博碩士論文 112821010 詳細資訊




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姓名 郭柏陞(Po-Sheng Kuo)  查詢紙本館藏   畢業系所 生命科學系
論文名稱 硫酸還原菌與脫鹵球菌共培養系統中生物性硫化亞鐵的生成與應用
(Synergistic Formation and Application of Biogenic Iron Sulfide in a Co-culture System of Sulfate-Reducing Bacteria and Dehalococcoides mccartyi)
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摘要(中) 多氯乙烯(Chlorinated ethenes, CEs)在台灣本土污染場址佔據數量之冠,尤其於富含硫酸鹽的地下水中,嚴重阻礙了整治進程。由硫酸還原菌產生的硫化氫(H2S)會影響 Dehalococcoides 的脫氯反應效率,使含氯有機物降解不完全,進而導致生物復育失敗。為了探究硫酸還原和脫氯之間的相互作用,本研究旨在利用亞鐵化合物來克服硫化氫的毒性,從而生成硫化亞鐵(FeS)。由於其高還原潛能, FeS 被廣泛應用於化學整治領域。首先,透過添加氯化亞鐵(FeCl2),能夠有效移除硫酸還原菌產生的硫化氫,進而促進對多氯乙烯的完全降解。其次,藉由添加與硫酸鹽濃度相等比例的 FeCl2 ,有效將硫化氫移除至僅剩 3.1% 並促進生物性 FeS 的形成。相比過量濃度 FeCl2 的添加, FeS 的晶體顆粒大小有效降低多達 10 倍,減緩了對現地整治中造成的堵塞問題。透過轉錄體分析證實,添加 FeCl2 不僅有效地消除了硫化氫的抑制作用,增強了有機鹵素呼吸相關酵素的活性,甚至上調了參與電子傳遞的關鍵基因表現,從而提高了脫氯效率和硫酸還原菌的活性。最後,利用臺灣本土原生脫氯菌群應證於硫酸鹽及 CEs 共存環境下,脫鹵球菌的增長趨勢與 FeS 的形成呈現正相關。由多樣性分析進一步證實了 FeS 的生成有助於恢復菌群結構,甚至增加了菌群的多樣性。本研究彰顯了在硫酸鹽和多氯乙烯共存環境下共培養系統對於生成生物性硫化亞鐵的潛力,同時移除了硫酸還原產物硫化氫對脫氯反應的抑制,並結合化學及生物整治工法以提升多氯乙烯的降解效率。
摘要(英) Chlorinated ethenes (CEs) constitute a predominant contaminant in Taiwan′s native polluted sites, particularly in groundwater inundated with sulfate salts that substantially impede remediation efforts. Hydrogen sulfide (H2S) produced by sulfate-reducing bacteria (SRB) impairs the dechlorination efficiency of Dehalococcoides, resulting in incomplete chloride degradation and thereby leading to the failure of bioremediation. In order to elucidate interactions between sulfate reduction and dechlorination, this study aims to utilize ferrous compounds to overcome H2S toxicity by synthesizing ferrous sulfide (FeS), which is commonly utilized in chemical remediation due to its high reduction potential. Initially, the addition of ferrous chloride (FeCl2) effectively removed H2S production from SRB and enhanced the degradation of trichloroethylene to ethene. Optimal efficiency in biogenic FeS generation was achieved by adding FeCl2 in equal ratio to sulfate concentration. This approach effectively reduced H2S and crystal particle sizes by up to 10 times compared to excessive FeCl2 dosages, mitigating clogging issues during in situ remediation. Transcriptomic analysis revealed that the addition of FeCl2 eliminated hydrogen sulfide inhibition, enhanced dehalogenase activity, and upregulated key electron transport genes, increasing dechlorination efficiency and sulfate-reducing bacteria activity. Finally, utilizing Taiwan′s indigenous dechlorinating consortium CW5 in a simulated sulfate and CEs co-contaminated environment, the growth trend of Dehalococcoides showed a positive correlation with the FeS formation. Biodiversity analysis confirmed that FeS formation facilitated microbial community structure restoration and increased diversity. This study validates the potential of the co-culture system in generating biogenic FeS under sulfate and CEs co-contamination, removing sulfate-reducing products and improving CEs remediation through integrated chemical and biological remediations.
關鍵字(中) ★ 多氯乙烯
★ 硫化氫抑制
★ Dehalococcoides
★ 生物性硫化亞鐵
★ 硫酸還原菌
關鍵字(英) ★ Chlorinated ethenes
★ Sulfide inhibition
★ Dehalococcoides
★ Biogenic iron sulfide
★ Sulfate-reducing bacteria
論文目次 國立中央大學圖書館學位論文授權書 I
國家圖書館學位論文延後公開申請書 II
論文指導教授推薦書 III
論文口試委員審定書 IV
摘要 V
Abstract VI
致謝 VII
目錄 VIII
圖目錄 XI
表目錄 XV
第壹章 緒論(Introduction) 1
1.1 三氯乙烯之污染 1
1.2 三氯乙烯污染場址整治 ⸻ 微生物整治 2
1.3 硫酸鹽及三氯乙烯之共同污染 4
1.3.1 硫化氫之生成抑制脫氯反應 4
1.3.2 硫化亞鐵於非生物整治之應用 4
1.4 生地化整治工法 ⸻ 結合微生物及非生物整治之優勢 5
第貳章 實驗目的及架構 7
第參章 實驗材料及方法(Materials and Methods) 9
3.1 菌株來源、繼代培養與保存 9
3.1.1 菌種及來源 9
3.1.2 Hungate絕對厭氧技術 9
3.1.3 繼代培養及短期保存 10
3.1.4 菌種長期保存 12
3.2 氣相層析火焰離子化偵檢器(Gas Chromatography - Flame Ionization Detector, GC-FID) 12
3.3 含氯有機物降解速率計算分析(Dechlorination kinetic: plot fitting and calculations) 13
3.4 細菌 Genomic DNA 萃取 14
3.5 全基因組定序(Whole genome sequencing) 15
3.6 親緣關係樹狀圖分析(phylogenetic analysis) 17
3.7 即時聚合酶連鎖反應(Real-time polymerase chain reaction, qPCR) 17
3.8 硫酸鹽濃度測定 18
3.9 硫化氫濃度測定 19
3.10 鐵離子濃度測定 20
3.11 硫化亞鐵形成之測定方法 21
3.12 高解析掃描式電子顯微鏡(Scanning Electron Microscope , SEM)及能量散射X射線譜(Energy-dispersive X-ray spectroscopy, EDS) 22
3.12.1 硫化亞鐵之固定 22
3.12.2 硫化亞鐵之結構與組成 23
3.13 細菌 RNA 萃取 24
3.14 轉錄組分析(Transcriptomics Analysis) 25
3.15 Full-length 16S rRNA基因擴增子分析(Full-length 16S rRNA gene amplicon analysis) 26
第肆章 實驗結果(Results) 29
4.1 以亞鐵化合物移除硫化氫對脫鹵球菌造成之毒性 29
4.2 不同濃度亞鐵化合物移除硫化氫之效率及 FeS 生成 30
4.3 以轉錄體分析硫酸還原菌及脫鹵球菌間之相互影響 32
4.4 臺灣本土地下水中菌群組成對FeS生成及硫化氫去除的影響 34
第伍章 討論(Discussion) 35
5.1 探討脫鹵球菌及硫酸還原菌共培養相互影響之關係 35
5.2 探討FeS生成的類型及環境影響因素 36
5.3 以基因表現差異比較FeS生成之優劣 38
5.3.1 解析 D. mccartyi CWV2 及 N. liaohensis KPS 間的相互影響 38
5.3.2 探究FeS生成對電子傳遞的促進作用及相關基因表達上升 41
5.4 共污染環境下原生菌群之菌相組成改變 42
第陸章 結論(Conclusion) 44
參考文獻(References) 45
圖表 60
附加資料(Supplementary File) 89
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指導教授 陳師慶(Ssu-Ching Chen) 審核日期 2024-7-26
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