重庆凉风垭天然气富集区上覆菜园土壤甲烷氧化细菌群落分析

METHANE OXIDIZING BACTERIAL COMMUNITIES IN SOILS WITH CH_4 SEEPING IN LIANGFENGYA,CHONGQING

对重庆市凉风垭地区一具有天然气泄漏的菜园土壤进行采样,利用16S rDNA克隆文库研究了该菜园表层及底层土壤样品中甲烷氧化细菌的群落结构,以期了解甲烷泄漏地区甲烷氧化菌的丰度、结构和组成状况,为深入探讨甲烷在该地区的循环及甲烷泄漏对环境的影响奠定基础。DNA序列分析结果显示,该天然气富集区上覆菜园地土壤中Ⅰ型甲烷氧化细菌以甲基细菌属(Methylobacter)占绝对优势,在底层样品与表层样品中所占比例存在较大差异,且底层样品中Ⅰ型甲烷氧化菌的多样性指数(1.99)比表层样品中的多样性指数(1.3)高。Ⅱ型甲烷氧化细菌仅在底层样品CD1中检测到,以甲基孢囊菌属(Methylocystis)占优势,其多样性指数为2.07,种类较Ⅰ型种类要多,这可能暗示Ⅱ型甲烷氧化细菌更能适应在土壤底层低氧、高浓度CH_4条件下生存。同时,系统发育树分析显示,大部分菜园底层土壤样品中的Ⅰ型甲烷氧化细菌序列无法同已知的甲烷氧化细菌种属聚在一起,暗示该天然气富集区上覆菜园土壤中甲烷氧化细菌具有特殊性。由于Ⅰ型和Ⅱ型甲烷氧化菌对CH_4的亲和力以及对CH_4氧化能力的差异,天然气富集区甲烷氧化菌的群落结构研究对深入了解陆地生态系统CH_4代谢具有重要意义。

Methane seeping has been observed in an area of several km2 since 1950s in Liangfengya, Chongqing, south western China. To investigate the community structure and diversity of methane oxidizing bacteria （MOB） , which play important role in methane cycle, surface （5 ～ 10cm）and bottom （20 -25cm）soil samples in a vegetable garden with CH4 seeping were collected in May, 2008. Surface sample （CB）and bottom samples （CD1, CD2）were subjected to 16S rDNA gene sequencing and clone library constructions for type I and type 11 MOB respectively. Results showed that type ｜ MOB is dominated by Methylobacter sp. in all soil samples. CD1 and CD2 shared a same Shannon-Wiener index of type I MOB with a value of 1.99 despite of their difference in species compositions. Type II MOB was only detected in CD1 ,which is dominated by Methylocystis sp. The Shannon-Wiener index（2.07） is higher than that of type I MOB （1.99）, indicating the preference of Type Ⅱ MOB under conditions with high CH4 concentration. Some of the type I MOB sequences showed high similarities with clones from various environments such as riverine sediments,soils,groundwater ect, which imply a ubiquitous distribution of these type I MOB in natural environments. However, most of the sequences of type I MOB in bottom soils show very low similarities with the known sequences in NCBI database, suggesting a unique MOB group in the CH4- seeping soils. Due to the great difference in CH4 oxidizing capacities of type I and Ⅱ MOB,our data will shed light on understanding the CH4 cycle in the terrestrial seeping sites.