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研究生:邱顯智
研究生(外文):Chiu, Sain-Jhih
論文名稱:臺中地區異形吸蟲科囊狀幼蟲之感染研究
論文名稱(外文):Infectivity Study of the Heterophyidae Metacercaria in Taichung Area
指導教授:李明憲李明憲引用關係
指導教授(外文):Li, Ming-Hsien
口試委員:范家堃廖朝財
口試委員(外文):Fan, Chia-KwungLiao, Chao-Tsai
口試日期:2014-06-30
學位類別:碩士
校院名稱:中臺科技大學
系所名稱:醫學檢驗生物技術系碩士班
學門:醫藥衛生學門
學類:醫學技術及檢驗學類
論文種類:學術論文
論文出版年:2014
畢業學年度:102
語文別:中文
論文頁數:121
中文關鍵詞:異形吸蟲科囊狀幼蟲溪流盛行率鑑定
外文關鍵詞:Heterophyidaemetacercariaestreamprevalenceidentification
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異形吸蟲科(Heterophyidae)是寄生於小腸的魚源性蠕蟲。寄生在人體內會導致非細菌性的腹瀉、腹痛或是因為異位感染所造成的腦膜炎以及感染性肉芽腫,寄生在魚類的鰓上則容易影響呼吸功能而死亡,這些症狀也稱為異形吸蟲症(Heterophyiosis)。在東南亞地區,異形吸蟲症的盛行影響著許多養殖業者以及食用淡水魚的民眾。在臺灣過去曾有自淡水魚體內分離出中華肝吸蟲(Clonorchis sinensis)及橫川吸蟲(Metagonimus yokogawai)等吸蟲類之囊狀幼蟲的相關研究報導,但是時間已久,因此本研究的目的主要調查臺中地區的三條溪流,異形吸蟲科之囊狀幼蟲時期在魚群間的盛行率,以瞭解目前異形吸蟲科的流行現況。本研究自2012年9月至2014年1月,針對臺中地區三條溪流,大里溪、旱溪以及大甲溪;採集九種淡水魚類,平頜鱲(Zacco platypus) 、粗首馬口鱲(Opsariichthys pachycephalus) 、臺灣石賓(Acrossocheilus paradoxus) 、臺灣馬口魚(Candidia scale barbata)、臺灣鏟頷魚(Varicorhinus barbatulus)、鯉魚(Cyprinus carpio carpio)、鯽魚(Carassius auratus auratus)、蝦虎(Gobiida)以及吳郭魚(Oreochromis mossambicus)共524尾,以人工胃液消化處理,沉降後在解剖顯微鏡下觀察並收集囊狀幼蟲,接著分析盛行率。最後再以SPSS統計軟體進行統計分析。結果顯示,大里溪、旱溪與大甲溪的魚群囊狀幼蟲盛行率分別為57.2%(178/311)、10.7%(13/121)與2.2%(2/92)。大里溪的魚群囊狀幼蟲盛行率高於其他兩條溪流並具有顯著性差異(P<0.05)。九種淡水魚的囊狀幼蟲盛行率分別為粗首馬口鱲62.7%(74/118)、平頷鱲44.8%(77/172)、臺灣馬口魚33.3(18/54)、臺灣石賓26.9%(21/78)、吳郭魚5.9%(1/17)、臺灣鏟頷魚3.3%(2/61)、鯉魚0%(0/14)、蝦虎0%(0/8)及鯽魚0%(0/2)。此外,在大里溪所採集的粗首馬口鱲則是擁有最高的囊狀幼蟲盛行率71.1%(64/90) (P<0.05)。最後,從魚體中共分離出四種囊狀幼蟲,其中有兩種可利用型態學及分子鑑定的方法鑑別出臺中異形吸蟲(Haplorchis taichui)以及臺灣異形吸蟲(Centrocestus formosanus)。本論文研究結果發現,大里溪的魚群囊狀幼蟲盛行率是三條溪流中最高的,而粗首馬口鱲則是在所有淡水魚類中囊狀幼蟲盛行率最高的。另外,我們可以從臺中地區的淡水魚類身上分離出臺中異形吸蟲與臺灣異形吸蟲的囊狀幼蟲。
Heterophyidae is a family of the fish-borne helminths that parasitizes in the small intestine. In human, it causes diarrhea and abdominal pain or meningitis and infectious granuloma caused by ectopic infection. In the gills of fishes, it affects respiratory function and cause death. Those symptoms are also known as heterophyiosis. In Southeast Asia, heterophyiosis affects many fish farmers and people who eat fish. In the past, there had reports about metacercariae of Clonorchis sinensis and Metagonimus yokogawai were isolated from freshwater fishes in Taiwan, but for a long time has passed. In this study, we investigate the prevalence of heterophyidae metacercariae infection in fishes from three rivers in central Taiwan to clarify the present prevalence status of Herterophyidae. From September 2012 to January 2014, for three rivers in Taichung area, Dali River, Han River and Dajia River, a total of 524 freshwater fish comprising of nine species such as Opsariichthys pachycephalus, Zacco platypus, Candidia scale barbata, Acrossocheilus paradoxus, Oreochromis mossambicus, Varicorhinus barbatulus, Cyprinus carpio carpio, Gobiidae and Carassius auratus auratus. Fish were individually grinded by the artificial digestive fluid method. Then, the sediment observed under a dissection microscope to check for the metacercariae. Statical analyses were performed by using SPSS software. Our result suggest that prevalence of metacercaria of Dali River, Han River and Dajia River was 57.2% (178/311), 10.7% (13/121) and 2.2% (2/92). Dali River had higher prevalence than others (P<0.05).The prevalence of metacercaria of nine species freshwater fish was Opsariichthys pachycephalus of 62.7% (74/118), Zacco platypus of 44.8% (77/172), Candidia scale barbata of 33.3% (18/54), Acrossocheilus paradoxus of 26.9% (21/78), Oreochromis mossambicus of 5.9% (1/17), Varicorhinus barbatulus of 3.3% (2/61), Cyprinus carpio carpio of 0%(0/14), Gobiidae of 0% (0/8) and Carassius auratus auratus of 0% (0/2). In addition, Opsariichthys pachycephalus of Dali River had the highest prevalence of 71.1% (64/90) (P<0.05). Finally, four kinds of metacercariae were isolated from fishes, two of which identify of Haplorchis taichui and Centrocestus formosanus was based on morphological and molecular identification. The highest prevalence of metacercaria was found in Dali River and Opsariichthys pachycephalus has the highest prevalence of metacercaria in all freshwater fish. Additionally, Haplorchis taichui and Centrocestus formosanus can be isolated from freshwater fish in the stream of Taichung.
摘要 I
Abstract III
縮寫字對照表 V
目錄 VI
表目錄 IX
前言 1
1 背景 1
2 異形吸蟲科之基本介紹 2
2.1 地理分佈與分類學 2
2.2 寄生宿主與生活史 3
2.3 各階段之形態學 4
2.3.1 蟲卵 4
2.3.2 纖毛幼蟲 4
2.3.3 胞狀幼蟲 5
2.3.4 雷氏幼蟲 5
2.3.5 尾動幼蟲 5
2.3.6 囊狀幼蟲 5
2.3.7 成蟲 6
2.4 臨床症狀 6
3 文獻回顧 7
4 研究動機與目的 8
5 採集地點溪流之介紹 8
5.1 大甲溪 8
5.2 烏溪 9
5.2.1 大里溪 9
5.2.2 旱溪 10
材料與方法 11
1 實驗材料及儀器 11
1.1 採集用器材 11
1.2 實驗用器材 11
1.2.1 材料 11
1.2.2 儀器 12
1.2.3 試藥套組 13
1.2.4 試劑及配方 13
2 檢體來源及採集 14
2.1 淡水螺螄 14
2.2 淡水魚類 15
2.3 實驗動物 15
3 方法 15
3.1 淡水螺螄之檢查-壓碎法 15
3.2 淡水魚類之檢查 16
3.2.1 鱗片與鰭之檢查-直接鏡檢法 16
3.2.2 鰓之檢查-壓片法 16
3.2.3 魚體之檢查-人工胃液消化法(artificial digestive fluid method) 17
3.3 實驗動物之感染 17
3.3.1 糞便檢查-福馬林乙醚沉澱法(formalin-ethyl concentration) 18
3.3.2 成蟲回收 19
3.3.3 蟲體的觀察 19
3.4 分子生物學鑑定 21
3.4.1 DNA萃取 21
3.4.2 測定DNA濃度及純度 22
3.4.3 聚合酶連鎖反應(Polymerase Chain Reaction) 23
3.4.4 瓊脂膠體電泳分析(Agarose gel electrophoresis) 25
3.4.5 核酸定序(DNA Sequencing) 26
4 統計分析 26
4.1 T 檢定 (student’s t test) 26
4.1.1 魚類大小與囊狀幼蟲感染之差異 26
4.2 卡方檢定 (chi-square test) 26
4.2.1 各溪流之魚群囊狀幼蟲盛行率之差異 26
4.2.2 魚群囊狀幼蟲在魚體與鰓部之盛行率 27
4.2.3 海拔差異與囊狀幼蟲感染之差異 27
結果 28
1 淡水螺螄檢查結果 28
2 淡水魚類檢查結果 29
3 異形吸蟲形態學鑑定 31
3.1 囊狀幼蟲之形態學 31
3.2 實驗動物感染 31
3.3 成蟲之形態學 32
4 分子生物學鑑定 32
5 統計分析 33
5.1 T 檢定 (student’s t test) 33
5.1.1 魚類大小與囊狀幼蟲感染之差異 33
5.2 卡方檢定 (chi-square test) 33
5.2.1 各溪流之魚群囊狀幼蟲盛行率之差異 33
5.2.2 魚群囊狀幼蟲在魚體與鰓部之盛行率 34
5.2.3 海拔差異與囊狀幼蟲感染之差異 34
討論 36
1 淡水螺螄以及淡水魚類採集與檢查 36
2 異形吸蟲形態學及分子生物學鑑定 38
3 統計學分析 39
3.1 魚類大小與囊狀幼蟲感染之差異 39
3.2 各溪流之魚群囊狀幼蟲盛行率之差異 39
3.3 魚群囊狀幼蟲在魚體與鰓部之盛行率 40
3.4 海拔差異與囊狀幼蟲感染之差異 41
表 42
圖 54
附錄 100
參考文獻 103

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