COLLABORATIVE RESEARCH: Active subsurface methanogenesis and anaerobic organic matter decomposition in a Devonian black shale

摘要微生物利用许多新的策略来获取能量和营养，并在这样做的过程中改变其环境的化学性质，这些方式对地质物质的形成和转化具有重要意义。我们建议调查这样一个战略：微生物甲烷生成在沉积盆地。页岩层沃茨中甲烷、二氧化碳和水的稳定同位素特征表明，几个天然气储量的微生物来源。然而，这些签名留下了有趣的问题未解决。厌氧烃类降解通过许多代谢途径进行，包括硫酸盐、硝酸盐和铁还原、发酵和甲烷生成，并且已经在石油储层、受污染的含水层和深海沉积物中被认识到。这项研究将通过探索地下沉积岩中有机物的厌氧分解来扩大对甲烷生成的理解，从而导致经济的甲烷积累。安特里姆页岩（晚泥盆世，美国密歇根州）是北美最大的页岩气储量之一。同位素研究表明，这种气体主要是微生物产生的。本项目将重点关注威尔斯井的南北向断面，通过主要产气趋势、地层沃茨取样、存档岩屑和新钻的井芯。Martini及其同事先前对Antrim地层沃茨的研究为了解该地区的地下物理和化学环境提供了一个坚实的框架。与密歇根州北方的能源公司和土地所有者建立了积极的工作关系，使我们能够以不同寻常的程度进入现场，包括在钻井期间收集新鲜材料的机会，以及将产量低的威尔斯井用作天然实验室。该研究综合了水地球化学、有机地球化学、热力学、分子生物学和微生物学的分析方法。我们将把我们的研究结果合成一个概念模型，解决关键的研究问题，包括：如何分析温度，盐度，营养物质和代谢底物揭示热力学和环境对这些生物体的活动的限制？沥青、干酪根和溶解有机物的化学特征揭示了在这种地下环境中产生甲烷的有机物的具体来源是什么？安特里姆页岩层沃茨中活性微生物的16 S和功能基因分子遗传学是什么，这些生物体发挥什么代谢作用？初步研究确定了两口Antrim威尔斯井的地层沃茨中产甲烷菌的高度多样性，包括一个全新的mcrA基因型簇和甲烷微生物目、甲烷微生物目和甲烷杆菌目内的新型16 S序列。基于16S rRNA的细菌多样性更有限，仅限于与产乙酸醋杆菌密切相关的序列以及在合养菌科和合养杆菌科内。令人惊讶的是，尚未发现硫酸盐或金属还原细菌的DNA证据。因此，而不是一套电子受体（O2，NO3，Fe，Mn和SO4），然后发酵和产甲烷（如在现代沉积物中发现），地下的安特里姆产甲烷环境可能更接近记录的互养社区，其中烃类分解为乙酸和H2，一个反应，这是积极的青睐，只有当乙酸和H2反过来迅速消耗产甲烷菌。我们的研究将探讨这两种假设。这项研究的广泛影响是多种多样的，影响深远。这项研究将确定一个与人类活动有关的不寻常的微生物群落。它还将详细说明岩石中的微生物活动如何混淆和叠加古代分子生物特征。此外，该研究将揭示地球大气中减少的气体的约束性差的来源，这对于理解地球表面储层中的大气成分，碳循环和碳转化的控制非常重要。我们的合作研究将增加教育和培训的机会，包括新的五所学院生物地球化学课程。我们还发起与盖洛德高中，区域公立高中服务于我们的外地居民的伙伴关系。由于天然气工业是当地经济的很大一部分，这将是宝贵的学生在这个中等收入地区获得实践经验与科学的努力发生在他们的后院。最后，通过广泛传播我们的结果，我们将提高对天然气储量来源、勘探指标和储量生产控制的认识。

ABSTRACTMicroorganisms employ many novel strategies to derive energy and obtain nutrients, and in doing so alter the chemistry of their environments in ways that are significant for formation and transformation of geologic materials. We propose to investigate one such strategy: microbial methane generation in sedimentary basins. Stable isotopic signatures of CH4, CO2 and H2O in shale formation waters indicate a microbial origin for several natural gas reserves. However, these signatures leave intriguing issues unaddressed. Anaerobic hydrocarbon degradation proceeds by many metabolic pathways including sulfate, nitrate, and iron reduction, fermentation, and methanogenesis, and has been recognized in petroleum reservoirs, polluted aquifers and deep marine sediments. This research will expand understanding of methane generation by exploring anaerobic decomposition of organic matter in subsurface sedimentary rocks leading to economic methane accumulations.The Antrim shale (Late Devonian, Michigan USA) is one of the largest shale gas reserves in North America. Isotopic studies have established that this gas is dominantly microbial in origin. This project will focus on a N-S transect of wells through the main gas-producing trend, sampling formation waters, archived cuttings, and freshly drilled well cores. Prior research on Antrim formation waters by Martini and co-workers provides a solid framework for understanding the subsurface physical and chemical environments within the region. A positive working relationship with energy companies and land owners in northern Michigan allows us an unusual degree of access to the field site, including opportunities to collect fresh material during drilling and to use poorly-producing wells as natural laboratories.Intellectual Merit. This research integrates analytical approaches from aqueous geochemistry, organic geochemistry, thermodynamics, molecular biology and microbiology. We will synthesize our results into a conceptual model that addresses key research questions including: How do analyses of temperature, salinity, nutrients and metabolic substrates reveal thermodynamic and environmental constraints on the activity of these organisms?What does chemical characterization of bitumens, kerogen, and dissolved organic matter reveal about specific sources of organic matter that fuel methane generation in this subsurface environment?What are the 16S and functional gene molecular phylogenies of active microorganisms within Antrim Shale formation waters, and what metabolic roles do these organisms play?Preliminary efforts have determined a high diversity of methanogenic Archaea in formation waters from two Antrim wells, including an entirely new cluster of mcrA genotypes and novel 16S sequences within the Methanomicrobiales, Methanosarcinales, and Methanobacteriales. 16S rRNA-based Bacterial diversity is more limited, confined to sequences closely related to acetate-producing Acetobacterium and within the Syntrophomoadaceae and Syntrophobacteraceae. Surprisingly, no DNA evidence for sulfate- or metal-reducing bacteria has yet been detected. Thus rather than a suite of electron acceptors (O2, NO3, Fe, Mn and SO4) followed by fermentation and methanogenesis (such as found in modern sediments), the subsurface methanogenic environment of the Antrim may more closely resemble documented syntrophic communities in which hydrocarbons are decomposed to acetate and H2, a reaction that is energetically favored only when acetate and H2 are in turn rapidly consumed by methanogens. Our research will explore both of these hypotheses.The broader impacts of this research are varied and far reaching. This study will define an unusual microbial community relevant to human activities. It will also detail how microbial activity in rocks may confound and overprint ancient molecular biosignatures. In addition, the study will shed light on a poorly-constrained source of reduced gases to Earth's atmosphere, important for understanding controls on atmospheric composition, carbon cycling and carbon transformations in earth surface reservoirs. Our collaborative research will increase opportunities in education and training, including a new Five Colleges Biogeochemistry course. We are also initiating partnership with Gaylord High School, the regional public high school serving residents of our field area. Because the gas industry is a large part of the local economy, it will be valuable for students in this modest-income region to gain hands-on experience with scientific endeavors occurring in their backyard. Lastly, by disseminating our results widely, we will improve understanding of gas reserve origins, indices for exploration, and controls on reserve production.