Improving Effects Of Astragalus Polysaccharides (APS) On Non-specific Immune Responses And Productive Performance Of Sea Cucumber (Apostichopus Japonicus)

The use of antibiotics and chemotherapeutics to combat sea cucumber infectious deseases has the risk of generating resistant pathogens,bioaccumulation and environmental pollution. Polysaccharides have been proved as a promising alternative to the drugs,chemicals and antibiotics currentenly to control desease because of more effective and fewer side effects. Astragalus polysaccharides(APS) has been reported to strengthen the defence system through different mechanisms and regulate the intestinal micreoflora to improve the productive performance and intestinal immunity of the fish and other vertebrate.However,the application of APS in sea cucumber,Apostichopus japonicus has not been investigated. Therefore,the application effects of APS were evaluated in vitro and in vivo.In the research,sea cucumbers elder than one year were used as donors and the effects of APS on non-specific immunity and productive performance were studied.The contents and results in detail were stated as follows:The effect of APS on gene expression of lysozyme-i in the coelmocytes was determined using semiquantitative reverse transcription-polymerase chaim reaction(RT-PCR) analysis in vitro.Lysozyme-i mRNA level increased in a dose-dependent manner in the coelmocytes following APS injection.A significant increase was observed with the dose of 5 mg/kg,and hige dose of APS(20 mg/kg) had no effect on lysozyme-i mRNA expression.Effect of APS on cell functions of phagocytic amoebocytes(PA) was studied in vitro. Phagocytosis and superoxide anion(O₂^-) production by PA from sea cucumber coelomic fluid were measured during incubation at 18℃,22℃,or 25℃with APS at different concentration. It suggested that immunocytes activity in sea cucumber decreased with the temperature increasing from 18℃to 25℃,and APS could be an effective immunostimulant to enhance phagocytic activity and O₂^- production.Compared with controls,including APS at 20μg/ml significantly increased the percentage of phagocytic capacity(PC) and phagocytic index(PI) at 18℃and 22℃,The concentration of 20μg/ml APS resulted in a significant increase in nitroblue tetrazolium(NBT) positive cells at different temperature,and when APS at 40μg/ml, they were all decrease.To further study the practicle application of APS in sea cucumber culture,oral administration of APS and the herbal medicine Astragalus membranaceus with different treatment were studied.Three ways were applied in processing A.membranaceus to make general grind powder(GP),superfinc powder(SP),and mechanochemical processed superfine powder(MP).Phagocytic capacity(PC),superoxide anion(O₂^-) production and the activities of phenoloxidase,lysozyme,acid phosphatase(ACP),alkaline phosphatase(ALP), lectin titer in serum were investigated 20,40 and 60 days after feeding,respectively.The activities of PC,ACP and O₂^- production significantly increased after 20 days of feeding with MP or APS,while lysozyme activity and lectin titer in APS group were increased significantly.After 40 days feeding,a significant enhancement of lysozyme and ACP activities in MP group were found,and a significant increase in the activities of PC,lysozyme, ALP and lectin titer were showed in APS group.On the day of 60,GP,SP and MP groups showed evident enhancement in lysozyme and ACP activities.Lectin titer in MP and APS groups was increased significantly.When challenged with Hbrio splendidu after 60 days feeding,the cumulative symptom rates in MP and APS groups were decreased by 83.33%and 75%compared to the controls.Productive performance was also measured in feeding trial.The results showed that SGR of groups MP and APS was higher than that of control group with an increase of 30.03%and 20.94%,respectively.There were no differences of viscera body wall ratio and nutrient contents of body walls between the trial group and control group.A significant enhancement of protease activity in groups of SP,MP,APS and alginase activity in groups of MP,APS were measured at the day of 40 and 60.In addition,an evident increase of amylase activity was showed in groups of GP,MP and APS after 40 days feeding.After oral administration of APS,the proportion of alginate producing stain in the bacterial populations in the intestinal walls showed two times of the other groups.In intestinal content, the proportion of alginate producing stain was 10 or even 100 times more than those of the other groups.In vitro,the addition of APS with the concentration of 0.1%in the medium could significantly increase the alginase production and the biomass of the stain compared to 0.05%and 2.0%groups.Consequently,the data indicate that APS improve immunity against pathogenic bacteria and increase productive performance of A.japonicus.It can be used as immunostimulant in sea cucumber culture.