| Chinese mitten crab(Eriocheir sinensis)is one of the traditional aquaculture anim als in China.Because of its good flavor and high nutritional value,it's not only regar ded as a delicious food,but also has high medicinal value.Therefore,it is very popul ar in China.The rice-crab breeding model is an efficient ecological planting method t hat has been popularized in recent years.However,the research on rice-crab model is not perfect,and there were still many problems to be solved.Actually,the ammonia i n the environment will increase due to various factors,especially the application of fe rtilizers and even at relatively low concentrations of ammonia stress,which will harm crustaceans and even be fatal.The growth safety and edible safety of Chinese mitten crabs mainly depend on the water where they grow.If there are environmental polluta nts,they may accumulate in crabs.As the key tissue for detoxification and metabolis m,hepatopancrea of Eriocheir sinensis was significantly affected by the environment.The purpose of this study is to determine the substances involved in metabolism and immune process in the hepatopancreas of Eriocheir sinensis,and to understand the infl uence of different ammonia concentration on the nutritional value and growth state of Eriocheir sinensis according to its changing trend,so as to provide theoretical support for the healthy growth and food safety of crustaceans.At the same time,proteomics was used to study the effect of different ammonia concentration on hepatopancreas of Eriocheir sinensis.The results not only help us to verify the change trend of the dete rmined substances at the molecular level,but also provide more research directions an d ideas for us to further understand the impact of ammonia on Eriocheir sinensis.In our experiments,60 mitten crabs were divided into three groups: control group(C,0g/L),low concentration ammonia treatment group(LT,0.04g/L),high concentrati on ammonia treatment group(HT,0.16g/L).Through seven days of simulating the env ironment with different concentrations of ammonia nitrogen,we studied the physiological characteristics of different concentrations of ammonia on juvenile Chinese mitten cr abs in consumption of energy and antioxidant activity of hepatopancrea.In order to fu rther understand the effect of ammonia stress on the hepatopancreas of Eriocheir sinen sis at the molecular level.Conclusion were showed as follow:In the seven-day experiment with different concentrations of ammonia stress,the protein content and ammonia concentration did not show a linear relationship,but fluc tuating;the glycogen content of hepatopancreas showed a decreasing trend with increa sing of ammonia concentration.After high ammonia stress,the accumulation of glycog en in hepatopancreas content was significantly reduced,which means that increase of ammonia concentration would increase the crabs metabolic consumption burden,which will affect the process of glucose metabolism.Prolonged stress might reduce the accu mulation of energy substances and reduce nutrition and flavor,which even increased it s chance of death.The content of pyruvate kinase activity became activated with the i ncrease of ammonia concentration,anaerobic metabolism increases,the energy metaboli sm pathway was affected and the glycolytic metabolism pathway might be activated.The effects of ammonia on the antioxidant system of Chinese mitten crabs were studied by measuring the activities of catalase(CAT),glutathione(GSH)and glutathio ne peroxidase(GSH PX)in hepatopancreas.During the concentration of ammonia stre ss,the activity of catalase(CAT),which was one of the antioxidant enzymes involved in eliminating active oxygen,was significantly inhibited with the increase of ammoni a concentration,which indicateed that the high concentration of ammonia might cause more active oxygen and limited activity of CAT.Both content of GSH and activity of GSH-Px showed a downward trend with the increase of concentration of ammonia,w hich affected its antioxidant activity.In short,the increase of the concentration of am monia may increase the production of active oxygen in the body,inhibit some enzym e activities and the content of active substances in the antioxidant system,resulting in an unbalance between the oxidation and the antioxidant system.In addition,2427 proteins were identified by proteomics,including 112 differentia lly expressed proteins between group C and lt,and 324 differentially expressed protein s(DEP)between group LT and HT(P < 0.05).Differentially expressed proteins are mainly involved in immune processes,such as C-type lectin(CTL),c-Jun N terminal kinase(JNK),and anti-lipopolysaccharide factor(ALF).Changes in glutathione peroxid ase(GPX),glutathione-S-transferase(GST),etc.suggested that ammonia stress may be caused by the destruction of oxidants and antioxidants.Balance,which causes oxidati ve stress,which in turn caused more adverse effects.In addition,we also found many different growth-related proteins,such as amylase,fatty acid synthase,fatty acid bindi ng protein(FABP),actin,etc.This showed that ammonia may affect the short-term gr owth process and related proteins.As the ammonia concentration changed,its expressi on showed a continuous change trend,while some proteins showed opposite changes u nder different concentrations of ammonia.The results of these changes indicated that t he effect of ammonia on Chinese mitten crab was multifaceted and the changes of the expression of some proteins were consistent with the changes in our previous measur ement results in Chapter Two.In addition,7 KEGG pathways were significantly chan ged between the two groups.These pathways were related to cell endocytosis,RNA tr ansport and immune function.In conclusion,compared with LT group,the concentration of ammonia in HT gro up has more significant effect on metabolism,immune system and other pathways of Eriocheir sinensis.In addition,we also got similar conclusion through proteomic meth od.Higher concentration of ammonia will not only increase the expression of innate i mmune related proteins,but also reduce the expression of related proteins involved in antioxidant process and production process.At the same time,we also found more pr oteins with different expression due to the influence of ammonia concentration,which is beneficial to us to fully understand the stress mechanism of ammonia... |
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