Studies on Toxicity of Freshwater Puffer Fish and Genetic Identification of Species Sold in Taiwan-1

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Project title

Code/計畫編號

NSC94-2313-B254-001

Translated Name/計畫中文名

台灣市售淡水河魨之毒性學和種屬基因鑑定法探討-1

Project Coordinator/計畫主持人

Deng-Fwu Hwang

Funding Organization/主管機關

National Science and Technology Council

Co-Investigator(s)/共同執行人

林欣榮

Website

https://www.grb.gov.tw/search/planDetail?id=1097460

Year

2015

Start date/計畫起

01-08-2015

Expected Completion/計畫迄

31-07-2016

Bugetid/研究經費

805千元

ResearchField/研究領域

漁業

近年來台灣觀賞魚業者大量引進淡水性之觀賞河魨，由於淡水河魨之毒性與物種親緣相關資料尚未建立，但在國外已有中毒案件發生，故為建立完整之淡水河魨毒性和種屬資料，本研究乃針對台灣市售觀賞用之淡水河魨，分析其毒性和毒成分，進一步應用生物技術的方式，建立有毒淡水河魨之基因鑑定資料。淡水河魨之毒性，經以河魨毒之生物毒性檢測法測定其皮膚、內臟和肌肉之毒性，得知淡水河魨中 Auriglobus silus ( 綠娃娃魚 ) 、 Carinotetraodon travancoricus (巧克力娃娃魚)、Tetraodon biocellatus (八字娃娃魚) 、 T. nigroviridis (金娃娃魚)及 T. turgidus 等五種類皆屬有毒種，毒性最高者為八字娃娃魚之外皮，毒性為 114.9± 93.1 MU/g。有毒淡水河魨之檢體其毒素經精製後，由 HPLC、GC-MS 和 LC-MS 鑑定皆屬河魨毒及其衍生物。另外，其中 7 種淡水河魨（ Auriglobus silus 、 Carinotetraodon irrubesco、C. lorteti、C. travancoricus、Tetraodon biocellatus、T. nigroviridis 和 T. turgidus 經以一對基因引子 H15149 和 L14735 可大量增幅 mtDNA 之 474 bp 基因片段，經由基因序列分析及親源分析結果其差異度為 2.77 - 29.67%，可區別各種魚種；另使用單一限制酶 Hae III 即可將上述 7 種淡水河魨之 474 bp 基因片段切割成各自不同長度的片段圖譜，顯示限制酶切位可順利區分 7 種淡水河魨之種別。 The purpose of this study is to establish the data of the toxicity and to identify the species of freshwater puffer in Taiwan. In the first part of this study, the toxicity of several freshwater puffers were determined using tetrodotoxin bioassay. It was found that Auriglobus silus, Carinotetraodon travancoricus, Tetraodon biocellatus, T. nigroviridis and T. turgidus contained moderate amounts of toxin. The highest toxicity score was 114.9±93.1 MU/g skin of Tetraodon biocellatus. The toxin was partially purified and analyzed and identified using HPLC, GC-MS and LC-MS showed that the toxin was tetrodoxin . On the other hand, Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) and sequence analysis were performed to establish genetic markers in cytochrome b gene for identifying seven species of freshwater puffer fish genus, including Carinotetraodon lorteti, C. irrubesco, C. travancoricus, Tetraodon nigroviridis, T. biocellatus, T. turgidus and Auriglobus silus. Polymerase chain reaction (PCR) prepared DNA for sequencing. No variation in sequences was detected among individuals within each species. Direct estimates of cytochrome b gene sequence divergence among seven freshwater puffer fishes were from 2.77 to 29.67%. In the restriction enzyme analysis, the restriction enzyme Hinf I could differentiate the species of C. lorteti, C. irrubesco, C. travancoricus and A. silus from seven freshwater puffer fishes. Using Hae III could differentiate the other three freshwater puffer species. Therefore, the polymorphisms obtained from restriction site analysis and sequence analysis have provided usable genetic markers for identifying the seven species of freshwater puffer fish genus.

Keyword(s)

淡水河魨
毒性
基因序列分析
Freshwater puffer fish
toxicity
sequence analysis

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Studies on Toxicity of Freshwater Puffer Fish and Genetic Identification of Species Sold in Taiwan-1

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