| Research one: Effects of dietary starch sources on the growth performance,body composition,antioxidant capacity of hybrid grouper(Epinephelus fuscoguttatus ♀ × E.lanceolatus ♂)Four iso-nitrogenous and iso-lipid diets(M4,P4,Y4 and G4)were formulated with cassava starch(M),pea starch(P),corn starch(C)and glutinous rice starch(G)and their levels of 4%,respectively,to estimate suitable starch source in diet for hybrid grouper,based on growth performance,feed efficiency,body composition,serum glucose,and antioxidant capacity.Each diet was randomly assigned to triplicate tanks of 35 fish with an average initial weight(23.46 ± 0.19g).Test fish were fed to apparent satiation twice daily(7:00 and 16:00)for12 weeks.During the experimental period,all tanks were provided with continuous aeration and the dissolved oxygen was ≥ 6 mg/L,water temperature was maintained at(28 ± 1)℃,and the tanks were maintained under a natural light cycle.The results of this research are presented as follows:1.Effects of starch sources on growth performance,body composition and utilization of feed nutrients of test fish.(1)With the inclusion level of 4% starch source had no significant effect on the survival rate of the pearl gentian grouper(P > 0.05).In terms of growth performance,there was no significant difference in the final body weight and specific growth rate of pearl gentian grouper fed with pea starch,cassava starch,corn starch and glutinous rice starch.There was no significant effect on the CF and VSI among four groups(P > 0.05).The HSI of pea starch group was significantly lower than the other three groups(P < 0.05).(2)In terms of fish body composition the result show that there was no significant difference in water,ash c and crude protein content among four different starch diets(P > 0.05).The crude fat content of pea starch,cassava starch and glutinous rice starch group was significantly higher than that of corn starch group(P > 0.05).There was no significant effect in water,crude fat and crude protein content in the liver of the tested fish(P > 0.05),but the glycogen content in the liver of the pea starch group was significantly lower than the other three groups(P < 0.05).There was no significant difference in water,glycogen and crude protein content in muscle of test fish(P > 0.05),but crude fat content in waxy rice starch group was significantly higher than that in other three groups(P < 0.05).(3)In terms of the feed utilization,with the inclusion level of 4% starch source had no significant effect on the feeding rate of the tested fish(P > 0.05).Four kinds of starch sources had significant effects on feed efficiency,protein efficiency,protein deposition rate and fat deposition rate of grouper(P < 0.05).The feed efficiency,protein efficiency and protein deposition rate of pea starch group were significantly higher than those of other three groups(P < 0.05),while the fat deposition rate of pea starch group was significantly lower than that of cassava starch group and glutinous rice starch group(P < 0.05).And the apparent digestibility of starch and fat in pea starch group was significantly lower than that in cassava starch,corn starch and glutinous rice starch group(P < 0.05),while the apparent digestibility of protein in pea starch group was significantly higher than that in other three groups(P < 0.05).3.Effects of starch sources on Antioxidant Enzyme Activity in liver of hybrid grouperHybrid grouper fed diets containing different starch sources for 12 weeks,the MDA content in liver of pea starch group was significantly lower than that of cassava starch,corn starch and glutinous rice starch group(P < 0.05).The activities of CAT and GSH-PX in liver of pea starch group were significantly higher than those of other three groups(P < 0.05).There was no significant difference in SOD activity in liver among the four groups(P > 0.05),and the highest SOD activity appeared in pea starch group.Research two: Effects starch sources and levels on growth performance,postprandial blood glucose and antioxidant ability of hybrid grouperNine iso-nitrogenous and iso-lipid diets(L5,L10,L15,W5,W10,W15,M5,M10 and M15)were formulated with cassava starch(L),pea starch(W),starch and Pullulan enzymatic cassava starch(M)and their levels of 5%,10% and 15%,respectively,to estimate suitable starch source and level in diet for hybrid grouper,based on growth performance,feed efficiency,body composition,serum glucose,and antioxidant capacity.Each diet was randomly assigned to triplicate tanks of 35 fish with an average initial weight(10.29 ± 0.04g).Test fish were fed to apparent satiation twice daily(7:00 and 16:00)over 12 weeks.During the experimental period,all tanks were provided with continuous aeration and the dissolved oxygen was ≥ 6 mg/L,water temperature was maintained at(28 ± 1)℃,and the tanks were maintained under a natural light cycle.The results of this research are presented as follows:1.Effects of starch sources and levels on growth performance,body composition and utilization of feed nutrients of test fish.(1)Starch sources and levels had no significant effect on the survival rate of the pearl gentian grouper(P > 0.05).In terms of growth performance,with the adding level of starch with the same kind increasing from 5% to 15%,the final body weight,specific growth rate and feeding rate of the test fish of cassava starch,pea starch and pullulan hydrolyzed cassava starch group were significantly decreased(P < 0.05).with the adding level of starch with the same kind increasing from 5% to 15%,the CF,VSI and HSI of test fish had significantly increased(P < 0.05).With the same inclusion level of dietary starch,the CF,VSI and HSI of pullulan enzymes starch group had significantly lower than those of the other two groups(P < 0.05).With the inclusion level of 10% starch source,the specific growth rate,final body weight and feed efficiency of the pullulan-digested tapioca starch were significantly higher than those of the tapioca starch and pea starch groups(P < 0.05).(2)In terms of fish body composition,the result show that,with the adding level of starch with the same kind increasing from 5% to 15%,the crude protein and crude fat content of the whole fish were significantly decreased(P < 0.05),while the moisture content of the whole fish was significantly increased(P < 0.05).However,there was no significant effect on the ash content of the whole fish.The crude fat content of fish muscle was significantly decreased(P < 0.05),while the glycogen content of muscle was significantly increased(P < 0.05).The crude protein content of muscle in cassava starch group was significantly decreased(P < 0.05),but had no significant effect on the crude protein content of muscle in pea starch and pullulan enzymatic cassava starch group(P > 0.05).The crude protein content and crude fat content in the liver of experimental fish was significantly decreased(P < 0.05),while the glycogen content in the liver increased significantly(P < 0.05).There was no significant effect on the liver water content(P > 0.05).(3)In terms of the feed utilization,with the adding level of starch with the same kind increasing from 5% to 15%,the feeding rate of test fish was significantly decreased(P < 0.05).The feed efficiency of cassava starch and pea starch groups decreased significantly(P < 0.05),while the feed efficiency of pullulan enzymatic cassava starch group was significantly higher than that of 5% and 15% groups at 10% level(P < 0.05).The apparent starch digestibility and fat digestibility of pullulan hydrolyzed cassava starch were significantly lower than those of cassava starch and pea starch(P < 0.05),while protein digestibility of pullulan hydrolyzed cassava starch was significantly higher than those of other two groups(P < 0.05).2.Effects of different starch sources and levels on postprandial blood glucose and liver antioxidant enzyme activity in hybrid grouperThe effect of postprandial blood glucose levels on the test fish was significant(P < 0.05).At the same time point(-3,1,3,6,9,12 and 24h),as the amount of the same kind of starch source in the feed increased(5%~15%),the postprandial blood glucose of the pearl gentian grouper The level was significantly increased(P < 0.05)and the postprandial blood glucose of the pullulan-digested cassava starch group was significantly lower than the other two groups(P < 0.05).The liver protein concentration,CAT and T-AOC activity of the test fish were significantly decreased(P < 0.05),while the SOD activity of the liver was significantly enhanced(P < 0.05).Research three:Molecular cloning,characterization,tissue expression and nutritional regulation of O-GlcNAc transferase gene in hybrid grouperIn the present study,the OGT gene of hybrid grouper was cloned and characterized,and its expression in response to dietary carbohydrate level and acute glucose treatment was investigated.The full-length of OGT(GenBank accession no.KY656469)was 4,063 bp,including a 302 bp 5’ untranslated terminal region(UTR),a 3,165 bp coding region that encoded 1,054 amino acids residues and a 596 bp 3’ UTR.The highly conservation of OGT gene between fish and mammals was also observed through multiple sequences alignment and phylogenetic analysis.O-GlcNAc transferase gene was ubiquitously expressed in all detected tissues with highest expressions in brain and liver,to a lesser degree,in eye,heart,kidney and intestine.The increasing dietary carbohydrate from 8.02% to 16.08% had no significant effect on the mRNA expression of OGT.However,the expression of OGT was slightly elevated at 6 h post-glucose injection,and the elevation became significant at 24 h time-point.These data may enhance our understanding on the nutritional regulation of OGT and O-GlcNAc modification in fish species.3.ConclusionDifferent starch sources had no significant effect on the growth performance of hybrid grouper,but pea starch significantly increased the feed efficiency,protein efficiency and protein deposition rate of hybrid grouper compared with cassava starch,corn starch and glutinous rice starch.The glycogen content in liver of pea starch group was significantly lower than the other starch groups.The antioxidant ability of test fish fed pea starch concluded diet was significantly improved in the liver,compared with cassava starch,corn starch and glutinous rice starch.The increase of starch level significantly reduced the growth performance and feed utilization rate of hybrid grouper,resulting in the accumulation of glycogen in the liver and the activity of antioxidant enzymes in the liver.With the inclusion level of 10% starch source,the pullulan hydrolyzed cassava starch group was better than cassava starch and pea starch.The full-length sequence of OGT gene was cloned.The increasing dietary carbohydrate from 8.02% to 16.08% had no significant effect on the mRNA expression of OGT.However,the expression of OGT was slightly elevated at 6 h post-glucose injection,and the elevation became significant at 24 h time-point.These data may enhance our understanding on the nutritional regulation of OGT and O-GlcNAc modification in fish species. |
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