The effect of dietary folic acid on biochemical parameters and gene expression of three heat shock proteins (HSPs) of blunt snout bream (<Emphasis Type="Italic">Megalobrama amblycephala)</Emphasis> fingerling under acute high temperature stress

The effects of dietary folic acid on biochemical parameters and gene expression of three heat shock proteins (HSPs) of blunt snout bream (Megalobrama amblycephala) fingerling under acute high temperature stress. Six dietary folic acid groups (0.0, 0.5, 1.0, 2.0, 5.0, and 10.0) mg/kg diets were designed and assigned into 18 tanks in three replicates each (300 l/tank) and were administered for 10 weeks in a re-circulated water system. The fingerlings with an initial weight of 27.0 ± 0.03 g were fed with their respective diets four times daily. At the end of the experiment, samples were collected before challenge, 0, 24, 72 h, and 7 days. Serum total protein (TP), alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), cortisol, glucose, complement C3 (C3), complement C4 (C4, immunoglobulin M (IgM) hepatic superoxide dismutase (SOD), catalase (CAT), malondialdehyde (MDA), and the expression of heat shock protein 60 (HSP60), 70 (HSP70), and 90 (HSP90) were studied. The results showed that fish fed with dietary folic acid between 1.0, 2.0, and 5.0 mg/kg significantly (P < 0.05) increased serum TP, C3, C4 hepatic SOD, CAT, and the expression of HSP60, HSP70, and HSP90 before and after temperature challenge of 32 °C. Also, serum ALP, cortisol, glucose, and hepatic MDA were significantly (P < 0.05) reduced by supplementation of dietary folic acid level 1.0, 2.0, and 5.0 mg/kg before and after the same temperature challenge of 32 °C. Before stress, 0, 24, 72 h, and 7 days significantly (P < 0.05) affects serum biochemical parameters, immune and antioxidant capacities, and expression level of three HSPs. Furthermore, there was no statistical evidence to show that dietary folic acid inclusion level and temperature duration have significant interactive effect on serum biochemical parameters, antioxidant parameters, and gene expression level (P > 0.05) of the three HSPs. However, there were statistical significant interactive effect between dietary folic acid inclusion level and temperature duration on serum C3 and C4 (P < 0.05) except IgM (P > 0.05). The present results indicate that supplementation of basal diet from 1.0 mg/kg; 2.0 and 5.0 mg/kg can enhance acute high temperature resistance ability in M. amblycephala fingerling to some degree and improve physiological response, immune and antioxidant capacities, and expression level of three HSPs.     

Effect of dietary betaine on growth performance,antioxidant capacity and lipid metabolism in blunt snout bream fed a high-fat diet

An 8-week feeding experiment was conducted to determine the effect of dietary betaine levels on the growth performance, antioxidant capacity, and lipid metabolism in high-fat diet-fed blunt snout bream (Megalobrama amblycephala) with initial body weight 4.3 ± 0.1 g [mean ± SEM]. Five practical diets were formulated to contain normal-fat diet (NFD), high-fat diet (HFD), and high-fat diet with betaine addition (HFB) at difference levels (0.6, 1.2, 1.8%), respectively. The results showed that the highest final body weight (FBW), weight gain ratio (WGR), specific growth rate (SGR), condition factor (CF), and feed intake (FI) (P < 0.05) were obtained in fish fed 1.2% betaine supplementation, whereas feed conversion ratio (FCR) was significantly lower in the same group compared to others. Hepatosomatic index (HSI) and abdominal fat rate (AFR) were significantly high in fat group compared to the lowest in NDF and 1.2% betaine supplementation, while VSI and survival rate (SR) were not affected by dietary betaine supplementation. Significantly higher (P < 0.05), plasma total cholesterol (TC), triglycerides (TG), low-density lipoprotein (LDL), aspartate transaminase (AST), alanine transaminase (ALT), cortisol, and lower high-density lipoprotein (HDL) content were observed in HFD but were improved when supplemented with 1.2% betaine. In addition, increase in superoxide dismutase (SOD), catalase (CAT), and reduced glutathione (GSH) in 1.2% betaine inclusion could reverse the increasing malondialdehyde (MDA) level induced by HFD. Based on the second-order polynomial analysis, the optimum growth of blunt snout bream was observed in fish fed HFD supplemented with 1.2% betaine. HFD upregulated fatty acid synthase messenger RNA (mRNA) expression and downregulated carnitine palmitoyltransferase 1, peroxisome proliferator-activated receptor α, and microsomal triglyceride transfer protein mRNA expression; nevertheless, 1.2% betaine supplementation significantly reversed these HFD-induced effects, implying suppression of fatty acid synthesis, β-oxidation, and lipid transport. This present study indicated that inclusion of betaine (1.2%) can significantly improve growth performance and antioxidant defenses, as well as reduce fatty acid synthesis and enhance mitochondrial β-oxidation and lipid transportation in high-fat diet-fed blunt snout bream, thus effectively alleviating fat accumulation in the liver by changing lipid metabolism.     

Effects of the supplementation of vitamin D<Subscript>3</Subscript> on the growth and vitamin D metabolites in juvenile Siberian sturgeon (<Emphasis Type="Italic">Acipenser baerii</Emphasis>)

The objective of this study was to evaluate the effects of the supplementation of vitamin D₃ on the growth, vitamin D metabolites, and osteocalcin secretion in juvenile Siberian sturgeon (Acipenser baerii). A 90-day growth trial was conducted with juvenile Siberian sturgeon (initial body weight 3.47 ± 0.14 g) fed seven isonitrogenous and isoenergetic practical diets (45% CP and 13% lipid) containing 60 (basal diet), 240, 450, 880, 1670, 3300, or 1.0 × 10⁵ IU/kg feed (D60~D 1.0 × 10⁵) vitamin D₃. The results showed that weight gain and specific growth rate increased as the dietary vitamin D₃ levels increased from 450 to 3300 IU/kg (P < 0.05). The fish fed with D1670 and D3300 diets had higher crude lipid and ash levels than the fish fed the D60 diet (P < 0.05). The fish fed D880, D1670, or D3300 diets had higher 25-OH-D₃ and 1,25-(OH)₂-D₃ levels than the fish fed the D60 diet (P < 0.05). The fish fed D880, D1670, D3300, or D1.0 × 10⁵ diets had higher osteocalcin levels than the fish fed the D60 diet (P < 0.05). Based on the broken line method analysis of weight gain and osteocalcin, the dietary vitamin D₃ requirement of juvenile Siberian sturgeon was estimated to be 1683.30 and 1403.27 IU/kg per diet, respectively.     

Replacement of fish oil with soybean oil in diets for juvenile Chinese sucker (<Emphasis Type="Italic">Myxocyprinus asiaticus</Emphasis>): effects on liver lipid peroxidation and biochemical composition

This study was designed to evaluate the effect of the replacement of fish oil (FO) by soybean oil (SO) on growth performance, liver lipid peroxidation, and biochemical composition in juvenile Chinese sucker, Myxocyprinus asiaticus. Fish (13.7 ± 0.2 g) in triplicate were fed five experimental diets in which 0% (FO as control), 40% (SO40), 60% (SO60), 80% (SO40), and 100% (SO100) FO were replaced by SO. The body weight gain of fish fed SO40, SO60, or SO80 diet was similar to FO group, but diets that have 100% soybean oil as dietary lipid significantly reduced fish growth (P < 0.05). Although the level of SO resulted in increasing crude lipid content of the liver, the level of SO did not significantly alter the hepatosomatic index (HSI). Indicators of peroxidation, such as vitamin E (V_E) and thiobarbituric acid-reactive substance (TBARS) contents, were changed as increasing dietary SO. It was shown that the inclusion of SO in the diets increased V_E concentrations, but reduced TBARS in the liver and total cholesterol (T-CHO) in the plasma. Linoleic acid (LA) and linolenic acid (LNA) significantly increased in fish liver fed diets that contained SO, but eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), and the ratio n-3/n-6 were significantly reduced by the inclusion of dietary SO (P < 0.05). Our results indicated that the inclusion of SO increased the hepatic V_E content and reduced lipid peroxidation in fish. However, diet containing 100% SO as dietary lipid could reduce growth performance. Thus, we recommended that 40–80% SO can be used as dietary lipid to replace FO for juvenile Chinese sucker.     

Effects of starvation and feeding on blood chemistry,fatty acid composition and expression of vitellogenin and fatty acid-binding protein genes in female swimming crab <Emphasis Type="Italic">Portunus trituberculatus</Emphasis> broodstock

Portunus trituberculatus broodstock were stocked in plastic tanks to evaluate the effects of starvation and feeding on gonadal development, blood chemistry, fatty acid composition, and expression of vitellogenin (Vtg) and fatty acid-binding protein genes (FABP) in females. Two treatments (starved and fed) were randomly assigned to triplicate groups of 90 swimming crab broodstock (approximately 230 ± 45 g). In the starved treatment, crabs were starved for 30 days, whereas in the fed treatment crabs were fed once a day with clams. The gonadosomatic index decreased significantly in starved crabs (P < 0.05), as did the serum glucose and cholesterol concentrations; conversely, the total protein concentration in serum significantly increased (P < 0.05). In the ovary, there was a significant relative decline of 18:0, 16:1n-7 and 20:1n-9 fatty acids and relative increases of 20:4n-6, 22:6n-3, 18:1n-9 and 20:5n-3 in starved crabs compared to fed crabs (P < 0.05). Relative expression of Vtg in the ovary decreased significantly in starved crabs (P < 0.05), while there was no significant difference in hepatopancreas Vtg expression between starved and fed crabs (P > 0.05). Starvation suppressed gonadal development in female swimming crab broodstock.     

Dietary supplementation of curcumin augments heat stress tolerance through upregulation of <Emphasis Type="Italic">nrf-2</Emphasis>-mediated antioxidative enzymes and <Emphasis Type="Italic">hsps</Emphasis> in <Emphasis Type="Italic">Puntius sophore</Emphasis>

Heat stress is one of the major environmental concerns in global warming regime and rising temperature has resulted in mass mortalities of animals including fishes. Therefore, strategies for high temperature stress tolerance and ameliorating the effects of heat stress are being looked for. In an earlier study, we reported that Nrf-2 (nuclear factor E2-related factor 2) mediated upregulation of antioxidative enzymes and heat shock proteins (Hsps) provide survivability to fish under heat stress. In this study, we have evaluated the ameliorative potential of dietary curcumin, a potential Nrf-2 inducer in heat stressed cyprinid Puntius sophore. Fishes were fed with diet supplemented with 0.5, 1.0, and 1.5% curcumin at the rate 2% of body weight daily in three separate groups (n = 40 in each group) for 60 days. Fishes fed with basal diet (without curcumin) served as the control (n = 40). Critical thermal maxima (CTmax) was determined for all the groups (n = 10, in duplicates) after the feeding trial. Significant increase in the CTmax was observed in the group fed with 1.5% curcumin- supplemented fishes whereas it remained similar in groups fed with 0.5%, and 1% curcumin-supplemented diet, as compared to control. To understand the molecular mechanism of elevated thermotolerance in the 1.5% curcumin supplemented group, fishes were given a sub-lethal heat shock treatment (36 °C) for 6 h and expression analysis of nrf-2, keap-1, sod, catalase, gpx, and hsp27, hsp60, hsp70, hsp90, and hsp110 was carried out using RT-PCR. In the gill, expression of nrf-2, sod, catalase, gpx, and hsp60, hsp70, hsp90, and hsp110 was found to be elevated in the 1.5% curcumin-fed heat-shocked group compared to control and the basal diet-fed, heat-shocked fishes. Similarly, in the liver, upregulation in expression of nrf-2, sod, catalase, and hsp70 and hsp110 was observed in 1.5% curcumin supplemented and heat shocked group. Thus, this study showed that supplementation of curcumin augments tolerance to high temperature stress in P. sophore that could be attributed to nrf-2-induced upregulation of antioxidative enzymes sod, catalase, gpx, and the hsps.     

Resveratrol supplementation improves lipid and glucose metabolism in high-fat diet-fed blunt snout bream

Here, we aimed to investigate whether resveratrol (RSV) can ameliorate high-fat diet (HFD)-induced metabolic disorder in fish. Blunt snout bream (Megalobrama amblycephala) with average weight 27.99 ± 0.56 g were fed a normal fat diet (NFD, 5% fat, w/w), a HFD (11% fat), or a HFD supplemented with 0.04, 0.36, or 1.08% RSV for 10 weeks. As expected, fish fed a HFD developed hepatic steatosis, as shown by elevated hepatic and plasma triglycerides, raised whole body fat, intraperitoneal fat ratio and hepatosomatic index, and increased plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST). RSV supplementation lessened increases in body mass, whole body fat, and intraperitoneal fat, and alleviated development of hepatic steatosis, elevations of plasma triglyceride and glucose, and abnormalities of ALT and AST in HFD-fed fish. RSV supplementation increased SIRT1 messenger RNA (mRNA) expression and consequently hepatic mRNA expression of adipose triglyceride lipase (ATGL), carnitine palmitoyltransferase (CPT1a), and microsomal triglyceride transfer protein (MTTP), implying upregulation of lipolysis, β-oxidation, and lipid transport, respectively, in the liver. Conversely, hepatic lipoprotein lipase (LPL), sterol regulatory element-binding protein 1 (SREBP-1c), peroxisome proliferator-activated receptor γ (PPARγ), and ATP citrate lyase (ACLY) mRNA expression were decreased, implying suppression of fatty acid uptake, lipogenesis, and fatty acid synthesis. Additionally, RSV downregulated glucokinase (GCK) and sodium-dependent glucose cotransporter 1 (SGLT1) and upregulated glucose transporter 2 (GLUT2) mRNA expression, thus restoring normal glucose fluxes. Thus, RSV improves lipid and glucose metabolisms in blunt snout bream, which are potentially mediated by activation of SIRT1.     

Effects of dietary <Emphasis Type="Italic">myo</Emphasis>-inositol on growth,chemical composition and plasma chemistry of Amur sturgeon <Emphasis Type="Italic">Acipenser schrenckii</Emphasis>

The present study was conducted to demonstrate the dietary myo-inositol requirement and its effects on the growth, proximate composition and blood chemistry of Amur sturgeon (Acipenser schrenckii). Triplicate groups of 30 fish (initial weight 11.90?±?0.12 g) were fed different diets containing graded levels of myo-inositol (28.75, 127.83, 343.83, 565.81, 738.15 and 936.28 mg kg^?1) until satiation for 56 days. The fish were weighed after a 24-h fast, and six fish were used for whole body composition analysis. Further, the liver and muscle were sampled from another six fish for lipid analysis. The blood and liver were sampled from the remaining six fish for haematology and fatty acid analysis. The weight gain of fish increased with myo-inositol content, from the 28.75- to 343.83-mg kg^?1 myo-inositol treatment groups, and then stabilised. The liver lipid content and hepatosomatic index decreased significantly from 21.91 to 19.14% and from 3.20 to 2.76% with increased dietary myo-inositol supplementation, respectively. The whole body lipid content generally decreased from 6.33 to 5.55%. The content of liver-saturated fatty acids decreased significantly (28.13%) in the 936.28-mg kg^?1 treatment group. The content of plasma non-esterified fatty acids increased with the increase in dietary myo-inositol supplementation from 0.77 to 1.17 mmol L^?1, whereas the content of triglycerides significantly decreased from 4.62 to 3.28 mmol L^?1. In conclusion, the optimum myo-inositol requirement was found to be 336.1 mg kg^?1, based on weight gain in a two-slope quadratic broken-line model.     

Dietary substitution of fishmeal by alternative protein with guanosine monophosphate supplementation influences growth,digestibility, blood chemistry profile,immunity, and stress resistance of red sea bream, <Emphasis Type="Italic">Pagrus major</Emphasis>

We determined the effects of complete fishmeal (FM) replacement by alternative protein (soy protein concentrate, SPC) with guanosine monophosphate (GMP) supplementation on growth, digestibility, immunity, blood chemistry profile, and stress resistance of juvenile red sea bream, Pagrus major. FM protein of a FM-based control diet (FM0) was replaced with 33.3 (FM33.3), 66.6 (FM66.7), and 100% (FM100) by SPC protein, and each replacement group was supplemented with 0.4% GMP to formulate four experimental diets. Each diet was randomly allocated to triplicate groups of fish (4.8 g) for 56 days. Results demonstrated that fish fed diet group FM33.3 had the significantly highest final weight, weight gain-specific growth rate, and feed intake. Meanwhile, in comparison to control, growth performance and feed utilization did not significantly differ with 66.7% FM replacement by SPC with GMP supplementation. Apparent digestibility coefficient of protein and lipid also followed a similar trend. All growth, feed utilization, and digestibility parameters were significantly lower in FM100 diet group. Blood urea nitrogen (BUN) and triglycerides (TG) increased (P < 0.05) with increasing FM replacement level by SPC. Interestingly, total cholesterol level reduces with the increasing level of FM replacement by SPC with GMP supplementation. Fish fed FM0 diet group showed the best condition of both oxidative and freshwater stress resistance. Meanwhile, FM33.3 and FM66.7 diet groups showed acceptable conditions. Innate immune responses enhanced with the increasing FM replacement level by SPC with GMP supplementation. In conclusion, FM could be replaced ≤66.7% by SPC with GMP supplementation in diets for red sea bream without any adverse effects on fish performances.     

Berberine attenuates oxidative stress and hepatocytes apoptosis via protecting mitochondria in blunt snout bream <Emphasis Type="Italic">Megalobrama amblycephala</Emphasis> fed high-fat diets

High-fat diets may have favorable effects on growth and cost, but high-fat diets often induce excessive fat deposition, resulting in liver damage. This study aimed to identify the hepatoprotective of a Chinese herb (berberine) for blunt snout bream (Megalobrama amblycephala). Fish were fed with a normal diet (LFD, 5 % fat), high-fat diet (HFD, 15 % fat) or berberine-supplemented diets (BSD, 15 % fat with berberine 50 or 100 mg/kg level) for 8 weeks. After the feeding, histology, oxidative status and mitochondrial function of liver were assessed. The results showed that HFD caused fat accumulation, oxidative stress and apoptosis in hepatocytes of fish. Hepatocytes in HFD group appeared to be hypertrophied, with larger liver cells diameter than these of LFD group. Berberine-supplemented diets could attenuate oxidative stress and hepatocytes apoptosis. HFD induced the decreasing mitochondrial complexes activities and bulk density and surface area density. Berberine improved function of mitochondrial respiratory chain via increasing the complex activities. Moreover, the histological results showed that berberine has the potential to repair mitochondrial ultrastructural damage and elevate the density in cells. In conclusion, our study demonstrated that berberine has attenuated liver damage induced by the high fat mainly via the protection for mitochondria.     

Physiological response,blood chemistry profile and mucus secretion of red sea bream (<Emphasis Type="Italic">Pagrus major</Emphasis>) fed diets supplemented with <Emphasis Type="Italic">Lactobacillus rhamnosus</Emphasis> under low salinity stress

Environmental stressors caused by inadequate aquaculture management strategies suppress the immune response of fish and make them more susceptible to diseases. Therefore, efforts have been made to relieve stress in fish by using various functional feed additives in the diet, including probiotics. The present work evaluates the effects of Lactobacillus rhamnosus (LR) on physiological stress response, blood chemistry and mucus secretion of red sea bream (Pagrus major) under low salinity stress. Fish were fed four diets supplemented with LR at [0 (LR0), 1 × 10² (LR1), 1 × 10⁴ (LR2) and 1 × 10⁶ (LR3) cells g^?1] for 56 days. Before stress, blood cortisol, urea nitrogen (BUN) and total bilirubin (T-BIL) showed no significant difference (P > 0.05), whereas plasma glucose and triglyceride (TG) of fish-fed LR2 and LR3 diets were significantly lower (P < 0.05) than those of the other groups. Plasma total cholesterol (T-CHO) of fish-fed LR3 diet was significantly (P < 0.05) lower than that of the other groups. Furthermore, total plasma protein, mucus myeloperoxidase activity and the amount of mucus secretion were significantly enhanced in LR-supplemented groups when compared with the control group (P < 0.05). After the application of the low salinity stress test, plasma cortisol, glucose, T-CHO and TG contents in all groups showed an increased trend significantly (P < 0.01) compared to the fish before the stress challenge. However, plasma total protein and the amount of secreted mucus showed a decreased trend in all groups. On the other hand, BUN, T-BIL and mucus myeloperoxidase activity showed no significant difference after exposure to the low salinity stress (P > 0.05). In addition, the fish that received LR-supplemented diets showed significantly higher tolerance against low salinity stress than the fish-fed LR-free diet (P < 0.05). The physiological status and the detected immune responses, including total plasma protein and mucus myeloperoxidase activity in red sea bream, will provide a more comprehensive outlook of the effects of probiotics to relieve stress in fish.     

Modulation of appetite,lipid and glucose metabolism of juvenile grass carp (<Emphasis Type="Italic">Ctenopharyngodon idellus</Emphasis>) by different dietary protein levels

This study was undertaken to explore the systemic metabolic strategies of juvenile grass carp (Ctenopharyngodon idellus) to maintain growth when fed with different dietary protein levels. The optimal growth group and two growing discomfort groups were selected through the basic data, to explain the growth difference from appetite regulation and lipid and glucose metabolism perspective. Three experimental diets were formulated with three dietary protein levels at 200.3, 296.1 and 442.9 g kg^?1, named P1, P2 and P3, respectively. Juvenile grass carp (initial body weight 12.28 ± 0.14 g) were fed with three diets with 3 replications per dietary treatment in an indoor recirculation system for an 8-week feeding trial. Fish fed with diet P2 dietary group showed significantly higher WG, SGR, FI and PER than other groups. Compared with other groups, mRNA expressions of NPY, Y8a and Y8b in fish fed with P2 significantly down-regulated, while the expressions of CCK and CART in fish fed with P3 significantly down-regulated (P < 0.05). With increasing dietary protein levels, G6Pase, GK, PK and PEPCK were all significantly inhibited (P < 0.05). For lipid metabolism, the mRNA expression of ACC in P1 dietary group was significantly higher than P3 dietary group; besides, LPL expression in P3 group was significantly higher than other two groups (P < 0.05). PPARα expression in P2 was significantly lower than other groups (P < 0.05). These results suggested that grass carp fed with P2 (296.1 g kg^?1 protein level) showed highest weight gain, contributed to more balanced nutrient metabolism and appetite regulation. Too high dietary protein (442.9 g kg^?1) should be avoided because it induced lowest PER, body lipid and liver lipid, and inhibited glucose and lipid metabolism in juvenile grass carp.     

Molecular cloning and expression analysis of <Emphasis Type="Italic">Megalobrama amblycephala</Emphasis> transferrin gene and effects of exposure to iron and infection with <Emphasis Type="Italic">Aeromonas hydrophila</Emphasis>

Transferrin (Tf) plays an important function in iron homeostasis and metabolism of organisms. In this study, we identified and characterized the Tf gene in Megalobrama amblycephala and evaluated its expression in basal conditions as well as after iron overload and experimental infection with Aeromonas hydrophila. Furthermore, we studied the iron binding properties of recombinant Tf. The full-length M. amblycephala Tf complementary DNA (cDNA) (GenBank accession no.: KX698308) of 2245 bp was cloned and contained a 1953 bp open reading frame (ORF) encoding 650 amino acid residues and flanked by a 68 bp 5′ and a 204 bp 3′ untranslated regions (UTR). Predicted conservative structure illustrated that M. amblycephala Tf consisted of two conservative Tf domains. Amino acid sequence alignment revealed that M. amblycephala Tf had high similarity with that of cyprinids deposited in Genbank, and phylogenetic analysis showed that M. amblycephala Tf clustered with Ctenopharyngodon idella and Hypophthalmichthys molitrix. Tissue expression pattern analyses demonstrated that the liver was the main Tf mRNA expressing organ, being significantly higher than other tissues (p < 0.05). In the liver, Tf mRNA expression in fish artificially injected with the pathogenic bacteria A. hydrophila was significantly upregulated, reaching a peak at 12 h post injection (hpi) and then decreasing afterward. The expression in FeCl₃-injected fish showed a similar tendency, but reached a peak at 8 hpi. Meanwhile, fish serum iron significantly decreased following A. hydrophila injection, but increased to peak at 4 hpi and then decreased in FeCl₃-injected fish. The recombinant M. amblycephala Tf showed iron binding capacity using CAS analysis. These results are helpful to understand the structure and regulation of expression of Tf, as well as the specific function of Tf for both immune responses and iron homeostasis.     

Effects of dietary supplementation with green tea waste on growth,digestive enzyme and lipid metabolism of juvenile hybrid tilapia, <Emphasis Type="Italic">Oreochromis niloticus</Emphasis> × <Emphasis Type="Italic">O. aureus</Emphasis>

An 8-week feeding trial was conducted to evaluate the effects of dietary supplementation with green tea waste (GTW) on growth, digestive enzyme and lipid metabolism of juvenile hybrid tilapia, Oreochromis niloticus × O. aureus. The fish (initial mean body weight, 12.63 ± 0.75 g) were fed five experimental diets that included 0 (control), 0.8, 1.6, 3.2 or 6.4 % of GTW in triplicate aquaria, twice daily. Growth performance, plasma metabolites content and liver and intestine digestive enzyme activities were determined. Fish accepted well all experimental diets during the trial, and no mortality was observed. The weight gain increased (P < 0.05) with the increase in GTW inclusion level up to 1.6 %, after which it decreased, but no significant differences between the control and high level (3.2 or 6.4 % of GTW) groups were observed. Moreover, fish fed on diets containing 0.8 and 1.6 % GTW had lower feed conversion ratio (FCR, 1.75 and 1.73, respectively) and had better protein deposition (higher protein efficiency ratio, PER, 1.73 and 1.71, respectively), compared to other treatments. No differences among groups were observed in whole body and dorsal muscle composition with the exception of lipid content which was lower in fish fed 6.4 % GTW diets, compared to other treatments. Lipase activities in liver or intestine were higher in fish fed GTW-supplemented diets with the exception of intestine lipase activities, which was unaffected, compared to the control. Similarly, liver lipoprotein lipase activities were also increased in fish fed diets supplemented a medium dose of GTW (1.6 or 3.2 %), compared to other treatments. However, intestine amylase activities were decreased in fish fed diets containing a high dose of GTW (3.2 and 6.4 %); while the liver amylase activities were unaffected by the GTW supplementation. Blood chemistry parameters were affected by GTW inclusion, except the values of triglycerides, which was unaffected. The values of total cholesterol, HDL cholesterol and LDL cholesterol increased with increasing GTW inclusion level up to 3.2 %, after which the values decreased. These results indicate that diets supplemented with appropriate concentration of GTW (from 0.8 to 1.6 %) may potentially serve as an effective functional food and additive for tilapia to improve growth performance, digestion efficacy and fat metabolism.     

Evaluation of different <Emphasis Type="Italic">Pyropia yezoensis</Emphasis> extracts as feed additives for growth and immunity of Japanese flounder <Emphasis Type="Italic">Paralichthys olivaceus</Emphasis>

We evaluated the effects of diets incorporating the red algae Pyropia yezoensis, prepared by several different extraction methods, on the growth of juvenile Japanese flounder Paralichthys olivaceus. We assessed growth performance, as well as the levels of amino acids, fatty acids, insulin-like growth factor I (IGF-I) and interleukins (ILs). Four experimental diets were developed based on different methods of processing P. yezoensis. A commercial feed, laver powder (P), high-pressure heat extraction of laver (HPHE) and acid hydrolysis extraction of laver were used as the experimental diets. Three experimental replicates were established for each diet (40 fish/group, body weight 123.7 ± 1.1 g), and the fish were fed for 6 weeks. We found no significant differences in weight gain, specific growth rate or feeding efficiency among the groups (P > 0.05); however, the fish fed HPHE had the greatest growth performance. Fish fed the laver extracts exhibited the highest protein efficiency ratio compared with the control and P groups. The experimental groups fed P. yezoensis extracts had significantly higher levels of IGF-I (P < 0.05) than those of the control group. High levels of IL-2 were found in the P and HPHE groups, IL-12 in the HPHE group, and IL-6 in all experimental groups. Therefore, these results suggest that a P. yezoensis extract improved the growth performance and immunity of Japanese flounder. In particular, the high-pressure heating process was a useful extraction method for preparing a P. yezoensis extract, which had beneficial effects as a dietary supplement in Japanese flounder.     

Dietary ascorbic acid reduced micronucleus and nuclear abnormalities in <Emphasis Type="Italic">Clarias gariepinus</Emphasis> (Burchell 1822) exposed to hospital effluent

Hospital effluents contain myriad of mutagens and genotoxins capable of increasing DNA damage in aquatic biota. African mudfish, Clarias gariepinus, are exposed to genotoxins when cultured in swamps and derelict water bodies often contaminated by effluents. Moreover, its DNA is susceptible to xenobiotic-induced lesions since it lacks l-gulonolactone oxidase and hence cannot synthesize l-ascorbic acid. This study investigated 96-h acute toxicity and protective effects of dietary ascorbic acid (AA) against micronucleus (MN) and abnormal nuclear (NAs) formation in C. gariepinus exposed to sub-lethal concentrations of hospital effluent. Six concentrations (0.5–3.0%) of the effluent were selected to determine the 96-h acute toxicity of the effluent in C. gariepinus, after range finding test. Fish were exposed to sub-lethal concentrations (0.08–1.30%) of the 96 h LC₅₀. Two other groups were exposed to the 96 h LC₅₀ (1.30%) of the effluent +50 and +100 mg/kg of dietary ascorbic for 7 days, and MN and NAs assessed in peripheral erythrocytes. The 96 h LC₅₀ (1.30%) was 1.18 times more toxic than the 24 h LC₅₀ (1.54%), indicating that the toxicity of the effluent increased with exposure duration. MN, nuclear bud, enucleated, fragmented nucleus (apoptosis), and necrotic erythrocytes significantly increase in effluent treated fish. Dietary AA reduced MN from 6.35-fold (1.30% treated group) to 3.72-fold (1.30% + 50 mg AA) and 3.54-fold (1.30% + 100 mg AA). Also, AA reduced total NAs from 2.26-fold (1.30%) to 1.40-fold (1.30% + 50 mg AA) and 1.06-fold (1.30% + 100 mg AA) compared to the control. Heavy metals and physicochemical parameters analyzed in the tested effluent possibly induced the mortality and cytogenotoxicity in C. gariepinus, and this was ameliorated by dietary AA.     

Characterization of the isoforms of type IIb sodium-dependent phosphate cotransporter (Slc34a2) in yellow catfish, <Emphasis Type="Italic">Pelteobagrus fulvidraco</Emphasis>, and their vitamin D<Subscript>3</Subscript>-regulated expression under low-phosphate conditions

In this study, two isoforms slc34a2 genes (type IIb sodium-dependent phosphate cotransporter), slc34a2a2 and slc34a2b, were cloned from intestine and kidney of yellow catfish (Pelteobagrus fulvidraco), with rapid amplification of cDNA ends. The structure differences and the regulation effects of dietary VD₃ under low phosphorus were compared among three isoforms of slc34a2 in yellow catfish. The predicted Slc34a2a2 and Slc34a2b proteins match 65 % and 53.8 % sequence identity, with Slc34a2a1, respectively. The membrane-spanning domains were different among these three isoforms. Intestinal Slc34a2a1 and Slc34a2a2 proteins had eight and eleven transmembrane domains, while renal Slc34a2b protein had nine. The tissue distribution study showed that same as slc34a2a1, slc34a2a2 mRNA was mainly distributed in intestine and slc34a2b mRNA in kidney. The effect of vitamin D₃ (VD₃) level on slc34a2 subfamily expression under low-phosphate conditions, induced by the addition of 0 (VD0), 324 (VD1), 1243 (VD2), 3621 (VD3), 8040 (VD4), or 22700 (VD5) IU VD₃/kg feed, was assessed by qPCR. The dose-responsive expression of intestinal slc34a2a2 and high expression of intestinal slc34a2a2 in VD5 together with peak expression of kidney slc34a2b in VD3 coincided with the accumulation of body phosphate content. These data suggested that appropriate level of dietary VD₃ up-regulated slc34a2a1, slc34a2a2, and slc34a2b mRNA levels, which increased phosphate retention. In conclusion, the current study provided another possible approach to improve dietary phosphate utilization by adding appropriate level of VD₃ to a low-phosphate diet to regulate intestinal and renal slc34a2 gene expression and thus minimize the excretion of phosphorus in yellow catfish.     

Feeding,somatic condition and survival of sand lance <Emphasis Type="Italic">Ammodytes</Emphasis> sp. larvae in Mutsu Bay,Japan

To clarify the recruitment process of sand lance Ammodytes sp., we investigated larval condition factor, relative gut fullness (%GF), prey abundance and oceanographic structure in Mutsu Bay, Japan, during 1999–2001. Ammodytes sp. larvae, which were collected by horizontal hauls of Motoda nets and a ring net at depths of 1, 10, 20, 30 and 40 m, were mainly distributed at 10–30 m. Larvae at the first feeding time until 12 mm in body length (BL) fed predominantly on copepod nauplii, whereas large larvae with BL of 12.1–14.0 mm fed on a mixture of copepod nauplii, copepodites and appendicularians from late February to April. A path analysis showed that difference in water density between 35- and 5-m depths negatively affected naupliar abundance at 10–30-m depth (standardised path coefficient β = ?0.71, p = 0.005 for 3.3–8.0-mm BL larvae and β = ?0.78, p < 0.001 for 8.1–12.0-mm BL larvae). Naupliar abundance positively affected the %GF of Ammodytes sp. larvae (β = 0.75, p < 0.001 for 3.3–8.0-mm BL larvae and β = 0.66, p < 0.001 for 8.1–12.0-mm BL larvae), whereas it was negatively affected by water temperature (β = ?0.45, p = 0.008 for 3.3–8.0-mm BL larvae and β = ?0.56, p = 0.002 for 8.1–12.0-mm BL larvae), and the temperature effect was weak compared with that of naupliar abundance. In turn, %GF positively affected larval somatic weight (β = 0.91, p < 0.001 for 6.0-mm BL larvae and β = 0.70, p = 0.005 for 10.0-mm BL larvae). The recruitment failure in 1999 was likely caused by a reduced condition factor, which resulted from low naupliar abundance. In contrast, the abundance of nauplii and Oithona similis copepodites was high in 2000 and 2001. It is possible that the higher recruitment success in 2001 was because of the higher water temperatures in Mutsu Bay, sustaining faster growth of the larvae than in 2000 under the high-prey abundance conditions.     

Supplementation of enzyme-treated soy protein saves dietary protein and promotes digestive and absorptive ability referring to TOR signaling in juvenile fish

This study was conducted to evaluate the effect of enzyme-treated soy protein (ETSP) supplementation in the low-protein diet on growth performance, digestive and absorptive capacities, and related signaling molecules’ gene expressions in juvenile Jian carp. The results showed that percent weight gain (PWG), specific growth rate (SGR), and feed intake (FI) were decreased by reducing dietary protein from 34 to 32% (P < 0.05). Supplying low-protein diet with optimal ETSP increased previously mentioned indices of juvenile Jian carp (P < 0.05), which also had no significant difference with the high-protein diet (34%CP) (P > 0.05). Compared with the low-protein diet, appropriate ETSP supplementation in the low-protein diet increased (P < 0.05) (1) the trypsin, lipase, and amylase activities in the hepatopancreas; (2) cholecystokinin concentration in the proximal intestine; (3) the γ-glutamyl transpeptidase (γ-GT), alkaline phosphatase (AKP), and Na⁺/K⁺-ATPase activities in all intestinal segments; and (4) the messenger RNA (mRNA) levels of trypsin, lipase, and amylase in hepatopancreas and γ-GT in the mid (MI) and distal (DI) intestine, alkaline phosphatase in MI, and Na⁺/K⁺-ATPase and target of rapamycin in all intestinal segments. At the same time, appropriate ETSP supplementation in the low-protein diet downregulated the mRNA levels of AKP in the DI and eIF4E-binding protein 2 in all intestinal segments (P < 0.05). In conclusion, adding 10 g ETSP/kg diet in the low-protein diet can restore the growth performance and digestive and absorptive abilities to the levels in group with 34% dietary protein. Supplementation of optimal ETSP in the low-protein diet enhanced the digestive and absorptive abilities and regulated the signaling molecules related to the TOR signaling pathway.