| The research on suitable lipid levels,lipid sources and glycolipid ratios for E.sinensis diets under different salinity conditions is extremely insufficient,which seriously restricts the precise compound diet for E.sinensis in saline-alkali land development.Therefore,this study firstly evaluated the optimal sources of lipid in the diet of E.sinensis under different salinity conditions,and determined the optimal level of lipid in the diet of juvenile crabs under different salinity conditions.On the basis,the appropriate glycolipid ratio and glycolipid metabolism mechanism of juvenile crabs in different salinity conditions and molting period were explored.The study results can provide a theoretical basis for optimizing the diet formula of the salt-alkali land crab and improving the comprehensive benefit of the salt-alkali land crab breeding.The main findings and conclusions of this study are as follows:(1)Effects of dietary lipid sources on the survival,growth,body composition,antioxidant capacity and expression of antioxidant and pro-inflammatory genes in juvenile Chinese mitten crab(Eriocheir sinensis)reared under three salinities: The purpose of this experiment was to investigate the survival,growth,body composition,antioxidant capacity and related gene expression of E.sinensis which were fed diets with different lipid sources under three salinities(0.5‰,2‰ and 6‰).5 isonitrogenous and lipid diets were formulated with one of the candidate oils(fish oil,soybean oil,linseed oil,colza oil or palm oil)and were named FO,SO,LO,CO or PO respectively,each diet was randomly fed to 3 groups of crabs(initial weight : 0.21± 0.02 g)under different salinity for 60 days.Results showed that the survival rate(SR)of E.sinensis showed a decreasing tendency as the salinity increased,but SR was not significantly affected by dietary lipid sources at each salinity(P>0.05).Weight gain rate(WGR)were highest in the treatment of FO/PO at a salinity of 0.5‰,CO/FO/SO at a salinity of 2‰ and LO at a salinity of6‰ respectively.At salinities of 0.5‰ and 2‰,the transcription of pro-inflammatory and antioxidant genes in LO was significantly higher than that in FO or PO(P<0.05).While at a salinity of 6‰,the transcription of detected genes in FO was significantly increased compared to that in the vegetable oil treatments(P<0.05).However,the docosahexaenoic acid(DHA)contents in E.sinensis fed the diets with FO were still significantly higher than those with vegetable oils(P<0.05).The above results indicate that the antioxidant capacity of E.sinensis were decreased by aquaculture water salinity increases and therefore negatively affected the survival and growth rates of experimental animals.The beneficial effects of FO on survival and growth performance were eliminated by increasing the salinity to 6‰,which could be due to the increased lipid peroxidation and inflammatory response caused by salinity stress.So,it was concluded that the optimal lipid source in E.sinensis diet were FO/PO at the salinity of 0.5‰,CO/FO/SO at the salinity of 2‰,and LO at the salinity of 6‰.(2)Effects of dietary lipid levels on the survival,growth,body composition,digestive enzyme activities and antioxidant capacity in juvenile E.sinensis reared under three salinities:The purpose of this experiment was to investigate the survival,growth,body composition and antioxidant capacity of E.sinensis which were fed diets with different lipid levels under three salinities(0.5‰,2‰ and 6‰).Six isonitrogenous diets with graded levels of lipids(5%,7%,9%,11%,13% and 15%)were formulated by including fish oil as the main lipid source,each diet was randomly fed to 3 groups of crabs(initial weight: 0.21± 0.02 g)under different salinity for 56 days.Results showed that the survival rate of E.sinensis fed diets with 7%～15% lipids just showed a decreasing tendency as the salinity increased.The highest weight gain rate(WGR)was found in the treatment with 11%,9% and 5% lipid at the salinity of 0.5‰,2‰ and 6‰respectively.At the salinity of 0.5‰,2‰ and 6‰,the highest activities of antioxidant enzymes(superoxide dismutase,catalase,glutathione peroxidase)were detected in the treatments with11%,13% and 7% lipid respectively.As the salinity increased,the protein and lipid contents showed an increasing trend,while the moisture content showed an opposite trend.At each salinity,arachidonic acid(ARA)and DHA increased,while saturated fatty acids and monounsaturated fatty acids decreased with the increase of dietary lipid level.The above results indicate that increasing salinity resulted in decreased antioxidant enzyme activities and increased malondialdehyde content in the juvenile E.sinensis,especially when they were fed diets with relatively higher lipid level.Based on SR,WGR,digestive enzyme activity and antioxidant capacity,the optimal lipid level in E.sinensis diet was estimated to be 8.28～13% at the salinity of 0.5‰,7.54～13% at the salinity of 2‰ and 5～8.89% at the salinity of 6‰.(3)Effects of dietary carbohydrate-to-fat ratio(glycolipid ratio)on growth performance,biochemical indexes,metabolic enzyme activities and differential gene expression of juvenile E.sinensis at different molting stages: The purpose of this experiment was to investigate the effects of dietary glycolipid ratio on the survival and growth of juvenile crabs under different salinities,as well as the effects on biochemical indicators,metabolic enzyme activities and transcriptome analysis at different molting stages.5 isonitrogenous diets were formulated with corn starch used as the main sugar source and soybean oil used as the main lipid source,and five different diets were formulated with glycolipid ratios of 1.34,2.39,3.59,5.52 and 9.42,respectively.Each diet was randomly fed to 3 groups of crabs(initial weight : 1.09± 0.01 g)under different salinity for58 days.The results showed that the SR of juvenile crabs was higher than 90%,and the SR of juvenile crabs in C/L3.59 group was the highest which was higher than the other groups,and there was almost no difference among the groups at 6‰ salinity.At each salinity,the C/L3.59 group had the highest WGR and was significantly higher than other groups(P<0.05).With the increase of dietary glycolipid ratio,the triglyceride content of juvenile crabs at the In M showed a decreased trend,and the triglyceride content of the juvenile crabs at the In M was higher than that at the Pr M,while the hepatic glycogen content was the opposite.With the increase of dietary glycolipid ratio,the glycogen synthase activity of juvenile crabs at each molting stage was significantly increased(P<0.05);the hexokinase and pyruvate kinase activities of juvenile crabs at the Pr M also showed an increased trend,while the activities of hexokinase and pyruvate kinase at the In M showed an opposite trend;the activity of fatty acid synthase in juvenile crabs at the In M showed a significant decreased trend(P<0.05),while the activity of fatty acid synthase at the Pr M showed first decreased and then increased.Through transcriptome analysis,a total of 118.16 Gb Clean Data was obtained.Through the enrichment of KEGG pathway,differential genes were enriched in fatty acid biosynthesis,fatty acid metabolism,oxidative phosphorylation pathway,pentose phosphate pathway and pyruvate metabolism.Through real-time fluorescence quantitative PCR verification,the quantitative expression of genes in each feed group was basically consistent with that in the transcriptome.When the stage were the In M,with the increase of dietary glycolipid ratio,the expression levels of genes Fasn,PCK,G6 PDH and Ndufa6 all showed an increased trend,while the genes Elovl6 and Aco expression level showed a decreased trend;when the stage were the Pr M,the expression levels of the genes Fasn,PCK and Acly showed a increased trend,while the expression levels of the genes G6 PDH and Aco showed a decreased trend.Compared with crabs at the Pr M,the C/L1.34 and C/L3.59 juvenile crabs at the In M had lower expression levels of CPT1α gene,but the opposite was true in the C/L9.42 group.The above results show that when the dietary glycolipid ratio is 3.59,it is more favorable for the growth of juvenile crabs.The mutual conversion between carbohydrates and lipids in juvenile crabs was more active at the In M than at the Pr M.The body lipid content of crabs at the Pr M decreased,while the accumulation of liver glycogen increased,which indicates that the crabs may mainly rely on lipid for energy during the transition from the In M to the Pr M,while the crabs at the Pr M to the molt stage mainly relied on the liver glycogen for energy supply. |
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