| (1)Phagocytosis of the coelomic fluid from Lianjiang and Dalian population of the sea cucumber with the relation with different water temperature(15℃,18℃,21℃ 24℃ 27℃ and 30℃)were studied by microscope.The results showed that coelomocytes had the abilities to phagocytize Staphylococcus aureus and Vibrio anguillarum,and the abilities of phagocytosis were correlated to the kind and size of the foreign bodies,and the abilities of phagocytosis were correlated to the varieties of the sea cucumber.The peak phagocytic percent at 21℃.In the assay of phagocytizing S.aureus,the Lianjiang population of the sea cucumber maximal phagocytosis(59.84%)and Dalian population of the sea cucumber maximal phagocytosis(58.48%)was observed 6 h;In the assay of phagocytizing V.anguillarum,the Lianjiang population of the sea cucumber maximal phagocytosis(43.64%)was observed 4 h,and the Dalian population of the sea cucumber maximal phagocytosis(38.66%)was observed 6 h.The phagocytic percentage of coelomocytes in Lianjiang population were higher than Daliang population of A.japonicus(P<0.05).The phagocytic percentage of coelomocytes on S.aureus was higher than V.anguillarum in Lianjiang and Dalian population of the sea cucumber.The phagocytic percentage and phagocytic index of coelomocytes after injection two bacteria had same change trend in Lianjiang and Dalian population of the sea cucumber.(2)After exposed under different sea water temperature,the activities of Superoxide dismutase,Catalase,Glutathione peroxidase,Lactate dehydrogenase,Alkaline phosphatase and the content of Malondialdehyde of the coelomic fluid and extracting fluid of visceral mass from Lianjiang and Dalian population of the sea cucumber,A.japonicus,were determined.The results showed that the temperature of highest activities of SOD,CAT,GSH-PX and LDH of extracting fluid of visceral mass from Lianjiang population were increased 3-9℃ than Daliang population of A.japonicus;under high temperature(24℃,27℃,30℃)stress,the activities of SOD,CAT and LDH of coelomic fluid and extracting fluid of visceral mass from Lianjiang population were higher than Daliang population of A.japonicus(P<0.01),the activities of GSH-PX of extracting fluid of visceral mass from Lianjiang population were higher than Daliang population of A.japonicus(P<0.01),and under 24℃ and 30℃,the activities of GSH-PX of coelomic fluid from Lianjiang population were also higher than Daliang population of A.japonicus(P<0.01);the temperature of highest activity of AKP of coelomic fluid and extracting fluid of visceral mass from the two populations of A.japonicus were all at 15℃ and 18℃,respectively;the content of MDA of the coelomic fluid and extracting fluid of visceral mass from Lianjiang population were significantly lower than Dalian population of A.japonicus at 18-30℃(P<0.05,P<0.01).(3)After exposed under different sea water temperature(15℃、24℃,27℃and 30℃),the gene hsp70 expression of Lianjiang and Dalian population of the sea cucumber,A.in response to the normal temperature and different stress temperature had analyze.The sea cucumbers were exposed to different high temperatures for 2 h,followed by 1 h recovery at 15℃,and the levels of hsp70 were measured using real-time quantitative PCR.The results showed that the expression of hsp70 had the highest expression at 30℃ of Lianjiang and Dalian population of the sea cucumber;at 27℃ and 30℃,the levels of hsp70 of Lianjiang population were higher than Daliang population of A.japonicus(P<0.01).Heat stress could accelerate the gene expression of hsp70,with the increase of temperature the expression of hsp70 mRNA elevated in Lianjiang and Dalian population of the sea cucumber.The phagocytosic assay showed that the coelomocytes’ abilities to phagocytize of the second generation of Lianjiang population of A.japonicus which exposed at high temperature during summer in Fujian province were higher significantly than Daliang population of A.japonicus from northeast China;The experiment of high temperature stress indicated that activities of SOD,CAT,GSH-PX and LDH of coelomic fluid and extracting fluid of visceral mass from the second generation of Lianjiang population of A.japonicus which exposed at high temperature during summer in Fujian province,southeast China,were higher significantly than Daliang population of A.japonicus from northeast China;contents of MDA from Lianjiang population were lower significantly than Daliang population of A.japonicus.The expression of hsp70 mRNA elevated were increased significantly of Lianjiang and Dalian population of the sea cucumber.The increasing of enzymes thermostability of the sea cucumber could be favorable to clean out excess free radicals of the organism under high temperature stress,and to protect the organism from damage caused by high temperature stress.Difference of heat resistance can be reflected by expression of heat shock protein in A.japonicus.It is showed that the adaptability of A.japonicus next generation group domesticated by high temperature was improved to high temperature stress in summer of Fujian Lianjiang,and the ability of the body to deal with the high temperature environment has been improved. |
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