- 學位論文
白點症病毒感染後白蝦血淋巴液中血糖、乳酸、三酸甘油酯分析與烯醇酶、醛縮酶、3-磷酸甘油醛脫氫酶及基因表現之分析
Analysis on the changes of hemolymph glucose, lactate and triglycerides, and the gene expressions of enolase, aldolase and glyceraldehyde 3-phosphate dehydrogenase in white shrimp, Litopenaus vannamei under WSSV infection
摘要
白點症病毒是一具有高侵襲性的蝦類病原體，俟至目前，有關於此病毒與宿主之間交互作用的研究仍極為有限。本研究首先分析病毒感染對白蝦血淋巴液中血糖、乳酸和三酸甘油酯的影響。結果發現，血淋巴液中乳酸量及三酸甘油酯量，於感染後逐漸下降，但血糖量卻在感染後呈現上揚之趨勢，12小時上升約1.74倍，推論白點症病毒藉由促進乳酸分解以利糖質新生作用進行，使得宿主血糖含量上升。本實驗也進一步以二維膠體電泳分析白點病毒感染白蝦胃部組織的蛋白質表現變化，觀察到烯醇酶 (Enolase)、醛縮酶 (Aldolase)、3-磷酸甘油醛脫氫酶 (Glyceraldehyde 3-phosphate dehydrogenase, GAPDH) 蛋白質含量確有明顯著的上升，並用西方轉印法分析烯醇酶，亦證實烯醇酶的蛋白質時序表現量亦有增加。 藉由比對白點症病毒感染前後草蝦表現序列標誌 (Expressed sequence tag, EST) 資料庫後，發現烯醇酶、醛縮酶、3-磷酸甘油醛脫氫酶基因表現量於病毒感染後上升。利用反轉錄酶-聚合酶鏈反應 (reverse transcriptase polymerase chain reaction, RT-PCR) 進行這三個基因在受到病毒感染蝦胃部組織的表現分析，亦證明證明這些基因的轉錄表現於病毒感染後的呈現明顯上升的趨勢。 本研究証實，白點症病毒感染蝦體後，會影響蝦類的醣類代謝途徑，促使血淋巴液中血糖的上升、乳酸及三酸甘油酯的下降，胃部組織細胞烯醇酶、醛縮酶、3-磷酸甘油醛脫氫酶基因表現的基因表現亦有變化，而能量代謝產物與醣類代謝相關酵素之相互關連性和其生理重要性，仍後續研究加以釐清。
並列摘要
White spot syndrome virus (WSSV) is known to be a virulent pathogen to penaeid shrimp. To date, information on the interaction between WSSV and hosts are still limited. The present study analyzed the temporal changes in the hemolymph glucose, lactate and triglyceride of the white shrimp under WSSV infection. The hemolymph lactate and triglycerides decreased after the challenge, while the hemolympyh glucose showed an increasing trend, peaked at 12 hr post-challenge, 1.74-folds increase over the control. The observations suggest that the hyperglycemic response of the shrimp is consequenced partly from the remetabolism of lactate in the process of glyconeogenesis. The protein expressions in the gastric tissue of the shrimp were further performed by 2-D gel electrophoresis, and the contents of enolase, aldolase and glyceraldehydes 3-phosphate dehydrogenase (GAPDH) were all markedly elevated. The increasing trend of the temporal expression of enolase also demonstrated by West-blotting method. The gene expression of enolase, aldolase, and GAPDH were notably increased after the WSSV infection to tiger shrimp, indicated in the differential display of expressed sequence tag of the species. These changes are further confirmed in the present study using the method of reverse transcriptase polymerase chain reaction (RT-PCR). It is therefore concluded that the carbohydrate metabolism and the expressions of enolsse, aldolase, and GAPDH genes in the gastric tissues are all influenced by WSSV infection. However, inter-relationship of the energy metabolites and the activities of carbohydrate metabolism-related enzymes, and its physiological significance remain for further elucidation.
參考文獻
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