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年度	104	專案性質	實驗性質
專案類別	研究專案	研究主題	調查
申請機構	國立成功大學	申請系所	環境工程學系
專案主持人	吳哲宏	職等/職稱	副教授
專案中文名稱	地下水污染環境法醫: 發展高通量微生物偵測技術
中文關鍵字	環境法醫、多氯乙烯、高通量微生物偵測技術、生物復育
專案英文名稱	Environmental Forensics for Groundwater Pollution Using High-throughput Microbiology Detection as a Toolbox
英文關鍵字	Environmental Forensics, Polychlorinated Ethene, High-throughput Biotechnology, Bioremediation
執行金額	1,250,000元
執行期間	2014/12/1 至 2015/11/30
計畫中文摘要	國內土水污染的環境法醫學主要以地層水文、化學性證據分析為主，尚缺乏生物性證據分析技術與相關知識。為填補此空白，本研究將導入高通量次世代定序(Next-generation high-throughput sequencing, HTS)與階層寡核苷酸引子延伸(Hierarchical Oligonucleotide Primer Extension, HOPE)等尖端分子生物分析技術，分析多氯乙烯地下水污染場址菌群結構與還原脫氯菌豐富度，以建立多氯乙烯污染的微生物指紋譜與污染指標菌群資料庫，結合多氯乙烯在厭氧還原脫氯過程微生物動態的關聯性，發展微生物追蹤法醫技術，相關研究結果並提供多氯乙烯生物整治所需的微生物學資訊。本計畫新設計偵測四屬脫氯菌的專一性引子，最佳化測試結果顯示HOPE分析技術能以低成本高通量的方式快速偵測於污染場址中相對豐富度高於0.4%的Dehalococcoides、Dehalobacter、Desulfitobacterium和Geobacter脫氯族群，或改以另一策略分析相對量更低的Dehalococcoides次族群，如與其他分生技術如分子選殖、定量PCR搭配運用，可快速提供場址整治過程脫氯菌群相關資訊。另一方面，本計畫導入土水微生物的高通量定序技術與大數據分析，除了全面性監測脫氯菌群之外，並探討土水微生物族群結構與含氯有機物的關聯性，提供地下水含氯有機物汙染微生物分析診斷的基礎。本計劃建立的整合性地下水微生物分析平台、資料庫及相關知識，將有助於提升國內多氯乙烯生物整治技術之水平，增加土水產業之國際競爭力。
計畫英文摘要	Currently, environmental forensics for groundwater pollutions is mainly relied on the analysis of hydrology and chemistry evidence. The microbiological evidence analysis is also compulsory but still in its infancy. This study applies the cutting-edge molecular biotechnology such as next-generation high-throughput sequencing and hierarchical oligonucleotide primer extension to analyze the microbial community structure and abundance of the anaerobic dechlorinators in the polychlorinated ethylene contaminating sites with an attempt to build up the relevant microbial fingerprint and indictor microorganism database. We have developed specific oligonucleotide primers and combined with hierarchical oligonucleotide primer extension (HOPE) to detect the common dechlorinators, including Dehalococcoides、Dehalobacter、Desulfitobacterium and Geobacter. With the optimal conditions obtained, the method can detect the targets at the abundance as low as 0.4% with respect to total bacterial population. Also, it can detect the three subgroup of Dehalococcoides. On the other hand, high-throughput sequencing (HTS) technology and big data analysis were used to study the microbial community structure in aquifers contaminated with chlorinated componds. Together with HOPE and HTS, the results obtained with the field samples suggest the efficacy of the methods used in this study to quantitative analysis of the dechlorinating populations, and providing relevant microbial diagnostic information for the improved management for polychlorinated contaminant bioremediation.