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專案計畫編號	EPA-107-E3S5-02-01
經費年度	107
計畫經費	1,900 千元
專案開始日期	2018/03/28
專案結束日期	2018/12/31
專案主持人	童心欣
主辦單位	環檢所
承辦人	林哲雄
執行單位	國立臺灣大學
專案分類	環境監測及檢測
中文關鍵字	硫酸銅、底泥、微生物菌群分布
英文關鍵字	copper sulfate, sediment, microbial community profile
協同主持人
共同主持人	于昌平；郭獻文
計畫聯絡人	周鈺錩
計畫聯絡電話	02-33664404#
計畫聯絡信箱	gold84double@gmail.com

中文摘要	本研究探討以微生物菌群變化作為污染源鑑識之可行性。本計畫第一年研究受硫酸銅影響之菌群變化，分為實驗室模擬反應槽暴露試驗及現地樣品測試。結果顯示Aeromonadaceae, Burkholderiales_incertae_sedis, Caldilineaceae, Chloroflexi, Firmicutes, Planctomycetaceae, Pseudomonodaceae, 及Xanthobacteraceae 皆重複出現在現場溪中底泥、溪水、及工廠放流水具有高濃度金屬銅的樣品中，有機會成為銅污染之普遍性指標判斷菌群。另外，由反應槽、老街溪河水及底泥菌相及樣品銅濃度交叉比對，結果顯示，Burkholderiaceae, Hydrogenophilaceae, Methylophilaceae, Chitinophagaceae, 及 Flavobacteriaceae等之豐富度消長與銅濃度相關。第二年研究以高通量次世代定序技術獲得菌株受硫酸銅污染下之基因表現變化。於老街溪工業區放流口上、下游共5個點位進行水質與底泥採樣，共取得5個水樣與5個底泥樣本。萃取底泥樣本中總DNA，並以MiSeq次世代定序分析16S-rRNA gene amplicon獲得菌群組成資訊。菌群分析結果除篩選出主要差異菌群外，並以PICRUSt運算模式評估菌群基因表現。另外，也同步利用illumnia shotgun sequencing分析各底泥樣品總基因體，取得與銅汙染相關的基因表現。結果顯示，工業區放流口下游有明顯增加趨勢的菌門為Microscillaceae及Geobacteraceae。而在受到工業區放流口影響趨勢較高的菌門則為Hydrogenophilaceae及Gallionaellaceae。此中，Hydrogenophilaceae菌門與第一年結果相符，在工業區放流口後豐度迅速減少，未來可能成為主要辨識污染之菌群。由不同菌群組成之基因表現預測(PICRUSt)分析，得到20種基因功能其相關基因豐度隨著上游至下游降低。大部分皆與代謝功能相關，次要則屬於細胞生化反應及遺傳信息處理，其中Calcium signal pathway 與Photosynthesis - antenna proteins 兩種基因豐度與底泥中重金屬銅、鉻、錳的濃度相關性非常高。這兩種的功能基因可能可作為未來篩選的標的基因。另外，以shot gun sequencing底泥基因型分析結果顯示，僅有少數基因在放流口下游樣品豐度較上游高。尤其是與銅、鋅、鎘在細胞膜間傳輸相關之ATPase activity (transporting or exporting)，未來也可能具有應用潛力。
中文關鍵字	硫酸銅、底泥、微生物菌群分布

中文摘要

本研究探討以微生物菌群變化作為污染源鑑識之可行性。本計畫第一年研究受硫酸銅影響之菌群變化，分為實驗室模擬反應槽暴露試驗及現地樣品測試。結果顯示Aeromonadaceae, Burkholderiales_incertae_sedis, Caldilineaceae, Chloroflexi, Firmicutes, Planctomycetaceae, Pseudomonodaceae, 及Xanthobacteraceae 皆重複出現在現場溪中底泥、溪水、及工廠放流水具有高濃度金屬銅的樣品中，有機會成為銅污染之普遍性指標判斷菌群。另外，由反應槽、老街溪河水及底泥菌相及樣品銅濃度交叉比對，結果顯示，Burkholderiaceae, Hydrogenophilaceae, Methylophilaceae, Chitinophagaceae, 及 Flavobacteriaceae等之豐富度消長與銅濃度相關。第二年研究以高通量次世代定序技術獲得菌株受硫酸銅污染下之基因表現變化。於老街溪工業區放流口上、下游共5個點位進行水質與底泥採樣，共取得5個水樣與5個底泥樣本。萃取底泥樣本中總DNA，並以MiSeq次世代定序分析16S-rRNA gene amplicon獲得菌群組成資訊。菌群分析結果除篩選出主要差異菌群外，並以PICRUSt運算模式評估菌群基因表現。另外，也同步利用illumnia shotgun sequencing分析各底泥樣品總基因體，取得與銅汙染相關的基因表現。結果顯示，工業區放流口下游有明顯增加趨勢的菌門為Microscillaceae及Geobacteraceae。而在受到工業區放流口影響趨勢較高的菌門則為Hydrogenophilaceae及Gallionaellaceae。此中，Hydrogenophilaceae菌門與第一年結果相符，在工業區放流口後豐度迅速減少，未來可能成為主要辨識污染之菌群。由不同菌群組成之基因表現預測(PICRUSt)分析，得到20種基因功能其相關基因豐度隨著上游至下游降低。大部分皆與代謝功能相關，次要則屬於細胞生化反應及遺傳信息處理，其中Calcium signal pathway 與Photosynthesis - antenna proteins 兩種基因豐度與底泥中重金屬銅、鉻、錳的濃度相關性非常高。這兩種的功能基因可能可作為未來篩選的標的基因。另外，以shot gun sequencing底泥基因型分析結果顯示，僅有少數基因在放流口下游樣品豐度較上游高。尤其是與銅、鋅、鎘在細胞膜間傳輸相關之ATPase activity (transporting or exporting)，未來也可能具有應用潛力。

中文關鍵字

硫酸銅、底泥、微生物菌群分布

類型	檔名	檔案大小	說明	最新版公開日期
期末報告	以菌群分布作為鑑識污染源技術建立期末報告.pdf	4MB	以菌群分布作為鑑識污染源技術建立(2/2)	2018/12/29 上午 12:00:00

摘要	This study investigates the possibility of applying microbial community profile changes to pollution source identification. In the first-year study, sediment microcosm was established and exposed to copper sulfate. In addition, water and sediments were sampled from a local river to verify the results. The results showed that Aeromonadaceae, Burkholderiales_incertae_sedis, Caldilineaceae, Chloroflexi, Firmicutes, Planctomycetaceae, Pseudomonodaceae, and Xanthobacteraceae can be found from microcosm, river sediments and factory discharge. And, the correlation analysis showed that Burkholderiaceae, Hydrogenophilaceae, Methylophilaceae, Chitinophagaceae, and Flavobacteriaceae were closely related to copper concentration. In the second year, sediments and water samples from both upstream and downstream of an actual industrial park discharging point were collected and analyzed. These samples will be used to verify the possibility of using the microbial change analysis as a pollution source identification. 16S-rRNA gene amplicon sequencing was performed for all 5 sediment samples collected from the river receiving industrial discharge and metagenomics function from each sample was estimated by PICRUSt (v1.1.3). In addition, whole sediment genome was also analyzed by illumnia shot gun sequencing. The amplicon sequencing results showed that Microscillaceae and Geobacteraceae were increased after discharging point. The Hydrogenophilaceae and Gallionaellaceae were two groups mostly affected by the industrial discharge. Especially, the Hydrogenophilaceae group is in agreement as the finding from the first year’s results. The results from PICRUSt estimation, there were 20 gene functions were affected by the industrial discharge, within those function, calcium signal pathway and photosynthesis – antenna proteins were the two major functions affected significantly by the industrial discharge, and may be considered as functional gene indicators in the future. From the whole genome analysis, the metal related (Cu, Zn, Cr) ATPase activity (transporting or exporting) gene expressed highest impact from the industrial discharge. It can also be considered and indicating functional gene for the future applications.
英文關鍵字	copper sulfate, sediment, microbial community profile

摘要

This study investigates the possibility of applying microbial community profile changes to pollution source identification. In the first-year study, sediment microcosm was established and exposed to copper sulfate. In addition, water and sediments were sampled from a local river to verify the results. The results showed that Aeromonadaceae, Burkholderiales_incertae_sedis, Caldilineaceae, Chloroflexi, Firmicutes, Planctomycetaceae, Pseudomonodaceae, and Xanthobacteraceae can be found from microcosm, river sediments and factory discharge. And, the correlation analysis showed that Burkholderiaceae, Hydrogenophilaceae, Methylophilaceae, Chitinophagaceae, and Flavobacteriaceae were closely related to copper concentration. In the second year, sediments and water samples from both upstream and downstream of an actual industrial park discharging point were collected and analyzed. These samples will be used to verify the possibility of using the microbial change analysis as a pollution source identification. 16S-rRNA gene amplicon sequencing was performed for all 5 sediment samples collected from the river receiving industrial discharge and metagenomics function from each sample was estimated by PICRUSt (v1.1.3). In addition, whole sediment genome was also analyzed by illumnia shot gun sequencing. The amplicon sequencing results showed that Microscillaceae and Geobacteraceae were increased after discharging point. The Hydrogenophilaceae and Gallionaellaceae were two groups mostly affected by the industrial discharge. Especially, the Hydrogenophilaceae group is in agreement as the finding from the first year’s results. The results from PICRUSt estimation, there were 20 gene functions were affected by the industrial discharge, within those function, calcium signal pathway and photosynthesis – antenna proteins were the two major functions affected significantly by the industrial discharge, and may be considered as functional gene indicators in the future. From the whole genome analysis, the metal related (Cu, Zn, Cr) ATPase activity (transporting or exporting) gene expressed highest impact from the industrial discharge. It can also be considered and indicating functional gene for the future applications.

英文關鍵字

copper sulfate, sediment, microbial community profile