The Study On Immune Effects,Feeding Physiology And Intestinal Health Of Soybean Antigen Protein On Giant Freshwater Prawn(Macrobrachium Rosenbergii)

1.The replacement of fish meal and soybean meal to fermented soybean meal and its effects on the growth performance,serum biochemical indices,and immune gene expression in giant freshwater prawn,Macrobrachium rosenbergiiAn experiment was conducted to investigate the effects of replacing fish meal(FM)and soybean meal(SBM)by fermented soybean meal(FSBM)on growth performance,serum biochemical indices,and immune genes in giant freshwater prawn,Macrobrachium rosenbergii,and to find a suitable proportion of FSBM to replace FM in diets.Five isonitrogenous and isoenergetic diets were formulated for giant freshwater prawn,among which a basal diet(T0)containing 30% FM and 18% SBM was treated as the control group,and the FM and SBM(2:1)in the basal diets replaced by 2%(T2),5%(T5),8%(T8)and 15%(T15)of FSBM respectively were treated as experimental diets.Giant freshwater prawns with initial average body weight of 0.17±0.02 g were cultured for 64 days in indoor net cages at a cement tank.Results showed that with increasing FSBM supplemental level,the growth rate and specific growth rate increased firstly and then decreased,and the highest value was found in group T8.Serum superoxide dismutase activity and malondialdehyde content firstly increased and then decreased.MDA of all experimental groups was higher than that of the control group.Serum aspartate transaminase,alanine aminotransferase activity and total protein content had decreasing trend followed by an increasing trend,and the values in all experimental groups were lower than the control group.Gills Toll receptor m RNA and NF-?B m RNA expression levels and hepatopancreas heat shock protein m RNA relative expression levels in group T15 were the highest among all groups.All these results indicated that the replacement of FM and SBM with FSBM would affect growth,antioxidant ability and immunity,and the optimum amount of FSBM was 8% in the diets in this experiment.2.The study on physiological,biochemical and immune effects of soybean antigen proteins on giant freshwater prawn(Macrobrachium rosenbergii)Soybean antigen proteins(Soybean glycinin and ?-conglycinin)are main antinutritional factors in soybean meal.Recent studies have shown that high level of glycinin and ?-conglycinin in soybean meal would break the intestinal tissue structure resulting in disorder of digestion and absorption,stall growth and allergic reactions with a variety of non-specific and specific immune responses in aquatic animals.In recent years,fermented soybean meal has been used increasingly in aquatic animal feed.Compared with soybean meal,the anti-nutritional factors,especially soybean antigen proteins in fermented soybean meal are drastically reduced,therefore,it may be a better plant protein.This study compared the effects of soybean meal(SBM)and fermented soybean meal(FSBM)on growth,serum biochemical indices,gene expression and oxidative status on Macrobrachium rosenbergii,and evaluated whether the effects were caused by soybean antigen protein.Practical diets(T15,TF)and purified diets(AP0,AP5)were formulated for feeding trial.Compared to T15(21.5% FM,15.45% SBM,15% FSBM),the soybean meal was completely replaced by fermented soybean meal in TF,while AP0 and AP5 were semi-purified diets which contained approximate content of soybean antigenic protein to TF(nearly 0% soybean antigen proteins)and T15(about 5% soybean antigen proteins)respectively and soybean antigen proteins were the sole anti-nutritional factor.Giant freshwater prawn with the initial average body weight of(0.17±0.02)g were cultured in net cages fixed indoor cement tank fed with above four diets for 64 days.Results showed that with the reduce of soybean antigen proteins,trypsin activity decreased in both series of diets,practical diet groups showed significantly decreased growth gain rate but purified diet groups didn't,it proved that 5% soybean antigen protein wouldn't affect the growth of Macrobrachium rosenbergii,while a significant reduction of soybean antigen protein is not conducive to growth in the practical diet.A certain amount of soybean antigen protein can increase trypsin activity to promote the digestion of feed,which in turn has a positive impact on growth.In the practical diet groups,there was no significant difference in serum MDA content and the expression of NF-?B m RNA in gills,activity of SOD in serum was significantly higher in TF,but the expression of Toll receptor in gills and HSP70 m RNA in hepatopancreas were significantly decreased;In the purified diet groups,there was no significant difference in serum and SOD activity,MDA content and the expression of HSP70 m RNA in hepatopancreas,while the expression of Toll receptor and NF-?B m RNA in gills was significantly higher in TF group.The results showed that fermented soybean meal could scavenge free radical through the synthesis of antioxidant enzymes,reduced the risk of oxidative stress and the degree of stress on the hepatopancreas,which may not be directly related to the reduction of soybean antigen protein content.Soybean antigen proteins and a variety of anti-nutritional factors together were likely to stimulate the body's immune system and achieved immune defense against the foreign body of the host by decreasing the Toll receptor and NF-?B m RNA expression levels.The TP in serum was significantly decreased in the practical diet groups,AST activity was significantly increased but TP and ALB were significantly decreased in the purified diet groups.The results proved that this concentration of antigen proteins was more conducive to the health of hepatopancreas,improved the body's amino acid and protein synthesis metabolism.Coupling with a more sensitive immune response to the body,the T15 group showed better growth performance.In conclusion,Macrobrachium rosenbergii has a certain tolerance to 5% of soybean antigen protein and is beneficial to its growth and health.There was a probable beneficial combination effect between soybean antigen protein and other anti-nutritional factors.It was not conducive to protein synthesis and metabolism of hepatopancreas and lead to decreased growth when the content of soybean antigen proteins was markedly lessened in the feed.All these results suggested that fermented soybean meal and soybean meal be mixed together with an appropriate proportion under normal feeding conditions.3.The study on growth,feeding preference and gastricemptying of fermented soybean meal and soybean antigenic protein on giant freshwater prawn(Macrobrachium rosenbergii)This study investigated the effects on growth,feeding preference,protease and gastric emptying of giant freshwater prawn when fish meal and soybean meal in diet were partly replaced by fermented soybean meal.The shrimp with initial weight(6.0±0.15)g were randomly allocated to 8 groups,fed with the above-mentioned 8 isonitrogenous and isoenergetic diets,cultured for 8 weeks.The percentage of wet weight of digesta in stomach was determined by continuous sampling method.Linear,exponential and square root models were used to simulate the emptying process.Food intake statistical test and rare earth element preference test were conducted to study the feeding preference and acclimation effects of giant freshwater prawn.After shrimp with a period of feeding with the mixture of 6 practical diets with unique rare earth element respectively to reach an equal inclusion,the feed preference and 8weeks acclimation effect were esitimated based on the amount of rare earth elements in the feed and faece.The results showed that the highest feeding preference index was observed in T0 group and the lowest in TF group,the feeding preference index decreased gradually with the decrease of the concentration of fermented soybean meal in the feed before and after domestication.High-concentration fermented soybean meal will result in feeding aversion and domestication could enhance the feeding preference with culturing for 8 weeks.The partial replacement of fermented soybean meal with fishmeal and soybean meal did not negatively affect the growth performance of giant freshwater prawn,except TF group.80% gastric emptying time,amylase activity and protease activity appeared the same tendency,namely increased firstly and then decreased with the increasing of fermented soybean meal,and the highest value was found in group T8.The influence on protease and gastric emptying of freshwater prawn,and their different dynamic changes may be related to soybean antigenic protein's single or combinational effect with other anti-nutritional factors.4.Effects of fermented soybean meal and soybean antigen protein on the intestinal morphology and bacterial community of giant freshwater prawn(Macrobrachium rosenbergii)As the substitution of fish meal,plant proteins have already been attracted more attention,however,there is negative effect when the inclusive level of plant proteins is excessive in diets.This test aimed to understand the relation among intestinal morphology and bacterial community of giant freshwater prawn(Macrobrachium rosenbergii),explore the role of fermented soybean meal as alternative of fish meal and soybean meal.The samples used in this experiment was taken after the end of the cultivation in Chapter 3.The result showed that,height of intestine fold and thickness of muscular layer decreased gradually with the decrease of the concentration of fermented soybean meal and T15 group was higher than that of the control group.There was no significant difference in the purified diet groups,while the height of intestine fold of T15 group was significantly higher than that of TF group.Sequencing of bacterial 16 s r RNA V3 region showed that Proteobacteria and Aermomas were the dominant bacteria in the intestine of giant freshwater prawn.CCA analysis showed a positive correlation between the height of intestine fold and Staphylococcus,Lactococcus,but a negative correlation of Acidovorax.Our results suggest that giant freshwater prawn has a great tolerance to 5% of soybean antigen protein in purified diet and did not have a significant negative effect on the intestinal morphology,but didn't showed in practical diet,it may imply a beneficial combination effect between soybean antigen protein and other anti-nutritional factors.It may hurt intestinal healthy and reduce height of intestine fold when the soybean antigen proteins markedly lessened in the practical diet.In addition,the intestinal bacterial population was highly affected by fermented soybean meal,soybean meal and soybean antigen protein,and Staphylococcus,Lactococcus and Acidovorax might play an important role in the height of intestine fold.