Effects Of Cottonseed Meal Protein Hydrolysate On Feeding Attraction,Growth And Immunity Of Chinese Mitten Crab,Eriocher Sinensis

In recent years,with the rapid development of Chinese mitten crab,Eriocheir sinensis industry,the research and application of artificial formulated feed will become an inevitable trend in the process of artificial culture.But as a result of the feed palatability and the behavior habit of crab resulting in eating food speed slowly,it is easy to cause feed waste and water pollution,and result in a reduction in disease resistance.Therefore,it is critical for us to look for the right solution.This experiment is conducted to find used in crab attractant and explore its effects on growth,feed intake and immunity of Chinese mitten crab.The experiment included three parts:Experiment 1:Study on the preference of five attractants for Chinese mitten crab,Eriocher sinensisThis study was conducted to screen the attractants for Chinese mitten crab,Eriocheir sinensis by pigment indication method.160 crabs（average initial weight:26.4±0.17 g）were equally divided into 20 concrete tankages.First,the basic feed was added pigment of different colors,taking the same number of particles for each color and put it into a specific area of the same concrete tankage.The aimed was to explor wheather different colors of basic feed affected the feed intake of crabs;then,we elected basal diet as control.Based on basal diet,the levels of supplemented with squid extract（SE）were 0.3%、0.6%、0.9%and 1.2%;the levels of supplemented with yeast hydrolysate（YH）were 0.2%、0.4%、0.6%and 0.8%;the levels of supplemented with cottonseed meal protein hydrolysate（CPH）were 0.2%、0.4%、0.6%and 0.8%;the levels of supplemented with betaine were 0.25%、0.5%、0.75%and 1%;the levels of supplemented with allicin were 0.03%、0.06%、0.09%and 0.12%.The feed of different levels of the same attractant were added pigment of different colors,taking the same number of particles for each color and put them into a specific area of the same concrete tankage.The aim was to elect the optimum supplement levels of five attractants;finally,we elected the optimum supplement levels of five attractants and then added pigments of different colors,using the same method to compare the preference of different attractants on crabs.The results showed that（1）There were no significant differences among the stimulant index of fed basic diet of different colors（P>0.05）;（2）The stimulant indexs of fed 0.3%SE,0.6%YH,0.6%CPH and 0.09%allicin diets were significantly（P<0.05）higher than that of the control diet;The highest stimulant index of betaine was observed in 1%group,but there was no significant difference among other groups（P>0.05）;（3）The highest stimulant index was observed in 0.6%CPH group,the stimulant indexs of 0.6%CPH and 0.6%YH diet were significantly（P<0.05）higher than those of the other groups;the stimulant index of 0.3%SE diet was significantly（P<0.05）higher than that the 1%betaine diet.In conclusion,there were no significant differences among the colors of feed on feeding of crabs;the optimum supplement levels of SE,YH,CPH,betaine and allicin for crabs were 0.3%,0.6%,0.6%,1%and 0.09%orderly,and diets preference were following:CPH>YH>SE>allicin>betaine.The 0.6%CPH or 0.6%YH had stronger attractive effect among the attractants,which can be used as attractants of crabs.Experiment 2:Effects of cottonseed meal protein hydrolysate on growth,feed intake and regulation of Chinese mitten crab,Eriocheir sinensisThis study was conducted to investigate the effects of dietary CPH on growth,feed intake,digestion ability and absorption function and appetite of Chinese mitten crab,Eriocheir sinensis.160 crabs（average initial weight:51.32 ± 0.08 g）were fed four experimental diets with CPH at 0%,0.3%,0.6%and 1.2%.Each diet was tested in five replicates for 60 days.The results showed that no difference（P>0.05）were observed in weight gain rate,specific growth ratio and feed efficiency ratio in crabs fed CPH diet compared to those fed the control diet,but dietary CPH significantly improved（P<0.05）the 1-hour feed intake.Moisture content was significantly reduced in crabs fed CPH0.6 diet compared to the control group.Protease activity of crabs fed CPH0.6 diet was significantly（P<0.05）higher than that of the control group whereas the difference was not significantly between the CPH0.3 and CPH0.6 groups（P>0.05）.Lipase,amylase,and Na+-K+ATPase activities were not significantly affected（P>0.05）among the CPH0,CPH0.3 and CPH0.6 groups.In thoracic ganglia,no significant different was observed in target of rapamycin（TOR）expression level.4E-binding protein（4E-BP）expression level of crabs fed CPH0.6 diet was significantly（P<0.05）higher than that of the control group.In cerebral ganglia,protein kinase B（Akt）and 4E-BP expression levels of crabs fed CPH0.6 diet was significantly（P<0.05）lower than that of the control group.In mid intestine,TOR and Akt relative expression level of crabs fed the diets containing CPH was significantly（P<0.05）lower than that of the control group.4E-BP expression level showed an opposite pattern.In hepatopancreas,TOR,Akt and 4E-BP expression level decreased as dietary CPH increase.Leptin receptor（LEPR）relative expression level of crabs fed CPH0.6 diet in thoracic ganglia and cerebral ganglia was significantly（P<0.05）lower than those of the other groups.In mid intestine,LEPR and cholecystokinin receptor relative expression level of crabs fed CPH0.3 and CPH0.6 diets was significantly（P<0.05）lower than that of the control group.The highest neuropeptide Y（NPY）expression level was observed in crabs fed CPH0.6 diet,and it was significantly（P<0.05）higher than CPH0 group in different tissues.The relative expression level of neuropeptide Y receptor presented the similar trend to that of NPY.In conclusion,our results indicated that supplementation of dietary CPH could stimulate appetite,improve the activity of digestive enzyme and quicken feeding rate via the TOR signaling pathway.Experiment 3:Effects of dietary cottonseed meal protein hydrolysate on antioxidants and immunity of Chinese mitten crab,Eriocheir sinensisThis study was performed to evaluate the effects of dietary CPH on antioxidants and immunity of Chinese mitten crab,Eriocheir sinensis.The experimental design and feeding trial was the same to the experiment 2.The results showed that superoxide dismutase and catalase activities of crabs fed CPH0.6 diet were significantly increased（P<0.05）whereas the difference was not significantly between the CPH0.3 and CPH0.6 groups.Malondialdehyde content of crabs fed CPH0.3 diet was significantly decreased（P<0.05）than that of the control group.Lysozyme and alkaline phosphatase activities of crabs fed CPH0.3 diet were significantly higher（P<0.05）than those of the other groups.Acid phosphatase activity of crabs fed CPH0.3 diet was significantly higher than that of the control group（P<0.05）.Glutamic-pyruvic transaminase activity of crabs fed CPH0.3 diet was significantly decreased compared to the control group（P<0.05）,glutamic-oxalacetic transaminease activity was no significantly difference among all groups.The relative expression levels of toll-like receptor 2（Toll2）,myeloid differentiation factor 88（MyD88）,lipopolysaccharide-induced TNF-α factor（LITAF）and interleukin-2 enhancer binding factor（ILF-2）of crabs fed CPH0.3 diet were significantly higher（P<0.05）compared to those fed other diets,the relative expression level of toll-like receptor 1（Toll1）of crabs fed CPH0.3 diet was significantly higher than that of the control group（P<0.05）.The expression level of suppressors of cytokine signaling 2（SOCS2）showed an opposite pattern.After acute CPH injection,the relative expression levels of Toll1,Toll2,MyD88,LITAF and ILF-2 in intestine and hepatopancreas were all significantly（P<0.01）affected by time,injection type and their interaction;the expression level of SOCS2 was significantly（P<0.001）affected by time and the interaction of time and injection type.The expression levels of SOCS2 and Toll1 in intestine at time 3 h and SOCS2 in hepatopancreas at time 18 h increased significantly to the highest value.The expression level of Toll2,MyD88,LITAF decreased at times 6 h,6 h,12 h,respectively,then increased gradually until basal level.The results of this study indicated that supplementation of dietary CPH could improve antioxidant capacity and immune function;the appropriate supplement dosage of CPH for crabs could be 0.3-0.6%.Furthermore,the short-term CPH stimulation could significantly affected the expression levels of immune-related genes.