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

摘要微生物采用许多新颖的策略来获取能量和营养物质，并以此改变其环境的化学性质，这对地质材料的形成和转化具有重要意义。我们建议研究这样一种策略：沉积盆地中的微生物甲烷生成。页岩地层水中 CH4、CO2 和 H2O 的稳定同位素特征表明多个天然气储量的微生物起源。然而，这些签名留下了一些有趣的问题没有得到解决。厌氧碳氢化合物降解通过许多代谢途径进行，包括硫酸盐、硝酸盐和铁还原、发酵和产甲烷，并已在石油储层、污染的含水层和深层海洋沉积物中得到认可。这项研究将通过探索地下沉积岩中有机物的厌氧分解导致甲烷经济积累来扩大对甲烷生成的认识。安特里姆页岩（美国密歇根州晚泥盆世）是北美最大的页岩气储量之一。同位素研究已证实这种气体主要是微生物来源的。该项目将重点关注贯穿主要产气趋势的南北向断面井，对地层水、存档岩屑和新钻探的井岩心进行取样。马蒂尼和同事之前对安特里姆地层水的研究为了解该地区的地下物理和化学环境提供了坚实的框架。与密歇根州北部的能源公司和土地所有者建立的积极的工作关系使我们能够以不同寻常的方式进入现场，包括在钻探过程中收集新鲜材料以及将产量较差的油井用作天然实验室的机会。智力优势。这项研究整合了水相地球化学、有机地球化学、热力学、分子生物学和微生物学的分析方法。我们将把我们的结果综合成一个概念模型，解决关键研究问题，包括：温度、盐度、营养物和代谢底物的分析如何揭示这些生物体活动的热力学和环境限制？沥青、干酪根和溶解有机物的化学表征揭示了地下环境中促进甲烷生成的有机物的特定来源？安特里姆页岩地层水中活性微生物的 16S 和功能基因分子系统发育是什么，以及这些生物体的代谢作用是什么初步努力已经确定了两口安特里姆井地层水中产甲烷古菌的高度多样性，包括甲烷微生物目、甲烷八叠球菌目和甲烷细菌目中全新的 mcrA 基因型簇和新的 16S 序列。基于 16S rRNA 的细菌多样性更加有限，仅限于与产醋酸醋杆菌密切相关的序列以及 Syntropomoadaceae 和 Syntropobacteraceae 内的序列。令人惊讶的是，尚未检测到硫酸盐或金属还原菌的 DNA 证据。因此，与一系列电子受体（O2、NO3、Fe、Mn和SO4）随后进行发酵和产甲烷作用（例如在现代沉积物中发现的）不同，Antrim的地下产甲烷环境可能更类似于记录的互养群落，其中碳氢化合物分解为乙酸盐和H2，只有当乙酸盐和H2又被产甲烷菌快速消耗时，这种反应才受到大力支持。我们的研究将探讨这两个假设。这项研究的更广泛影响是多种多样且深远的。这项研究将定义一个与人类活动相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覆盖古代分子生物特征。此外，这项研究还将揭示地球大气中受约束不良的还原气体来源，这对于了解地球表面碳库的大气成分、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。我们还与盖洛德高中建立合作伙伴关系，这是一所为我们现场地区居民提供服务的地区公立高中。由于天然气行业是当地经济的重要组成部分，因此对于这个中等收入地区的学生来说，在自家后院进行的科学活动中获得实践经验将非常有价值。最后，通过广泛传播我们的研究结果，我们将增进对天然气储量来源、勘探指数和储量生产控制的了解。

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.