Yanjiao Zhang

Effects of potential probiotic Bacillus subtilis T13 on growth, immunity and disease resistance against Vibrio splendidus infection in juvenile sea cucumber Apostichopus japonicus

A feeding experiment was conducted to determine influences of potential probiotic Bacillus subtilis T13 (isolated from intestine of healthy sea cucumbers) on growth, immunity and disease resistance against Vibrio splendidus infection in juvenile sea cucumbers Apostichopus japonicus. Animals were fed with diets containing B. subtilis T13 at 0, 10(5), 10(7) and 10(9) CFU/g for 30 days, respectively. At the end of the growth trial, fifteen sea cucumbers from each aquarium were sampled for immune indices measurement. Then twenty sea cucumbers from each replicate were challenged with V. splendidus. Results showed that administration of B. subtilis T13 had significant effect on the specific growth rates (SGR) of sea cucumbers (P < 0.05). Phagocytosis, respiratory burst activity and total nitric oxide synthase (T-NOS) activity were significantly improved in coelomocytes of sea cucumbers fed with T13 at 10(9) CFU/g diet (P < 0.05). The highest values of the total coelomocytes counts (TCC) and superoxide dismutase (SOD) activity were found in sea cucumbers fed diet containing T13 at 10(9) CFU/g. The cumulative mortality after V. splendidus challenge decreased significantly in sea cucumbers fed with T13 at dose of 10(9) CFU/g (P < 0.05). The present study confirmed the potential beneficial effects of B. subtilis T13 as dietary probiotic in juvenile A. japonicus.

Regulation of tissue LC-PUFA contents, Δ6 fatty acyl desaturase (FADS2) gene expression and the methylation of the putative FADS2 gene promoter by different dietary fatty acid profiles in Japanese seabass (Lateolabrax japonicus).

The present study was conducted to evaluate the influences of different dietary fatty acid profiles on the tissue content and biosynthesis of LC-PUFA in a euryhaline species Japanese seabass reared in seawater. Six diets were prepared, each with a characteristic fatty acid: Diet PA: Palmitic acid (C16:0); Diet SA: Stearic acid (C18:0); Diet OA: Oleic acid (C18:1n-9); Diet LNA: α-linolenic acid (C18:3n-3); Diet N-3 LC-PUFA: n-3 LC-PUFA (DHA+EPA); Diet FO: the fish oil control. A 10-week feeding trial was conducted using juvenile fish (29.53±0.86 g). The results showed that Japanese seabass had limited capacity to synthesize LC-PUFA and fish fed PA, SA, OA and LNA showed significantly lower tissue n-3 LC-PUFA contents compared to fish fed N-3 LC-PUFA and FO. The putative gene promoter and full-length cDNA of FADS2 was cloned and characterized. The protein sequence was confirmed to be homologous to FADS2s of marine teleosts and possessed all the characteristic features of microsomal fatty acid desaturases. The FADS2 transcript levels in liver of fish fed N-3 LC-PUFA and FO were significantly lower than those in fish fed other diets except LNA while Diet PA significantly up-regulated the FADS2 gene expression compared to Diet LNA, N-3 LC-PUFA and FO. Inversely, fish fed N-3 LC-PUFA and FO showed significantly higher promoter methylation rates of FADS2 gene compared to fish fed the LC-PUFA deficient diets. These results suggested that Japanese seabass had low LC-PUFA synthesis capacity and LC-PUFA deficient diets caused significantly reduced tissue n-3 LC-PUFA contents. The liver gene expression of FADS2 was up-regulated in groups enriched in C16:0, C18:0 and C18:1n-9 respectively but not in the group enriched in C18:3n-3 compared to groups with high n-3 LC-PUFA contents. The FADS2 gene expression regulated by dietary fatty acids was significantly negatively correlated with the methylation rate of putative FADS2 gene promoter.

Cloning and characterization of SREBP-1 and PPAR-α in Japanese seabass Lateolabrax japonicus, and their gene expressions in response to different dietary fatty acid profiles

In the present study, putative cDNA of sterol regulatory element-binding protein 1 (SREBP-1) and peroxisome proliferator-activated receptor α (PPAR-α), key regulators of lipid homoeostasis, were cloned and characterized from liver of Japanese seabass (Lateolabrax japonicus), and their expression in response to diets enriched with fish oil (FO) or fatty acids such as palmitic acid (PA), stearic acid (SA), oleic acid (OA), α-linolenic acid (ALA), and n-3 long-chain polyunsaturated fatty acid (n-3 LC-PUFA), was investigated following feeding. The SREBP-1 of Japanese seabass appeared to be equivalent to SREBP-1a of mammals in terms of sequence feature and tissue expression pattern. The stimulation of the mRNA expression level of SREBP-1 in liver of Japanese seabass by dietary fatty acids significantly ranked as follows: PA, OA>SA, ALA, and n-3 LC-PUFA>FO. A new PPAR-α subtype in Japanese seabass, PPAR-α2, was cloned in this study, which is not on the same branch with Japanese seabass PPAR-α1 and mammalian PPAR-α in the phylogenetic tree. Liver gene expression of PPAR-α1 of Japanese seabass was inhibited by diets enriched with ALA or FO compared to diets enriched with PA or OA, while the gene expression of PPAR-α2 of Japanese seabass was up-regulated by diets enriched with ALA or n-3 LC-PUFA compared to diets enriched with OA or FO. This was the first evidence for the great divergence in response to dietary fatty acids between PPAR-α1 and PPAR-α2 of fish, which indicated probable functional distribution between PPAR-α isotypes of fish.

Effects of dietary nucleotides on growth, non-specific immune response and disease resistance of sea cucumber Apostichopus japonicas.

A 9-week feeding trial was conducted to investigate the effects of dietary nucleotides (NT) on growth, immune response and disease resistance of sea cucumber Apostichopus japonicas (initial weight: 5.87 ± 0.03 g). Four graded levels of dietary NT were designed as 0, 150, 375 and 700 mg/kg, respectively. After the feeding trial, sea cucumbers were challenged with Vibrio splendidus for the determination of disease resistance. The results showed that the specific growth rates were significantly higher in sea cucumber fed the diet with 375 mg/kg NT than those fed the basal diet without NT supplementation (P < 0.05). The highest total coelomocytes counts in coelomic fluid were found in the treatment with 150 mg/kg of dietary NT (P < 0.05). Compared to those fed with the basal diet, sea cucumber fed diets with nucleotides (≥ 375 mg/kg) had significantly higher phagocytic activities in coelomic fluid (P < 0.05). Respiratory burst activities in coelomic fluid significantly increased with increasing dietary NT supplementations up to 700 mg/kg (P < 0.05). No significant differences in the activities of superoxide dismutase, total nitric oxide synthase and acid phosphatase in coelomic fluid were found among all the treatments (P > 0.05). After being challenged with V. splendidus, the cumulative mortalities of sea cucumber fed diets with 150 and 375 mg/kg NT were significantly lower than that in the treatment without dietary nucleotide supplementation (P < 0.05). Under the experimental conditions, the present results confirmed that a diet supplemented with 375 mg/kg NT is able to enhance both non-specific immune response and growth of sea cucumber in vivo. In conclusion, it was showed that dietary NT does increase the growth performance, non-specific immunity and disease resistance of sea cucumber. The optimum dietary NT supplementation level for sea cucumber was found to be 375 mg/kg. The application of dietary NT may present a novel strategy for health management in sea cucumbers aquaculture.

Effects of dietary stachyose on growth performance, digestive enzyme activities and intestinal morphology of juvenile turbot (Scophthalmus maximus L)

A 12-week feeding trial was conducted to evaluate the effects of dietary stachyose on the growth performance, digestive enzymes activities and intestinal structures of juvenile turbot (Scophthalmus maximus L). Five isonitrogenous (49.58% crude protein) and isolipidic (10.50% crude lipid) diets were formulated to contain 0 (Control), 0.625% (S-0.625), 1.25% (S-1.25), 2.5% (S-2.5) and 5% (S-5) stachyose, respectively. With the increase of stachyose level, the growth performance and feed utilization of turbot, such as the specific growth rate, final mean body weight, weight gain rate and feed efficiency, increased significantly (P< 0.05) and then stabilized. The feed intake of fish fed S-5 was significantly higher (P< 0.05) than that of fish in other groups. The activities of trypsin, intestinal caseinolytic, stomach and intestinal amylase were significantly influenced by stachyose (P<0.05). The highest values of trypsin and intestinal caseinolytic activities were observed in group S-1.25, while the highest activity of stomach amylase and the lowest activity of intestine amylase were observed in group S-5. No lesion or damage was found on the distal intestine structures of fish from all treatments, while the height of simple folds in the distal intestine was significantly increased (P< 0.05) when 1.25% or 2.5% stachyose was added in the diets. These results indicated that moderate level of stachyose (1.25%) improves the growth performance, feed utilization, digestive enzyme activities and the distal intestine structures of juvenile turbot.

Effects of dietary corn gluten meal on growth performance and protein metabolism in relation to IGF-I and TOR gene expression of juvenile cobia (Rachycentron canadum)

A growth experiment was conducted on cobia (Rachycentron canadum, initial weight 108.2 g ± 3.0 g) to investigate the effects of dietary corn gluten meal (CGM) levels on the fish growth, whole body composition and protein metabolism in relation to specific gene expression. Five isonitrogenous (crude protein 45%) and isoenergetic (gross energy 20 kJ g−1) practical diets were formulated by replacing 0% (the control), 17.5%, 35.0%, 52.5%, and 70.0% of fish meal (FM) protein with CGM protein. No significant differences were observed in the survival, feed intake (FI), specific growth rate (SGR), feed efficiency (FE) and protein productive value (PPV) among fish fed diets with 0%, 17.5%, 35.0%, and 52.5% of CGM protein. However, these indices were significantly lower in fish fed the diet with 70.0% of CGM protein than those in fish fed the control diet (P < 0.05). The whole-body crude protein and lipid contents were significantly lower while the whole-body moisture content was significantly higher in fish fed the diet with 70.0% of CGM protein compared with the control group (P < 0.05). When 70.0% of FM protein was replaced by CGM, plasma total protein and cholesterol contents were significantly lower than those in the control group (P < 0.05). Fish fed the diet with 70.0% of CGM protein had significantly lower hepatic insulin-like growth factor I (IGF-I) expression levels than those in the control group (P < 0.05). However, no significant differences were observed in hepatic target of rapamycin (TOR), dorsal muscle IGF-I and TOR expression levels among dietary treatments. Results of the present study indicated that 52.5% of FM protein could be replaced by CGM in the diets without significant influences on the growth, feed utilization and protein metabolism of juvenile cobia. The present results might be useful for developing cost effective and sustainable cobia dietary formulations.

Effects of dietary soy isoflavones on feed intake, growth performance and digestibility in juvenile Japanese flounder (Paralichthys olivaceus)

An 8-week feeding trial was conducted to investigate the effects of dietary soy isoflavones on feeding intake, growth performance, and digestion of juvenile Japanese flounder (Paralichthys olivaceus). Four isonitrogenous (49% crude protein) and isoenergetic (20.1 MJ kg−1) diets were formulated to contain four graded levels of soy isoflavones, namely, 0, 1, 4 and 8 g soy isoflavones in 1 kg of diet. Each diet was randomly fed to triplicate tanks of fish (Initial average weight: 2.58 g ± 0.01 g), and each tank was stocked with 35 fish. No significant difference was observed among diets with levels of 0, 1 and 4 g kg−1 soy isoflavones in feed intake, weight gain, feed efficiency ratio (FER), proximate composition of fish whole body and apparent digestibility coefficients (ADC) of nutrients and energy (P>0.05). However, high dietary soy isoflavones level (8 g kg−1) significantly depressed weight gain, FER, whole-body crude lipid content of fish and ADC of nutrients (P<0.05). These results indicate that high level of dietary soy isoflavones (above 4 g kg−1) significantly depresses growth responses and FER of Japanese flounder. However, as the content of soy isoflavones in soybean meal is around 1 to 3 g kg−1, the adverse effects might be neglected when soybean products are used as a fish feed ingredient.

Wnt/β-catenin signaling participates in the regulation of lipogenesis in the liver of juvenile turbot (Scophthalmus maximus L.).

In this study, the mechanism that Wnt/β-catenin signaling inhibits lipogenesis was investigated in the liver of juvenile turbot (Scophthalmus maximus L.) by LiCl or XAV939 treatment. Wnt/β-catenin signaling was activated by LiCl treatment or inhibited by XAV939 treatment through regulating the expression of glycogen synthase kinase-3β (GSK-3β) and Wnt10b. In addition, the expression of lipoprotein lipase (LPL), fatty acid synthetase (FAS), peroxisome proliferator-activated receptor γ (PPARγ), and CCAAT/enhancer binding protein α (C/EBPα) was inhibited by LiCl treatment, but induced by XAV939 treatment. In the plasma of juvenile turbot, the level of nonesterified fatty acid (NEFA), glycerol, triglyceride (TG), total cholesterol (TC), and low density lipoprotein cholesterol (LDL-C) was decreased by LiCl treatment, which was related to the decrease of the activity of LPL and FAS. Thus the inhibitory effect of Wnt/β-catenin signaling on lipogenesis was associated with the expression of key enzymes and transcriptional factors. Wnt/β-catenin signaling may participate in inhibiting lipogenesis by inhibiting the expression of PPARγ and C/EBPα in the liver of juvenile turbot.

Tumour necrosis factor-α inhibits hepatic lipid deposition through GSK-3β/β-catenin signaling in juvenile turbot (Scophthalmus maximus L.)

In this study, the mechanism that TNFα inhibits lipid deposition through GSK-3β/β-catenin signaling was investigated in the liver of juvenile turbot (Scophthalmus maximus L.) by injection of TNFα or TNFα inhibitor pomalidomide (POM). It was found that TNFα inhibited the expression of GSK-3β and induced β-catenin expression. TNFα inhibited the expression of peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer binding protein α (C/EBPα), as well as the activity of lipoprotein lipase (LPL) and fatty acid synthetase (FAS). In addition, the level of triglyceride (TG), total cholesterol (TC), nonesterified fatty acid (NEFA), and glycerol was decreased by TNFα treatment in the liver. In the plasma, the level of TG, TC, low density lipoprotein cholesterol (LDL-C), NEFA, and glycerol was decreased, but high density lipoprotein cholesterol (HDL-C) was increased by TNFα treatment. However, compared to TNFα, POM had the opposite effect on the biochemical indexes and genes related to lipid deposition in the liver. The results indicated that TNFα may regulate hepatic lipid metabolism and fat distribution through GSK-3β/β-catenin signaling as well as transcription factors PPARγ and C/EBPα in juvenile turbot.

GSK-3b participates in the regulation of hepatic lipid deposition in large yellow croaker (Larmichthys crocea).

In this study, the participation of glycogen synthase kinase-3β (GSK-3β) in the lipid deposition was investigated in the liver of large yellow croaker (Larmichthys crocea) by LiCl treatment. It was found that the expression of GSK-3β and peroxisome proliferator-activated receptor-γ (PPARγ) was inhibited, but the expression of β-catenin was induced by LiCl treatment. Furthermore, the gene expression and activity of fatty acid synthetase (FAS) and lipoprotein lipase (LPL) in the liver was inhibited by LiCl treatment. The content of total cholesterol (TC), triglyceride (TG), and non-estesterified fatty acid in the liver, as well as TC, TG, and low-density lipoprotein cholesterol in plasma, was decreased by LiCl treatment. However, high-density lipoprotein cholesterol in plasma was increased, and the number of lipid droplets in the liver was decreased by LiCl treatment. The results indicate that GSK-3β/β-catenin may participate in regulating LPL and FAS through PPARγ in the liver of large yellow croaker, which will lead to the inhibition of hepatic lipid deposition.