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

这项研究项目将确定在日本大阪湾和Muko River消耗氨基酸的微生物种群。这一目标将使用拉曼显微光谱来测量从环境中提取氨基酸并将其用于生长的细胞。溶解氨基酸是水生微生物产生能量和生物量的重要有机物质来源，因此，微生物对这一底物的周转和循环是世界各地海岸边缘碳循环的关键步骤。这项研究将与日本神户关西学园大学著名的拉曼光谱生物和化学分析专家Shinsuke Shigeto博士合作进行。很少有实验室配备拉曼显微光谱仪来扫描活的生物细胞，因此这次合作是将PI项目和日本合作者的专业知识结合在一起的独特机会。这项研究项目将测量13C标记氨基酸(AAs)在日本大阪湾和木库河自然微生物群落中单个浮游细菌细胞生物量中的掺入情况。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相结合的方法来实现。在13C标记的氨基酸与群落孵育后，将通过红移测量13C标记的AAs对优势种群单个细胞生物量的掺入。它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技术，从利用特定底物的自然微生物群落中识别单个细胞。原核生物的16S rRNA基因扩增也将在孵化前后进行测序，以将群落水平的数据与种群和单细胞水平的AA掺入数据相关联。该奖项由东亚和太平洋暑期学院项目资助一名美国研究生的暑期研究，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.