CAREER: Microbial Mineral Oxidation in a Temperate Marine Sediment: Quantifying the importance of extracellular electron transfer to sediment sulfur biogeochemistry

众所周知，许多海洋沉积物微生物可以循环利用固定的碳基质，产生二氧化碳。然而，越来越多的证据表明，许多海洋沉积物也可以作为二氧化碳的汇。为这种“暗”碳固定提供燃料的微生物过程知之甚少。矿物氧化，如铁或硫氧化，被认为是暗碳固定的重要驱动因素，但对这些代谢的不良表征意味着我们缺乏必要的工具来识别和量化这些生理领域。对于常用的基于测序的检测方法来说尤其如此，这种方法依赖于具有标记基因或蛋白质来告知过程的代谢潜力。这项工作是研究不同微生物谱系中矿物氧化的生物标志物，从而扩大我们目前对矿物氧化机制的广度和多样性的理解，以及它们在海洋沉积物中碳循环和固存中的作用。除了这些生物和过程的环境相关性外，氧化固相矿物的能力通常与氧化还原活性表面（如定电位电极）相互作用。因此，了解这些机制也为微生物电极技术的潜力提供了见解。该基金正在研究几种海洋沉积物微生物用于矿物氧化的氧化细胞外电子转移（EET）的基因和途径。所研究的微生物以前是从卡塔利纳港分离出来的，已知具有独特的电化学性质。在基因组上，它们与已知的EET通路缺乏同源性。根据所研究的特定菌株，高通量遗传技术（Tn-Seq）或基因表达研究（RNA-seq）的应用正在用于基因鉴定。在这项工作中发现的假定生物标志物正在遗传学上得到证实。本资助的教育部分使用其中一个菌株作为第二学期入门生物学实验室的课程研究经验。迄今为止进行的生理学和基因组研究表明，EET在各种硫氧化代谢中具有潜在作用。研究硫矿物在支持微生物代谢中的作用，正在使用微生物合成代谢与生物正交非规范氨基酸标记（BONCAT）并行进行光谱技术（拉曼，XRD等）或荧光活化细胞分选耦合宏基因组测序在卡塔利纳港沉积物中进行。由于从沉积物中分离出的许多微生物能够与硫进行岩石营养相互作用，因此假设硫物种在支持微生物代谢和该系统中的暗碳固定中发挥着重要作用-可能使用EET使用或储存固相硫矿物，和/或开发导电网络以连接氧化剂和还原剂之间的空间间隙。该项目由生物海洋学、地球生物学和低温地球化学项目共同资助。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Many marine sediment microbes are known to recycle fixed carbon substrates, producing CO2. However, there is building evidence that many marine sediments can also serve as a sink for CO2. The microbial processes that fuel this “dark” carbon fixation are poorly understood. Mineral oxidation, such as iron or sulfur oxidation, are thought to be important drivers of dark carbon fixation, but poor characterization of these metabolisms means we lack the tools necessary to identify and quantify these physiologies in the field. This is especially true for detection with commonly used sequencing-based approaches, which depend on having marker genes or proteins that inform the metabolic potential for a process. This work is investigating biomarkers for mineral oxidation in distinct microbial lineages, thus expanding our current understanding of the breadth and diversity of mineral oxidation mechanisms, and the role they play in carbon cycling and sequestration in marine sediments. In addition to the environmental relevance of these organisms and processes, the ability to oxidize solid phase minerals, often lends itself to interaction with redox active surfaces such as poised potential electrodes. As such, understanding these mechanisms also provides insight into the potential for microbe electrode technologies. This grant is investigating the genes and pathways used by several marine sediments microbes for oxidative extracellular electron transfer (EET) for mineral oxidation. The subject microorganisms were previously isolated from Catalina Harbor and are known to have unique electrochemical properties. Genomically, they lack homology to known EET pathways. Application of either high throughput genetic techniques (Tn-Seq), or gene expression studies (RNA-seq), depending on the specific strain studied is being used for gene identification. Putative biomarkers identified in this work are being confirmed genetically. The education component of this grant uses one of these strains in a course based undergraduate research experience for a second semester Intro-biology lab. The physiologic and genomic work performed thus far point to a potential role for EET in various sulfur-oxidizing metabolisms. Investigating the role of sulfur minerals in supporting microbial metabolism, is occurring using microbial anabolism with bio-orthogonal non-canonical amino acid tagging (BONCAT) in parallel with either spectroscopic techniques (Raman, XRD, etc.) or fluorescent activated cell sorting coupled to metagenomic sequencing in Catalina harbor sediments. As many of the microbes isolated from this sediment are capable of lithotrophic interaction with sulfur, it is hypothesized that sulfur species are playing an important role in supporting microbial metabolism, and dark carbon fixation in this system—likely using EET to use or storing solid phase sulfur minerals, and/or developing a conductive network to bridge spatial gaps between the oxidants and reductants. This project is cofunded by the Biological Oceanography and Geobiology and Low-temperature Geochemistry Programs.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.