EAPSI: Defining Resource Preferences of Single Cells from Aquatic Microbial Communities using Raman Microspectroscopy

该研究项目将确定日本大坂湾和Muko河中消耗氨基酸的微生物种群。这一目标将通过使用拉曼显微光谱来测量从环境中获取氨基酸并用于生长的细胞来实现。溶解氨基酸是水生微生物生产能量和生物质的重要有机物质来源，因此，微生物对这种底物的周转和再循环是世界各地海岸边碳循环的关键步骤。这项研究将与Shinsuke Shigeto博士合作进行，Shinsuke Shigeto博士是日本科比关西学院大学使用拉曼光谱进行生物和化学分析的著名专家。很少有实验室配备了拉曼显微光谱仪来扫描活的生物细胞，因此这次合作代表了一个独特的机会，联合收割机结合项目PI和日本合作者的专业知识。该研究项目将测量13 C标记的氨基酸（AAs）掺入到来自日本大坂湾和Muko河的天然微生物群落中的单个浮游细菌细胞的生物量中。这一目标将使用稳定同位素探测，荧光原位杂交和拉曼显微光谱的组合来实现。孵育后的社区与13 C-标记的氨基酸，掺入的同位素到生物量的个别细胞从占主导地位的人口将被测量？红移在他们的拉曼光谱中的苯丙氨酸峰。这项研究将开发一种非破坏性的技术，以确定单个细胞从一个自然的微生物群落，利用特定的基板。还将在孵育前后对prostaglandin 16 S rRNA基因扩增子进行测序，以将群落水平数据与群体和单细胞水平AA掺入数据相关联。东亚和太平洋夏季研究所计划下的这个奖项支持美国研究生的夏季研究，由NSF和日本科学促进会共同资助。

This research project will determine the microbial populations that are consuming amino acids in Osaka Bay and Muko River, Japan. This objective will be accomplished using Raman microspectroscopy to measure cells that have taken amino acids from the environment and used them for growth. Dissolved amino acids are an important organic matter source for both energy and biomass production in aquatic microorganisms, thus, turnover and recycling of this substrate by microbes is a critical step in the carbon cycle of coastal margins around the world. This research will be conducted in collaboration with Dr. Shinsuke Shigeto, a noted expert in biological and chemical analysis using Raman spectroscopy, at Kwansei Gakuin University in Kobe, Japan. Few laboratories are equipped with a Raman microspectrometer set up to scan living biological cells, thus this collaboration represents a unique opportunity to combine the expertise of the project PI and the Japanese collaborator. This research project will measure the incorporation of 13C-labeled amino acids (AAs) into the biomass of single bacterioplankton cells in natural microbial communities from Osaka Bay and the Muko River, Japan. This objective will be accomplished using a combination of stable isotope probing, fluorescence in-situ hybridization, and Raman microspectroscopy. After incubating communities with 13C-labeled AAs, incorporation of the isotope into biomass of individual cells from dominant populations will be measured by a ?red-shift? of the phenylalanine peak in their Raman spectra. This study will develop a non-destructive technique to identify single cells from a natural microbial community that utilize a specific substrate. Prokaryotic 16S rRNA gene amplicons will also be sequenced before and after incubations to correlate community-level data with population- and single-cell level AA incorporation data. This award under the East Asia and Pacific Summer Institutes program supports summer research by a U.S. graduate student and is jointly funded by NSF and the Japan Society for the Promotion of Science.