全球每年之總甲烷釋出量約為110Tg，約有21 %由濕地系統中釋放自大氣環境，而甲烷的分解依賴甲烷氧化菌。本研究擬建立台灣地區不同濕地系統之甲烷氧化菌群落結構，比較自然濕地、人工濕地土壤以及廢水處理廠樣品之甲烷氧化速率，並馴養分離出甲烷氧化菌，測定各菌株的甲烷氧化能力，並利用甲烷氧化菌16S rDNA引子進行PCR反應，依定序結果進行各菌種之親緣關係。
甲烷氧化能力實驗中，添加0.1%甲烷濃度，四草自然濕地土壤之甲烷氧化率最高；添加20%甲烷濃度下，則以廢水處理廠之水樣的甲烷氧化率最高。從各樣區篩選分離出之甲烷氧化菌HNW-1、HNW-2、SNW-2、SCW-4、ECW-3、R-4、W-4、W-5 均具有甲烷氧化能力，其中SCW-3及R-4之甲烷氧化能力較高。其中以革蘭氏染色結果SNW-2、ECW-3、W-4、W-5為革蘭氏陽性菌，HNW-1、HNW-2、SCW-4、R-4革蘭氏陰性菌。分析16S rDNA之序列所繪製出親緣樹狀圖中顯示，HNW-1和HNW-2與Type I為同一次群，其可信度達99%，推測自然濕地中所分離出之菌株為甲烷氧化菌之Type I。W-4 與Methylorhabdus可信度達99%；而SNW-2、SCW-4、R-4、W-5、ECW-3、SA-4可能為甲烷氧化菌Type I與Type II之外的單系群，推測可能為Type X，且可能為台灣本土新屬種。另分離自二行人工溼地的ECW-3，與自南沙崙現齡掩埋區所分離之SA-4有極高的相似度，顯示雖為不同的二種環境，然相近之地理關係仍可在演化上形成較為相近之親緣關係。 Methane emission from wetlands is estimated to 110 Tg yr-1, and it is one of the largest natural sources of atmospheric methane, accounting for about 21% of the current global annual methane buget. Methanotrophs are important regulators of methane fluxes from the wetlands to atmosphere. There is very limited information presently available on the community structure of methanotrophs in wetlands in Taiwan. The study is an investigation on the community structure of methanotrophs as revealed by molecular method in different wetland soils. The methanotrophic strains were isolated by enrichment culture from natural wetlands, constructed wetlands paddies and wastewater treatment plant, and thereby subsequent loss of a variety of other methanotroph. Comparisons of methane oxidation potential of various wetland soils and different isolated were done in this study. Furthermore, the pylogenic relationship was constructed by 16S rDNA.
The maximum methane oxidation rates was found from Szu-tsao natural wetland in addition of 0.1% CH4, while the maximum was found from wastewater treatment plant in addition of 20% CH4. All the isolated strains (HNW-1、HNW-2、SNW-2、SCW-4、ECW-3、R-4、W-4、W-5) owned high oxidation potential of methane. SNW-2、ECW-3、W-4、W-5 are Gram-positive strains；HNW-1、HNW-2、SCW-4、R-4 are Gram-negative strains. Phylogenetic analysis of the test strains was identified by 16S rDNA gene fragment from NCBI database and MEGA software. The result indicated that HNW-1 and HNW-2 are clustered in one subgroup from the phylogenic tree constructed by 16 S rDNA and belong to the Type I. W-4 is the most closely related to a group Methylorhabdus. SNW-2, SCW-4, R-4, W-5, ECW-3 and SA-4 are monophly group distincted from Type I and Type II, inferred to TypeX and might be a new genus from Taiwan. The highly silimilarity between ECW-3 (from Erh-Hang constructed wetland) and SA-4 (from Nan-Sa-Lun landfill) was shown in this study. It was indicated that the closely phylogenetic relationship was depended on the adjacent sampled sites geographically.
