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年度	110	專案性質	實驗性質
專案類別	研究專案	研究主題	整治
申請機構	國立中正大學	申請系所	地球與環境科學系
專案主持人	許昺慕	職等/職稱	教授
專案中文名稱	含氯污染場址之現地微生物整治技術開發與本土菌群篩選
中文關鍵字	現地整治技術開發,本土菌群篩選,含氯污染場址
專案英文名稱	The development of in-situ microbial remediation techniques and functional bacteria isolation from local chlorine-contaminated sites
英文關鍵字	In-situ microbial remediation technical development, native functional bacteria isolation ,chlorine-contaminated sites
執行金額	1,019,975元
執行期間	2021/1/1 至 2021/12/31
計畫中文摘要	國內土壤及地下水含氯有機溶劑污染事件以四氯乙烯(PCE)和三氯乙烯(TCE)最為常見。本研究團隊擬定六年期生物復育整治計畫，預期達成下列最終目標: (1)評估污染場址自然衰減所需期程。(2)使用本土菌株添加，進行生物強化(Bioaugmentation)整治。(3)使用基質改良法，並利用高通量檢驗技術評估生物刺激(Biostimulation)之成效。評估現地生物復育技術可行性其中最重要條件之一是場址本身須具有可降解污染物之微生物，任何單一菌種微生物是無法完成所有的代謝反應進而達成含氯有機汙染物的降解。需要透過不同的細菌群落，共同交互執行的代謝反應才可完成。因此，快速、深入分析菌相變動及其可能的生物降解代謝途徑為評估現地生物復育的關鍵步驟。本年度計畫重於:(1)土壤及地下水現地整治策略開發。(2)泥火山風化土壤的甲烷及含氯汙染物共代謝菌株/群篩選與場址評估。(3)以第三代定序技術、化學分析、生物分子分析、磷脂質脂肪酸分析法，了解污染場址之整治現況與探討後續整治策略。(4)使用Biolog EcoPlate™結合菌相組成代謝分析，提供評估現地整治汙染物代謝方式及其代謝效率之方法。本研究完成Ecoplate與高通量定序分析之代謝結果預測整合，能有效提供汙染現地背景資料與後續整治策略菌群資訊。本研究調查泥火山樣本帶有甲烷營養菌以及對氯代謝有關之菌種，做為含氯污染場址之菌群添加試驗之研究基礎。研究發現，在噴發初期樣本中甲烷營養菌以及對氯代謝有關之菌種比較多，但甲烷營養菌的佔比較低，但隨著時間拉長，在該樣本中仍可發現甲烷營養菌的存在。另外，針對南臺灣不同區域的泥火山樣本中甲烷營養菌以及對氯代謝之KEGG代謝預測發現，花蓮及高雄甲仙泥火山相關代謝基因比例較高，可作為後續整治場址之參考依據。綜整兩年場址檢測結果發現，已進行過生物及化學整治場址，其相關氯代謝有關之菌種多樣性及相關基因檢測結果均較為豐富。後續本研究團隊將以微生物電化學之觀點評估該技術對含氯汙染場址之整治效益。
計畫英文摘要	Many domestic cases regarding soil and groundwater contaminated by volatile organic compounds have been reported to date. The most frequently detected volatile compounds in these cases include the two most pervasive chlorinated solvents namely Perchloroethene (PCE) and trichloroethene (TCE). The six-year project will be proposed for bioremediation of the PCE and TCE contaminated sites with the following three aims: (1) Assessing the efficacy for natural attenuation of the contaminated sites; (2) Isolation the indigenous bacteria for bioaugmentation; (3) Assessing the efficacy of biostimulation by high-throughput metagenomic technologies. One of the most key points for assessing the viability of bioremediation technology is that there are microorganisms with the potential to degrade contaminants. Any single microorganism cannot complete all the metabolisms to degrade contaminants, and only complex bacterial communities can perform all of the metabolic pathways to biodegrade contaminants. Therefore, the key step of bioremediation technology is to predominate bacterial communities’ variation and metabolic pathways, and bacterial communities are also the indicators for natural attenuation. This year's plan focuses on: (1) Development of on-site soil and groundwater remediation strategies; (2) Screening and site assessment of methane and TCE co-metabolizing strains/groups in the weathered soil of mud volcano; (3) Using the third-generation sequencing technology, chemical analysis, biomolecular analysis, and phospholipid fatty acid analysis methods to understand the current situation of the remediation of contaminated sites and discuss the follow-up remediation strategies; (4) Using Biolog EcoPlate™ combined with bacterial composition and metabolism analysis to provide a method to evaluate the metabolic mode and efficiency of on-site remediation of pollutants. This study completed the integration of the metabolic results prediction of Ecoplate and high-throughput sequencing analysis, which can effectively provide the bacterial information of the contaminated sites and the follow-up remediation strategy. The study investigates the methanotrophic bacteria and the chlorine-metabolized bacteria in mud volcano samples. The results will be the research basis for the microflora addition test of chlorine-contaminated sites. The study found that there were more methanotrophs and chlorine-metabolized bacteria in the samples collected at the beginning of the eruption. As time went on, the methanotrophs were still found in the sample. In addition, the KEGG metabolism prediction for methanotrophs and chlorine-metabolized bacteria in mud volcano samples found that the proportion of metabolic genes is relatively high in Jiaxian and Hualian mud volcanoe samples which can be used as a reference for subsequent research. Based on the two-year site study, the biological and chemical remediation sites carried out high bacterial species diversity and chlorine metabolic genes. In the follow-up, we will evaluate the effect of microbial electrochemistry for bioremediation.