MO: Collaborative Research: Transitions in the Surface Layer and the Role of Vertically Stratified Microbial Communities in the Carbon Cycle- An Oceanic Microbial Observatory

这项提案的重点是浮游细菌微生物群落分层在海洋碳循环中的作用。复杂的生物、化学和物理过程控制着碳从真光带海洋转移到深海的效率，在深海，碳有可能被封存。从真光层输出的大多数有机碳永远不会离开500米的表面，大约90%的输出有机物质在中层带(140-1000米)重新矿化。微生物群落在海洋中垂直分层，特别是在表层(0-300米)，它横跨深层混合事件区域，并从真光层过渡到中上层--碳再矿化活动最高的区域。这一提议的前提是，分层的浮游细菌分支从事可通过综合海洋学和微生物学方法确定的专门的生物地球化学活动。具体地说，这项建议的目的是评估中层微生物群落是否依赖于真光层中微生物过程产生并通过下沉或混合进入中层上层的成岩蚀变有机质和亚细胞碎片。在过去的努力中，该微生物观测站在培养真光区微生物群落成员方面取得了更大的成功，并揭示了SAR11分支的甲蛋白细菌对还原硫化合物的意外生长要求。基因组信息显示，对底物的激烈竞争对浮游细菌施加了权衡--在N维营养空间的某些区域，专家们获胜了。我们推测，特定的生长要求可能解释了在中上层浮游细菌群落中观察到的一些规律性的时空格局，以及培养其中一些生物体的困难。这个项目的具体目标是：1)从光合作用浮游生物培养物中生产13C和15N标记的亚细胞(例如，可溶性、细胞壁和膜)和DOM组分，并使用稳定的同位素探测来确定表层和中层上层微生物群落中吸收不同组成和能力的部分的特定分支。2)使用荧光原位杂交方法监测从SIP和HTC实验中确定的特定微生物种群的时间和空间变异性。为了提高分辨率，我们将使用卡片-鱼协议。3)测定浮游细菌群落的蛋白质组，以确定表层和中上层高度翻译的基因，以及群落对季节性营养限制的反应。4)和，通过高通量培养(HTC)培养这些生物，通过追求他们需要特定的营养因素和/或成岩改变的有机底物的假设。从关键生物中寻找完整的基因组序列，并将其作为查询，研究与生物地球化学重要功能相关的基因和种群的自然变异模式。该项目将使科学界获得新的浮游细菌和基因组序列的培养。这项研究的结果可以直接用于食物链和海洋碳循环模型。这项研究的教育部分通过暑期课程为许多学科的学生提供微生物海洋学培训，并为直接参与研究的研究生和本科生提供专门培训。

The focus of this proposal is the role of bacterioplankton microbial community stratification in the ocean carbon cycle. Complex biological, chemical and physical processes control the efficiency of carbon transfer from the euphotic zone ocean to the deep sea, where sequestration is a possibility. Most organic carbon exported from the euphotic zone never leaves the surface 500 m, with approximately ninety percent of the exported organic matter being remineralized in the mesopelagic zone (140 - 1000 m). Microbial communities are vertically stratified in the oceans, particularly in the surface layer (0-300 m), which spans the region of deep mixing events and transition from the euphotic zone to the upper mesopelagic - the region of highest carbon remineralization activity. The premise of this proposal is that stratified bacterioplankton clades engage in specialized biogeochemical activities that can be identified by integrated oceanographic and microbiological approaches. Specifically, the objective of this proposal is to assess if the mesopelagic microbial community rely on diagenetically altered organic matter and subcellular fragments that are produced by microbial processes in the euphotic zone and delivered into the upper mesopelagic by sinking or mixing. In past efforts this microbial observatory had greater success cultivating members of the euphotic zone microbial community, and revealed an unanticipated growth requirement for reduced sulfur compounds in alphaproteobacteria of the SAR11 clade. Genomic information showed that intense competition for substrates imposes trade-offs on bacterioplankton - there are regions of N dimensional nutrient space where specialists win. We postulate that specific growth requirements may explain some the regular spatial and temporal patterns that have been observed in upper mesopelagic bacterioplankton communities, and the difficulties of culturing some of these organisms.The specific objectives of this project are: 1) to produce 13C and 15N labeled subcellular (e.g., soluble, cell wall, and membrane) and DOM fractions from photosynthetic plankton cultures and use stable isotope probing to identify specific clades in the surface and upper mesopelagic microbial community that assimilate fractions of varying composition and lability. 2) to use fluorescence in situ hybridization approaches to monitor temporal and spatial variability of specific microbial populations identified from the SIP and HTC experiments. To increase resolution we will use CARD-FISH protocols. 3) to measure the proteomes of bacterioplankton communities to identify highly translated genes in the surface layer and upper mesopelagic, and community responses to seasonal nutrient limitation. 4) and, to cultivate these organisms via high throughput culturing (HTC) by pursuing the hypothesis that they require specific nutrient factors and/or diagenetically altered organic substrates. Complete genome sequences from key organisms will be sought and used as queries to study patterns of natural variation in genes and populations that have been associated with biogeochemically important functions.This project will make cultures of novel bacterioplankton and genome sequences available to the scientific community. Findings from this research may be used directly in foodweb and ocean carbon cycle models. The educational component of this research brings microbial oceanography training to students from many disciplines, through a summer course, and specialized training to graduate and undergraduate students involved directly in research.