Research And Application On Intermediate Culture Technology Of Juvenile Sipunculus Nudus Based On Formulated Diet

The development of nutritionally appropriate formulated diet is the key to achieve high-density and large-size seedlings production of Sipunculus nudus.In this thesis,based on the D1 diets formulated by the group in the early stage,we optimized the formulation of S.nudus particulate diets and explore the corresponding culture technology based on the optimized diets inputs,and initially establish the technical scheme for the intermediate culture of S.nudus seedlings,with the following main findings.1.Iso-fat and iso-energetic D2 series formulated diets with different carbohydrate/protein levels(C/P)were formulated:D2-1(39.46%/20.67%),D2-2(34.97%/24.02%),D2-3(30.48%/27.37%),D2-4(25.99%/30.72%)and D2-5(21.50%/34.07%),S.nudus juveniles were carried out for culture experiments.After90 d of culture,there were significant differences in the final body weight(TBW),weight gain rate(WGR),specific growth rate(SGR)and survival rate(SR)of S.nudus raised on different diets(P<0.05),with the best TBW,WGR,SGR and SR of S.nudus in the EG2 group(D2-2 diet);the worst growth of S.nudus in the EG5 group(D2-5 diet).Body composition analysis showed that different diets had significant effects on the moisture of the body wall of S.nudus juvenile and the crude protein,crude fat and ash content of their dry matter(P<0.05);16 amino acids(AA)were detected,the differences between groups in total AA,total essential AA,and total presenting AA were not significant(P>0.05),and there were significant differences between groups in the content of nine amino acids,including methionine,isoleucine,and valine(P<0.05).Biochemical assays showed that different diets had significant effects on the antioxidant index and digestion-related enzyme activities of the S.nudus juvenile(P<0.05),with the EG2 group(D2-2 diet)having higher reduced glutathione content as well as amylase and lipase activity.2.The above D2 series diets were used again to conduct experiments on the culture of S.nudus juveniles to verify their production and to analyze the metabolic effects of the diets on the S.nudus.The results after 90 d of culture showed that the different diets had significant effects on the TBW,WGR,SGR and SR of the S.nudus(P<0.05),with the eg2 group(D2-2 diet)having the best culture effect and the eg5group(D2-5 diet)having the worst culture effect.The results of metabolomic analysis showed that 83 significantly different metabolites(POS:59;NEG:24)were present in the body cavity fluids of the eg2 and eg5 groups of S.nudus,which were involved in22 metabolic pathways(POS:9;NEG:13),including the biosynthesis of tyrosine,phenylalanine and tryptophan;phenylalanine metabolism;D-glutamate and D-glutamine metabolism;proline and arginine metabolism;aspartate,alanine,and glutamate metabolism;and aminyl-t RNA biosynthesis.3.Several single-factor experiments were set up to compare the effects of different environmental conditions on the culture effect of S.nudus juveniles using D2-2 diet as the nutrient source.The results showed that after 30 d of culture,(1)the experimental temperatures(15°C,20°C,25°C,30°C,35°C)had significant effects on the survival rate,growth indexes,digestive enzyme activity and other assays of S.nudus juveniles(P<0.05),and the regression equation of total production and culture temperature was:y=-0.0239x~3+1.6059x~2-34.035x+285.31(R2=0.9927),and the maximum value of total yield was obtained when the temperature was 27.59°C.(2)The experimental salinity(15‰,20‰,25‰,30‰,35‰)had a significant effect on the above detection indexes(P<0.05),and the regression equation of total yield versus culture salinity was:y=0.0174x~3-1.3609x~2+33.51x-190.12(R2=0.9512),and the maximum total yield was obtained when the salinity of seawater was 19.93‰;(3)the light conditions(natural light,semi-shade,light,full shade)had a significant effect on the above detection indexes(P<0.05),in which the S.nudus feeding activity was the most vigorous in the natural light group,and the survival rate,growth rate and total yield were significantly higher than the rest of the groups(P<0.05).4.Several single-factor experiments were set up to compare the effects of different management measures on the culture effect of S.nudus juveniles using D2-2diet as the nutrient source.The results showed that(1)after 30 d of culture,the experimental feeding amounts(1.5%,3.0%,4.5%,6.0%)had significant effects on the survival rate,growth indexes,digestive enzyme activity and other detection indexes of S.nudus juveniles(P<0.05),and the regression equation of total production and feeding amount was:y=6.0576x~3-81.535x~2+315.8x+605.63(R2=1),when the feeding amount is 2.83%,the total production obtains the maximum value;(2)after 30 d of culture,the experimental culture density(2000 ind/㎡,2500ind/㎡,3000 ind/㎡,3500 ind/㎡,4000 ind/㎡)has a significant effect on the above detection index(P<0.05),the regression equation of total production and culture density is:y=0.0307 x+18.607(R2=0.9944),within the specifications of 0.08-0.10g/ind,the culture density of S.nudus juveniles can reach 4000 ind/㎡,and the total yield obtains the maximum value;(3)after 60 d of culture,the frequency of tilling(0d,7 d,15 d,30 d)has no significant effect on S.nudus juveniles TBW,WGR,SGR,SR,total yield,ingestion rate and satiation rate were not significantly affected(P>0.05).5.Referring to the results in Chapter 3,a trial group(D2-2 diet as the nutrient source)and a control group(existing culture technology of the enterprise)were set up for seedling mesoculture trial production,and after 140 d of culture,the results showed that the TBW,WGR and SGR of the trial group were significantly higher than those of the control group(P<0.05);compared with the control group,the survival rate,yield and production value of the trial group were increased by 7.04%,92.17%and 46.35%,respectively,compared with the control group.