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会循环利用固定的碳底物，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。人们对这种“黑暗”固碳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，但对这些代谢过程的糟糕描述意味着我们缺乏必要的工具来在该领域识别和量化这些生理现象。对于常用的基于测序的方法的检测尤其如此，这种方法依赖于具有告知过程新陈代谢潜力的标记基因或蛋白质。这项工作正在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的碳循环和封存中所起的作用。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有助于与氧化还原活性表面(如稳定的电位电极)相互作用。因此，了解这些机制也有助于洞察微生物电极技术的潜力。这笔赠款正在研究几种海洋沉积物微生物用来进行氧化细胞外电子转移(EET)以进行矿物氧化的基因和途径。研究对象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。从基因上看，它们与已知的EET途径缺乏同源性。高通量遗传技术(TN-SEQ)或基因表达研究(RNA-SEQ)的应用，取决于所研究的特定菌株，正被用于基因鉴定。这项工作中确定的假定生物标记物正在得到基因确认。这笔助学金的教育部分在基于课程的本科生研究经验中使用了其中一种菌株，用于第二学期的生物入门实验室。到目前为止进行的生理和基因组工作表明，EET在各种硫氧化代谢中具有潜在的作用。研究硫矿物在支持微生物代谢中的作用，正在利用微生物合成代谢与生物正交非典型氨基酸标记(BONCAT)结合任何一种光谱技术(拉曼、X射线衍射等)进行。或在卡塔利纳港沉积物中将荧光激活的细胞分类与元基因组测序相结合。由于从该沉积物中分离出的许多微生物能够与硫相互作用，推测硫物种在支持微生物新陈代谢和暗碳固定方面发挥着重要作用-可能使用EET来使用或储存固相硫矿物，和/或发展导电网络来弥合氧化剂和还原剂之间的空间差距。该项目由生物海洋学和地球生物学以及低温地球化学项目共同资助。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势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.