Quantifying the contribution of the deep biosphere in the marine sediment carbon cycle using deep-sea sediment cores from the Baltic Sea

海洋沉积物中包含的微生物数量足以与地球海洋的微生物数量相媲美，但关于这个庞大的群落的许多情况尚不清楚。细胞总数计数方法的创新改进了对细胞浓度的估计，但对具体的分类群一无所知。同位素数据提供的证据表明，大多数地下微生物通过分解有机物生存，但特定微生物类群与其有机物质底物之间的可测量联系尚未得到测试。拟议的工作克服了这些限制，特别侧重于蛋白质和碳水化合物的降解，它们构成了可分类沉积有机质的主体。该项目将把特定的类群与单个微生物细胞基因组中潜在的胞外酶活性联系起来，应用新发现的最佳方法，使用催化报告沉积荧光原位杂交(CARD-FISH)来计数属于特定类群的活细胞，并原位测量它们的酶的潜在活性。由此产生的数据将为地下生命用来克服极端能源限制和促进长期碳循环的策略提供关键证据。首席调查人员正在使用新的、改进的方法来量化海洋次表层中特定分类群的细胞，并确定那些未培养的分类群的生物地球化学功能，包括：1)确定未培养的、数量占优势的次表层微生物的单个细胞基因组中的有机碳降解途径。2)利用对卡片鱼现有技术的改进，对深层地下的活细菌和古菌进行量化。3)测量波罗的海深海沉积物中酶的潜在活性(Vmax值)，并利用产酶微生物的丰度来计算特定细胞Vmax的深度剖面。该项目将这些方法结合在一起，以识别和量化波罗的海深层沉积物中能够降解有机物质的细胞，这些细胞是从综合海洋钻探计划(IODP)探险347获得的。这些结果将极大地扩展我们对地下深层未培养微生物的功能和活性的认识。

Marine sediments contain a microbial population large enough to rival that of Earth's oceans, but much about this vast community is unknown. Innovations in total cell counting methods have refined estimates of cell concentrations, but tell us nothing about specific taxa. Isotopic data provides evidence that a majority of subsurface microorganisms survive by breaking down organic matter, yet measurable links between specific microbial taxa and their organic matter substrates are untested. The proposed work overcomes these limitations, with a particular focus on the degradation of proteins and carbohydrates, which comprise the bulk of classifiable sedimentary organic matter. The project will link specific taxa to potential extracellular enzyme activity in the genomes of single microbial cells, apply newly-identified, optimal methods for counting viable cells belonging to specific taxa using catalyzed reporter deposition fluorescent in situ hybridization (CARD-FISH), and measure the potential activity of their enzymes in situ. The resulting data will provide key evidence about the strategies subsurface life uses to overcome extreme energy limitation and contribute to the long-term carbon cycle. The Principal Investigators are employing novel,improved methods to quantify cells of specific taxa in the marine subsurface and to determine the biogeochemical functions of those uncultured taxa, including: 1)Determine the pathway of organic carbon degradation in single cell genomes of uncultured, numerically dominant subsurface microorganisms. 2)Quantify viable bacteria and archaea in the deep subsurface using an improvement on the existing technology of CARD-FISH. 3)Measure the potential activities (Vmax values) of enzymes in deep Baltic Sea sediments, and use the abundances of enzyme-producing microorganisms to calculate depth profiles of cell-specific Vmax values. The project combines these methods in order to identify and quantify the cells capable of degrading organic matter in deep sediments of the Baltic Sea, obtained from Integrated Ocean Drilling Program (IODP) expedition 347. These results will greatly expand our knowledge of the function and activity of uncultured microorganisms in the deep subsurface.