Stubner S, Liesack W
Max-Planck-Institut fur terrestrische Mikrobiologie, D-35043 Marburg, Germany.
Appl Environ Microbiol. 1998 Dec;64(12):4983-9. doi: 10.1128/AEM.64.12.4983-4989.1998.
Because excised, washed roots of rice (Oryza sativa) immediately produce CH4 when they are incubated under anoxic conditions (P. Frenzel and U. Bosse, FEMS Microbiol. Ecol. 21:25-36, 1996), we employed a culture-independent molecular approach to identify the methanogenic microbial community present on roots of rice plants. Archaeal small-subunit rRNA-encoding genes were amplified directly from total root DNA by PCR and then cloned. Thirty-two archaeal rice root (ARR) gene clones were randomly selected, and the amplified primary structures of ca. 750 nucleotide sequence positions were compared. Only 10 of the environmental sequences were affiliated with known methanogens; 5 were affiliated with Methanosarcina spp., and 5 were affiliated with Methanobacterium spp. The remaining 22 ARR gene clones formed four distinct lineages (rice clusters I through IV) which were not closely related to any known cultured member of the Archaea. Rice clusters I and II formed distinct clades within the phylogenetic radiation of the orders "Methanosarcinales" and Methanomicrobiales. Rice cluster I was novel, and rice cluster II was closely affiliated with environmental sequences obtained from bog peat in northern England. Rice cluster III occurred on the same branch as Thermoplasma acidophilum and marine group II but was only distantly related to these taxa. Rice cluster IV was a deep-branching crenarchaeotal assemblage that was closely related to clone pGrfC26, an environmental sequence recovered from a temperate marsh environment. The use of a domain-specific oligonucleotide probe in a fluorescent in situ hybridization analysis revealed that viable members of the Archaea were present on the surfaces of rice roots. In addition, we describe a novel euryarchaeotal main line of descent, designated rice cluster V, which was detected in anoxic rice paddy soil. These results indicate that there is an astonishing richness of archaeal diversity present on rice roots and in the surrounding paddy soil.
由于切除并洗净的水稻（Oryza sativa）根在缺氧条件下培养时会立即产生甲烷（P. Frenzel和U. Bosse，《FEMS微生物生态学》21:25 - 36，1996），我们采用了一种不依赖培养的分子方法来鉴定水稻植株根上存在的产甲烷微生物群落。通过PCR从总根DNA中直接扩增古菌小亚基rRNA编码基因，然后进行克隆。随机选择32个古菌水稻根（ARR）基因克隆，并比较约750个核苷酸序列位置的扩增一级结构。环境序列中只有10个与已知产甲烷菌相关；5个与甲烷八叠球菌属相关，5个与甲烷杆菌属相关。其余22个ARR基因克隆形成了四个不同的谱系（水稻簇I至IV），它们与古菌的任何已知培养成员都没有密切关系。水稻簇I和II在“甲烷八叠球菌目”和甲烷微菌目的系统发育辐射范围内形成了不同的分支。水稻簇I是新的，水稻簇II与从英格兰北部沼泽泥炭中获得的环境序列密切相关。水稻簇III与嗜热栖热菌和海洋类群II出现在同一分支上，但与这些分类单元的关系较远。水稻簇IV是一个深分支的泉古菌组合，与从温带沼泽环境中回收的环境序列克隆pGrfC26密切相关。在荧光原位杂交分析中使用域特异性寡核苷酸探针表明，古菌的活成员存在于水稻根表面。此外，我们描述了一个新的广古菌主要谱系，命名为水稻簇V，它在缺氧的稻田土壤中被检测到。这些结果表明，水稻根和周围稻田土壤中古菌的多样性惊人地丰富。
