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研究生: 王子境
Wang, Zi-Jing
論文名稱: 底層型蚯蚓土後腔環蚓應用於鉻污染土壤之生物整治
Bioremediation of chromium-contaminated soil using endogeic earthworm Metaphire posthuma
指導教授: 黃榮振
Huang, Jung-Chen
學位類別: 碩士
Master
系所名稱: 工學院 - 環境工程學系
Department of Environmental Engineering
論文出版年: 2024
畢業學年度: 112
語文別: 中文
論文頁數: 93
中文關鍵詞: 蚯蚓整治植物整治生物整治蚯蚓
外文關鍵詞: bioremediation, phytoremediation, vermiremediation, Chromium, earthworm
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  • 本研究分為兩種面向進行實驗,第一種面向為探討土後腔環蚓Metaphire posthuma 在鉻污染土壤整治之角色。研究結果顯示,當土壤鉻濃度為60 mg/kg,蚯蚓的致死率與重量損失分別為84.4%、39.1%。經蚯蚓攝入的土壤形成蚓糞後,鉻含量減少了33.6%,並顯著增加鉻生物可利用性。蚯蚓體內鉻濃度從11.1 mg/kg上升至41.7 mg/kg。然而由於較少的蚯蚓數量與較短的整治期,土壤的整體鉻濃度並沒有變化。微生物群落分析顯示,鉻略減少蚯蚓腸道中的微生物多樣性,然而在6種功能菌屬中有5種表現出豐富度上升的趨勢,其中以Pseudomonas (~32.24%) 與Aeromonas (~34.02%) 為優勢屬。在鉻污染土壤發現16種功能菌屬,其中9種功能菌屬在蚯蚓活動下增加了相對豐富度。此外,蚯蚓在鉻污染土壤貢獻2種未在無蚯蚓土壤發現的功能菌屬。Bacillus pseudomycoides為蚓糞中篩選出之鉻還原菌,具有一定的鉻抗性,在300 mg Cr6+/L之LB培養基中仍能存活,並在100 mg Cr6+/ L之LB培養基中表現出最佳的還原能力 (三天後還原95.46%)。總結來說,本實驗結果顯示M. posthuma在鉻污染土壤中有著作為功能菌載體或刺激劑之潛力,可能透過間接促進鉻還原菌在土壤的優勢,增進鉻在土壤中的安定化以減少毒性,提供該整治技術在底層型蚯蚓之應用於鉻污染土壤之發展基礎。
    第二種面向為探討底層型蚯蚓在植物整治鉻污染土壤中的作用。本實驗以向日葵 (Helianthus annuus) 作為植物。當植物在鉻濃度100 mg Cr/kg 之土壤生長一個月後,蚯蚓顯著促進植物根部與葉子之生長,分別增加了147% 與 158% 之生物量,莖部增加48%,但未顯著。蚯蚓也增進植物對鉻的吸收,根部鉻濃度與鉻累積量分別上升50.1 % 與272 %,但莖、葉部位的鉻濃度微乎其微。在蚯蚓活動下,生長三個月之植物與一個月之植物相比,其根部之鉻濃度減少了11.7%,而莖、葉之鉻濃度顯著上升至0.69與 1.38 mg Cr/kg,花與種子的鉻濃度為1.55與1.62 mg Cr/kg。無論哪一種實驗組,植物皆未影響土壤鉻濃度,但提升了鉻生物可利用性。在根際微生物群落中,鉻使微生物多樣性上升,蚯蚓則略減少微生物多樣性。蚯蚓活動增進11種鉻還原菌屬、3種PGPR屬以及5種CRPGPR屬 (Cr-reducing & PGPR) 在根際微生物群落中的優勢。整體來看,蚯蚓不僅促進植物生長、鉻組織濃度與累積量,同時提升多種PGPR與鉻還原菌在根際間的優勢,進一步證實M. posthuma具有強化植物整治以及功能菌刺激劑與載體的作用,透過提升植物對鉻的吸收以及鉻還原菌在根際間的優勢,增進土壤鉻的去除與安定化的效果,提供底層型蚯蚓整合植物整治之發展基礎。

    This study aimed to explore the role of the endogeic earthworm Metaphire posthuma in the remediation of chromium-contaminated soil. First, the Cr toxicity tests show that the survival rate and the weight loss of earthworms reached 84.4% and 39.1%, respectively, at the soil chromium concentration of 60 mg/kg. After digestion by worms, the Cr concentration in the castings decreased by 33.6% and significantly increased Cr bioavailability. Cr concentration in the earthworm biomass increased from 11.1 mg/kg to 41.7 mg/kg. The microbial community analysis reveals that there were 16 functional bacterial genera in chromium contaminated soil, of which 9 functional bacterial genera and 2 functional bacterial genera were not found in the worm-free soil, suggesting their richness was enhanced by earthworm activity. Moreover, Bacillus pseudomycoides screened from earthworm feces survived at 300 mg Cr/L, showing great chromium resistance as well as the profound reduction ability (95.46% has been reduced on the third day).
    Meanwhile, the study further investigated effects of M. posthuma on the phytoremediation of the chromium contaminated soil by sunflowers (Helianthus annuus). The results suggest the presence of earthworms apparently promoted the growth of the plant in the Cr-contaminated soil at 100 mg Cr/kg. Compared with the plants grown in the Cr-contaminated soil without earthworms, the biomass of roots, stems and leaves increased by 147%, 48% and 158%, respectively, for the plants growing in the Cr-contaminated and worm-inhabited soil. Over an experimental period of 30 days, the Cr uptake by the plant was also enhanced in the presence of earthworms, with a 50.1 % and 272 % increase in Cr concentration and accumulation in roots, compared to those in the absence of earthworms. Moreover, earthworm activity seemed to have enhanced the dominance of 11 genera of Cr-reducing bacteria, 3 genera of PGPR, and 5 genera of CRPGPR (Cr-reducing & PGPR) in the rhizosphere microbial community.
    Overall, our study demonstrated that M. posthuma could strengthen the phytoremediation of Cr-contaminated soil by sunflowers particularly through the Cr accumulation in the root. Meanwhile, the earthworm could function as a functional bacterial carrier or stimulant in Cr-contaminated soils and rhizospheres, prompting Cr immobilization and reducing Cr toxicity in the soil. The findings provide a solid foundation for the application of vermiremediation by endogeic earthworms and endogeic earthworm-mediated phytoremediation of chromium-contaminated soils.

    摘要 I Abstract III 誌謝 VIII 目錄 X 表目錄 XII 圖目錄 XIII 第一章 前言 1 1.1 研究動機 1 1.2 研究目的 3 第二章 文獻回顧 4 2.1 鉻 4 2.2 鉻污染整治技術 5 2.3 蚯蚓整治 (Vermiremediation) 5 2.3.1 蚯蚓吸收土壤重金屬之途徑 6 2.3.2 蚯蚓對重金屬毒性的防禦機制 6 2.3.3 土壤重金屬在蚯蚓作用下之宿命 7 2.3.4 影響蚯蚓整治重金屬之因子 7 2.3.5 底層型蚯蚓應用於蚯蚓整治重金屬土壤之相關研究 10 2.4 植物整治 (phytoremediation) 11 2.4.1 鉻在土壤-植物的轉移 12 2.4.2 鉻超富集植物 12 2.4.3 植物對鉻毒性的防禦機制 13 2.4.4 蚯蚓整合植物整治重金屬土壤之相關研究 14 第三章 研究方法 16 3.1 實驗架構 16 3.2 實驗材料 16 3.3 實驗方法 17 3.3.1 蚯蚓之鉻毒性試驗 17 3.3.2 底層型蚯蚓對鉻污染土壤之作用 17 3.3.3 蚯蚓對植物整治鉻污染土壤之作用 18 3.4 總鉻分析 19 3.5 鉻之生物可利用性 19 3.6 蚯蚓、植物之生長表徵與重金屬累積指標 20 3.7 鉻功能菌篩選及鉻還原效率測試 22 3.8 微生物群落分析 22 3.9 鉻還原菌種之鑑定 23 3.10 數據分析 23 第四章 結果與討論 24 4.1 底層型蚯蚓在鉻污染土壤之整治作用 24 4.1.1 鉻對蚯蚓之毒性 24 4.1.2 蚯蚓對鉻宿命之作用 27 4.1.3 蚯蚓與鉻對微生物群落之影響 31 4.1.4 鉻還原菌之篩選與其鉻還原效率 39 4.2 蚯蚓對植物整治鉻污染土壤之作用 42 4.2.1 蚯蚓對暴露於鉻污染土壤之植物生長特徵之影響 42 4.2.2 蚯蚓對鉻在植物整治下之宿命之作用 44 4.2.3 蚯蚓與鉻對植物根際微生物群落之影響 49 第五章 結論與建議 57 5.1 結論 57 5.2 建議 58 參考文獻 60

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