not vary significantly in the D2 and D3 groups during the whole trial of this study compared to the control.
In September, there was no significant difference in HSP-70 between the trial groups and the control. From October to the end of the study, the concentration of HSP-70 in the D4 group was the highest, it was significantly higher than the control group. Also, a high concentration was observed in the D2 group (Fig.
Growth performance, mean final weight, weight gain, coefficient of variation, size heterogeneity, specific growth rate, and Fulton’s condition factor are presented in Table
Growth performance of juveniles of sea trout, Salmo trutta, subjected to long-term dietary administration of β-glucan and BGN-2 in experimental groups (treatments) by the end of trial.
Group | Parameter | |||||||
---|---|---|---|---|---|---|---|---|
W I | W F | L F | W G | CV | SH | SGR | K | |
D1 | 2.50 ± 0.21a | 11.84 ± 0.78c | 9.33 ± 0.24a | 9.34 | 28.76 | 0.90 | 0.73 | 1.60 ± 0.11a |
D2 | 2.59 ± 0.17a | 13.31 ± 0.84bc | 9.50 ± 0.24a | 10.72 | 27.50 | 1.10 | 0.77 | 1.58 ± 0.10a |
D3 | 2.67 ± 0.21a | 16.18 ± 0.77abc | 11.12 ± 0.32ab | 13.51 | 20.79 | 0.69 | 0.85 | 1.18 ± 0.12ab |
D4 | 2.67 ± 0.17a | 16.76 ± 0.64ab | 11.91 ± 0.28b | 14.09 | 16.55 | 0.66 | 0.86 | 0.99 ± 0.04b |
D5 | 2.76 ± 0.16a | 19.88 ± 0.74a | 12.81 ± 0.26b | 17.12 | 16.15 | 0.73 | 0.93 | 0.94 ± 0.04b |
Initially, there were no statistically significant differences in the initial mean weight among the diet groups. However, at the end of the study, it was observed that the final mean weight in groups D4 and D5 was significantly higher compared to the control group (D1). Consequently, the weight gain of fish from groups D4 and D5 was higher.
The same was observed for the specific growth rates (SGR) where fish from D4 and D5 displayed an increased SGR compared to other diet groups. The final mean length in D4 and D5 was significantly higher compared to D1 and D2.
Fulton’s condition factor (K) was significantly affected by dietary treatment. The lowest K was recorded in D4 and D5, while the highest value was observed in D1 and D2. The results obtained at the end of the study showed that fish fed the D2 and D3 did not show a statistically significant difference (P < 0.05) from fish fed the control diet. The results showed that the growth performance of sea trout in the control group was at the lowest level and showed a significant difference (P < 0.05) with the D4 and D5 diet groups.
Results of other aspects of these experiments have been reported in previous papers (
The use of natural immunostimulants offers important advantages for aquaculture. These substances enhance the immune response and strengthen disease resistance, contributing to improved health and overall well-being in aquatic organisms. Furthermore, these products are environmentally friendly, biodegradable, and safe for humans (
In teleost fishes, such as Atlantic salmon, Salmo salar and rainbow trout, Oncorhynchus mykiss (Walbaum, 1792), the administration of β-glucans enhances the regenerative capacity of immunosuppressed cells, thereby bolstering their ability to combat infectious diseases (
While the exact mechanism by which β-glucan influences the fish’s immune system remains uncertain, there is substantial evidence indicating its ability to boost phagocytic activity and increase the expression of cytokines in different types of immune cells, including macrophages, dendritic cells, and neutrophils (Di
The results of our studies are consistent with other authors (
The inclusion of β-glucan at a dose of 1 g kg–1 (D2) in the diet leads to a significant rise in TNF-α levels during the months of September and October. In subsequent months, the expression of TNF-α in all groups remains without significant statistical difference. Similar results were obtained by other researchers (
GH plays a crucial role in regulating various physiological functions of fish such as development, growth, osmoregulation, immune systems, reproduction function, etc. (
More authors demonstrated that dietary β-glucan has played a major role in regulating stress- and immune-related factors through expressing HSP-70 (
This study highlights the beneficial outcomes resulting from the oral administration of the BGN- 2 product at doses of 6 g kg–1 and 14 g kg–1 on the growth performance of sea trout. These findings align with previous research (
This study adds to the growing body of evidence supporting the efficacy of β-glucan, specifically the BGN-2 product, in promoting the growth of sea trout. The results suggest that the administration of β-glucan products at the specified doses can positively influence the growth parameters of sea trout, contributing to the development of more efficient and sustainable aquaculture practices.
In summary, all of our research (
The presently reported study was supported financially by the research project “Strengthening the Scientific Capacity of LLU” No. Z-27 – The application of β-glucan to ensure sea trout health. The authors would like to express their gratitude to the Institute of Food Safety, Animal Health, and Environment “BIOR” for the support in carrying out this study. This manuscript is supported by the Institute of Food Safety, Animal Health, and Environment BIOR.