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中國畜牧學會會誌
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中文摘要
自東海大學農牧場之荷士登乳牛（Holstein）瘤胃內分離出使用單一碳源之嗜甲烷菌，經物化分析 顯示其係革蘭氏陰性好氧短桿菌，最適生長條件為 37℃ 與 pH 6.5，甲烷及甲醇最適濃度為 10% 與 0.5%。經 16S rDNA 序列分析，確立此菌屬於Methylomonas 的菌種（暫名為Methylomonas strain THUA）。本研究擬對THUA 及其突變菌株（THUA-2）進行生長條件及碳源（甲烷及甲醇） 降解最佳化分析，以利未來大量培養此微生物與進行應用。將 nitrate mineral salts（NMS）培養液 分別添加瘤胃液 1%〜5%，共 5 組，碳源為 10% 甲烷；或 0.5〜3 g/L 酵母萃取物（yeast extract， YE），共 6 組，碳源為 0.5% 甲醇。於 37℃ 恆溫培養 24 小時，並於單位時間取出以分光光度 計監測菌種生長。試驗結果顯示瘤胃液對菌體（THUA 與THUA-2）之比生長速率影響不顯著， 而添加 1 g/L 之 YE 的THUA 組生長速率較未添加組提高 7.8 倍、THUA-2 組可提高 12 倍； 添加 3 g/L 之 YE 可使THUA 提高 4.2 倍、THUA-2 提高 10.6 倍。當碳源由 0.5％ 甲醇改為 10％ 甲烷時，於單位時間採集並經氣相層析儀測定甲烷濃度以計算降解速度。結果顯示添加 1 g/L 之 YE 可使THUA 生長速率提高 6.9 倍、THUA-2 變為 5.1 倍，甲烷的降解速度也提高 1.4 倍; 3 g/L 之 YE 可使THUA 及THUA-2 增為 5 倍。綜合以上結果顯示 1 g/L 酵母萃取物可同時使 THUA 及其突變株THUA-2 迅速生長，而使用 0.5％ 甲醇作為碳源時，其生長速率比使用 10% 甲烷更具效能。
英文摘要
A novel strain of Methanotrophic bacteria (strain THUA) isolated from rumen fluids of a Holstein cow at Tunghai university, were physicochemically determined as Gram-negative. Its optimal growing conditions were determined at 37 ℃ and at the pH value of 6.5. Single carbon sources such as methane and methanol were most effectively at 10% and 0.5%, respectively. The phylogenetic analysis based on its 16S rRNA gene sequence showed that the strain THUA belongs to the genus Methylomonas (thereof named Methylomonas strain THUA). In this study, THUA and its derived mutant strain THUA-2 were tested for the optimal conditions of cell growth and degradation of two carbon sources (methane and methanol). Nitrate mineral salts (NMS) medium was added with 1 ~ 5% of rumen fluids and 10% methane or with 0.5 ~ 3 g/L yeast extract (YE) and 0.5% methanol, respectively. The cultures were incubated at 37 ℃ for 24 hours and monitored by spectrophotometer. The results showed no significant differences in cell numbers with the supplementation of rumen fluids, but the specific growth rate at 1 g/LYE increased by 7.8 times for THUA and 12 times for THUA-2 when compared to the control group. At 3 g/L, the specific growth rate increased by 4.2 times for THUA and 10.6 times for THUA-2. Alternatively, the carbon source was switched to 10% methane along with yeast extracts supplementation. The results showed that the growth rates at 1 g/L increased by 6.9 times for THUA and 5.1 times for THUA-2. At 3 g/L, both THUA and THUA-2 amplified 5 times faster. Taken together, supplementation of 1 g/L YE to the NMS medium greatly improved the culture growth of THUA and THUA-2. Furthermore, methanol was a more suitable carbon source than methane for the strain THUA.