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研究生:丁彥文
研究生(外文):Yen-Wen Ting
論文名稱:應用反轉錄聚合酶鏈反應技術檢測鴨小RNA病毒及台灣地區鳥禽血清學調查
論文名稱(外文):The Detection of Duck Picornavirus by Reverse Transcription Polymerase Chain Reaction and Serological Survey among Bird Species in Taiwan
指導教授:蔡向榮蔡向榮引用關係
指導教授(外文):Hsiang-Jung Tsai
學位類別:碩士
校院名稱:國立臺灣大學
系所名稱:獸醫學研究所
學門:獸醫學門
學類:獸醫學類
論文種類:學術論文
論文出版年:2004
畢業學年度:92
語文別:中文
論文頁數:70
中文關鍵詞:鏈反應反轉錄聚合酶血清學調查鴨小病毒鴨病毒性肝炎
外文關鍵詞:serological surveyduck virus hepatitisduckreverse transcription polymerase chain reactionduck picornavirus
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鴨病毒性肝炎(DVH)在小鴨是一種高死亡率的疾病,可分為三型,其中第1、3型病原為小RNA病毒(picornavirus),台灣分別於1972及1990年曾爆發DHV大流行,在本研究中,針對1990年在高雄分離的Tw90A株鴨小RNA病毒(duck picornavirus; DPV)之3D核酸部位設計特異性引子,建立DPV之Nested-RT-PCR之快速診斷技術,經檢測其敏感性可達100.025 TCID50,口服接種感染無DPV抗體之一日齡北京鴨,接種後定期採集檢體,並以Nested-RT-PCR偵測DPV特異性核酸片段,結果發現DPV於接種後8小時到48小時於小鴨體內呈全身性分佈,直至接種後21天尚可由小腦、食道、心肌、腿肌、肝臟、脾臟、腎臟、胰臟等器官檢測出病毒核酸反應,其中以肝、脾、心肌、腿肌、腎臟等檢體之反應訊息最強。小鴨接種DPV其增重明顯小於控制組 (p<0.01)。調查台灣多種鳥禽之DPV中和抗體分布情形,其抗體陽性率分別為鴨32.95%、雞1.50%、鴿22.50% 和鵝7.06%。而鴨隻陽性率顯著高於其他鳥禽類 (p<0.01)。而vDHV及DHV之分別為1.1%和62.3%,DHV鴨隻抗體陽性率顯著高於DPV及vDHV。

Duck viral hepatitis (DVH) is a high mortality disease in ducklings. Three types of DHV have been reported, and type 1 and type 3 caused by viruses belonging to Picornavidiae. In Taiwan, outbreaks of DVH were reported in 1972 and 1990, respectively. A strain of the duck pricornavirus (DPV), Tw90A, isolated in the 1990 Kaohsiung outbreak was studied. Two pairs of primers were designed based on the conserved region of 3D and were used in nested reverse transcription polymerase chain reaction (Nested-RT-PCR) to detect DPV. Day-old to 21 days-old Peking ducks without DPV antibody were inoculated with Tw90A isolate. Various organs were collected from 8 hours to 21 days after inoculation and Nested-RT-PCR was used to detect the presence of the DPV genome. The results showed that DPV persisted in the infected ducklings for at least 21 days. Most of the other organs, such as the liver, spleen, heart muscle, skeletal muscle, and kidney, became are tested positive in samples collected between 8 to 48 hours after inoculation. The detection of the DPV genome was limited to the cerebrum, esophagus, heart muscle, skeletal muscle, liver, kidney, spleen, and pancreas at 21 days post-inoculation (DPI). The weight gains of duckling inoculated with DPV were significantly lower than the un-inoculated control ducklings (p<0.01). Seroprevalence of DPV among bird species in Taiwan was also studied. The results showed positive rates of 32.95% in duck, 1.50% in chicken, 22.50% in pigeon, and 7.06% in goose. DPV positive rate was significantly higher in duck than in other species (p<0.01). Seroprevalence of vDHV and in duck were 1.1% and 62.3% respectively. DHV positive rate was significantly higher in duck than DPV and vDHV (p<0.01).

致謝………………………………………………………………ii
中文摘要……………………………………………………iii
英文摘要………………………………………………………...iv
目次………………………………………………………………..vi
表次……………………………………………………………..vii
圖次……………………………………………………………..viii
第一章 緒言……………………………………………………..1
第二章 文獻回顧
第一節 Picornavirus簡介…………………………………….2
第二節 鴨病毒性肝炎簡介………………………………………..3
第三節 鴨病毒性肝炎2型簡介……………………………………9
第四節 鴨病毒性肝炎3型簡介………………………………11
第三章 材料與方法
第一節 病毒定性試驗……………………………………14
第二節 設計RT-PCR檢測引子………………………….16
第三節 DPV台灣分離株動物人工感染試驗………………….18
第四節 DPV、DHV及vDHV台灣分離株之血清抗體調查…………20
第四章 結果
第一節 病毒增殖與定性………………………………………..22
第二節 Nested-RT-PCR檢測引子之建立………………………..22
第三節 DPV台灣分離株動物人工感染試驗…………………….24
第四節DPV、DHV及vDHV台灣分離株之血清抗體調查…………25
第五章 討論……………………………………….....27
表、圖…………………………………………. ……....31
參考文獻…………………………………………………. 59
附錄…………………………………………………. 67


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