Lin-Yuan Huang , Yi-Fan Liu , Wan-Qi Qin , Shi-Yi Wang , Yuan-Hao Wang , Lei Zhou , Shi-Zhong Yang , Ji-Dong Gu , Bo-Zhong Mu
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引用次数: 0
Abstract
Obtaining pure cultures and enrichment systems of methylotrophic anaerobic microorganisms capable of utilizing methyl compounds is critical to studying the carbon cycle in subsurface anoxic oil reservoirs. Culture-independent methods have been instrumental in uncovering the rich diversity of microorganisms in oil reservoirs. However, there remains a notable scarcity of methylotrophic microorganisms from oil reservoirs obtained by culture-dependent methods. In this study, we used five different methyl compounds, namely methanol, methylamine, dimethylamine trimethylamine, and methyl sulfide, as the sole substrates to isolate methyl-utilizing microorganisms from oil reservoirs, and H2 was also added together with each of these methyl compounds in a separate isolation experiment, to facilitate H2-dependent methylotrophic growth and metabolism. Notably, the highest colony numbers in roll tubes were achieved when using methanol as the substrate. A total of 306 pure strains, representing eight genera were obtained, and these microorganisms have rarely been reported for their ecological roles in oil reservoir systems. Following isolation, each strain was tested for utilization of methanol as the sole carbon and energy sources after a second transfer, including headspace gas assay and microbial cell observations. In addition, enrichment cultures amended with each of the five different methyl compounds with or without the addition of H2 gas were established from four oil reservoir samples. Further experiments showed that the archaea enriched with methyl substrates from different oil reservoirs were almost all Methanobacteria, but after adding H2, the H2-dependent methylotrophic methanogen Ca. Methanomethylica was enriched in most of the enrichment cultures. On the contrary, the addition of H2 has less impact on the bacterial communities. The isolation of pure cultures has significantly enhanced our understanding of the diversity and ecophysiology of methylotrophic microorganisms in oil reservoirs. This study has provided useful insights into methyl-based methane generation within oil reservoirs, contributing to further understanding of the microbial ecology and carbon cycle in anoxic oil reservoirs.
获得能够利用甲基化合物的甲基厌氧微生物的纯培养物和富集系统对于研究地下缺氧油藏的碳循环至关重要。不依赖培养的方法有助于发现油藏中微生物的丰富多样性。然而,通过依赖培养的方法从油藏中获得的养甲微生物仍然非常稀少。在本研究中,我们使用了五种不同的甲基化合物(即甲醇、甲胺、二甲胺三甲胺和甲基硫醚)作为唯一底物,从油藏中分离出甲基利用微生物,并在单独的分离实验中将 H 与这些甲基化合物一起加入,以促进 H 依赖性的甲基营养生长和代谢。值得注意的是,使用甲醇作为底物时,辊管中的菌落数量最多。这些微生物在油藏系统中的生态作用鲜有报道。分离后,对每个菌株进行了二次转移后的甲醇作为唯一碳源和能源的利用测试,包括顶空气体检测和微生物细胞观察。此外,还从四个油藏样本中建立了用五种不同的甲基化合物分别进行添加或不添加 H 气体的富集培养物。进一步的实验表明,不同油藏中以甲基底物富集的古细菌几乎都是甲烷菌,但添加 H 后,大多数富集培养物中都富集了依赖 H 的甲烷菌 Methanomethylica。相反,添加 H 对细菌群落的影响较小。纯培养物的分离大大加深了我们对油藏中养甲烷微生物的多样性和生态生理学的了解。这项研究对油藏中基于甲基的甲烷生成提供了有益的见解,有助于进一步了解缺氧油藏中的微生物生态学和碳循环。
期刊介绍:
International Biodeterioration and Biodegradation publishes original research papers and reviews on the biological causes of deterioration or degradation.