Collaborative Research: Next generation physiology: a systems-level understanding of microbes driving carbon cycling in marine sediments

合作研究：下一代生理学：对驱动海洋沉积物碳循环的微生物的系统级理解

• 批准号: 1817381
• 负责人: Andreas Teske
• 金额: $ 26.78万
• 依托单位: University of North Carolina at Chapel Hill
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2022-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1817381&HistoricalAwards=false
• 关键词: Collaborative; Research; Next; generation; physiology

It is currently impossible to grow the vast majority (99%) of deep-sea microbes under laboratory conditions. Because of this recalcitrancy of most microbes to cultivation, linking specific genotypes with their in situ phenotypes has been a problem in microbiology for decades - it has been very difficult to figure out which members of microbial communities are doing what, due to the complexity of these communities. The new techniques used in these studies allow for a relatively straightforward labeling of the microbial cells that are active under different conditions, allowing for these "next generation physiology" techniques to be used to identify the individual cells actively consuming different compounds. In this technique metabolically active cells are labeled with aa compounds with the detection of these cells by fluorescence staining. This allows for the metabolism of hundreds of taxa to be followed under dozens of different conditions in parallel. These studies will focus on the hydrothermally active sediments of Guaymas Basin in the Gulf of California. There, a broad range of phylum-level lineages of uncultured archaea and bacteria are hypothesized to engage in the degradation of complex organic molecules, including hydrocarbons. The microbes that assimilate and break down these molecules will be identified, and their metabolism will be correlated with their genomes, allowing for an understanding of how these microbes are able to live on a diet of hydrocarbons, and helping us to further understand how these compounds break down in the environment. The investigators make use of topical blogs, public talks and magazine articles to educate and inform the general public on the importance of microbes in creating the world around us, and their laboratories will provide undergraduate researchers with exciting research opportunities. The aim of this project is to investigate physiological controls of microbial carbon cycling in marine sediments and to link specific genotypes with their in situ phenotype. The investigators recently developed a new, highly parallelizable, and adaptable approach that allows for the studying microbial function and metabolic interactions in uncultured microbes at unrivaled throughput. To achieve this goal they combine the (1) labeling of translationally active cells via bioorthogonal non-canonical amino acid tagging (BONCAT), (2) sorting of individual, active cells by fluorescence activated cell sorting, and (3) study of the identity and genetic makeup of active cells by massive parallel gene sequencing. This results in a readout of the translational activity of hundreds of uncultured taxa in parallel, which allows for the functional activity of hundreds of taxa under dozens of different conditions to be determined. Guaymas Basin sediments were chosen for these studies due to their steep gradients in temperature, variety of electron donor identity and availability, and the presence of hydrocarbon seeps supplying aliphatic and aromatic hydrocarbons to the system. This means that these sediments are likely to have highly diverse metabolic potential, and their study will allow for a deeper understanding of carbon-cycling and other geochemical processes in marine sediments. The researchers will identify those microbes in environmental samples that are involved in hydrocarbon, complex substrate and heterotrophic/autotrophic activity, and follow this with targeted whole genome shotgun sequencing to directly identify the microbes actively utilizing those lifestyles, and provide the genetic makeup of cells that are able to exploit specific substrates. This work will provide a benchmark study for the developing field of deep-sea sediment microbiology and lay the foundation for future physiological studies in environmental microbiology.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.

目前不可能在实验室条件下培养绝大多数（99%）深海微生物。由于大多数微生物对培养的这种不确定性，几十年来，将特定基因型与其原位表型联系起来一直是微生物学中的一个问题-由于这些群落的复杂性，很难弄清楚微生物群落的哪些成员在做什么。这些研究中使用的新技术允许在不同条件下活跃的微生物细胞的相对直接的标记，允许这些“下一代生理学”技术用于识别活跃消耗不同化合物的单个细胞。在该技术中，代谢活性细胞用aa化合物标记，并通过荧光染色检测这些细胞。这允许在几十种不同的条件下并行地跟踪数百个分类群的代谢。这些研究将集中在加州湾瓜伊马斯盆地的热液活动沉积物。在那里，一个广泛的门级谱系的未培养的古菌和细菌被假设从事复杂的有机分子，包括碳氢化合物的降解。同化和分解这些分子的微生物将被识别，它们的代谢将与它们的基因组相关，从而了解这些微生物如何能够以碳氢化合物为食，并帮助我们进一步了解这些化合物如何在环境中分解。研究人员利用专题博客，公开讲座和杂志文章来教育和告知公众微生物在创造我们周围世界的重要性，他们的实验室将为本科研究人员提供令人兴奋的研究机会。该项目的目的是调查海洋沉积物中微生物碳循环的生理控制，并将特定基因型与其原位表型联系起来。研究人员最近开发了一种新的，高度并行化和适应性强的方法，可以以无与伦比的通量研究未培养微生物中的微生物功能和代谢相互作用。为了实现这一目标，他们结合了联合收割机：（1）通过生物正交非规范氨基酸标记（BONCAT）标记免疫活性细胞，（2）通过荧光激活细胞分选对单个活性细胞进行分选，以及（3）通过大规模平行基因测序研究活性细胞的身份和遗传组成。这导致并行地读出数百个未培养的分类群的翻译活性，这允许确定数百个分类群在数十种不同条件下的功能活性。选择瓜伊马斯盆地沉积物进行这些研究，是因为它们的温度梯度很陡，电子供体的身份和可用性多种多样，以及存在向系统提供脂肪族和芳香族烃的烃渗漏。这意味着这些沉积物可能具有高度多样的代谢潜力，对它们的研究将有助于更深入地了解海洋沉积物中的碳循环和其他地球化学过程。研究人员将识别环境样本中参与碳氢化合物，复杂底物和异养/自养活动的微生物，并通过有针对性的全基因组鸟枪测序直接识别积极利用这些生活方式的微生物，并提供能够利用特定底物的细胞的遗传组成。这项工作将为发展中的深海沉积物微生物学领域提供基准研究，并为今后环境微生物学的生理学研究奠定基础，该奖项反映了国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

项目成果

Large-scale protein level comparison of Deltaproteobacteria reveals cohesive metabolic groups

• DOI: 10.1038/s41396-021-01057-y
• 发表时间: 2021-07-30
• 期刊: ISME JOURNAL
• 影响因子: 11
• 作者: Langwig, Marguerite V.;De Anda, Valerie;Baker, Brett J.
• 通讯作者: Baker, Brett J.