OCE-PRF Track 1: Resolving the Advantages of Motility and Chemotaxis in Oceanic Crust

海洋地壳构成了一个巨大的循环流体含水层，是一个重要的，但相对未开发的生物圈。从胡安·德富卡海岭（JFR）东侧采集的地壳流体样本的基因组观察揭示了丰富的运动和趋化基因。该项目的目标是研究这些独特的地下生物体的趋化潜力，利用上述基因组数据集，以便在实验室中分离和详细研究能动微生物。该项目的预期意义将是从根本上了解地下海洋地壳中微生物生命的适应策略，这将进一步影响对这一广阔环境中生物地球化学循环的解释。推广工作包括培训来自STEM领域代表性不足的群体的本科生掌握新的基因组技术，并在社区研讨会上和通过K-12教育项目与公众分享成果。运动被视为一种能源昂贵的功能，对于在能源匮乏的环境中生存的生物体来说是不切实际的。尽管如此，富集文化已经建立了从地壳流体的JFR基因组数据和代谢的解释的基础上。显微镜检查证实了这些富集物中存在活性能动生物。这些培养物代表了如何利用基因组解释来确定实验室生长的营养需求的一个典型例子。本研究的目的包括：（1）在纯培养中分离运动细胞，并对这些纯分离物的基因组进行测序，以识别它们在地壳环境中的代谢潜力。鉴于这些富集培养物是第一批成功从地下地壳流体中生长出来的，任何生理或代谢观察对于了解大洋地壳中的生命都将是新颖和重要的。第二个目的是（2）调查哪些化学引诱剂对目标1中分离的培养物最有吸引力。将在装有含硬琼脂营养塞的0.15%低熔点琼脂糖的梯度管中观察趋化反应。趋化反应将评价为朝向或远离硬化学引诱物栓的生长。本研究的最终目的是测量地壳流体生物在（3）最佳生长条件和（4）原位JFR营养条件下的营养消耗速率、它们的平均和最大速度以及它们对营养的反应时间。将使用定制的微流体显微镜载玻片研究运动性。该载玻片的微通道将填充根据目标1和2确定的各种营养引诱剂。将使用倒置明视野显微镜监测细胞行为和对引诱剂的运动性，并进行数字记录以计算和模拟2D速度。总的来说，这套独立但互补的实验室和基因组分析旨在更好地了解海洋地壳运动的作用，最终解决在这种环境中生存的独特适应性，并推进海洋地壳地球化学循环的知识。

The oceanic crust constitutes a vast aquifer of circulating fluids and represents a significant, but relatively unexplored biosphere. Genomic observations of crustal fluids sampled from the eastern flank of the Juan de Fuca Ridge (JFR) have revealed an abundance of motility and chemotactic genes. The goal of this project is to investigate the chemotactic potential of these unique subsurface organisms, leveraging the aforementioned genomic datasets to enable the isolation and detailed study of motile microorganisms in the laboratory. The expected significance of this project will be the fundamental understanding of adapted strategies of microbial life in subsurface oceanic crust, which will further impact interpretations of biogeochemical cycling in this vast environment. Outreach efforts include training undergraduate students from underrepresented groups in STEM in novel genomic techniques and sharing results with the public at community seminars and through K-12 educational projects.Motility is viewed as an energy expensive function and impractical for organisms surviving in energy starved environments. Nevertheless, enrichment cultures have already been established from crustal fluids of the JFR based on genomic data and metabolic interpretations. Microscopy confirms the presence of active motile organisms within these enrichments. These cultures represent a prime example of how genomic interpretations can be utilized to determine nutrient requirements for laboratory growth. The objectives of this study include (1) isolating motile cells in pure culture, and sequencing the genomes of these pure isolates in order to recognize their metabolic potential within crustal environments. Given that these enrichment cultures are some of the first to be successfully grown from subsurface crustal fluids, any physiological or metabolic observations will be novel and important for understanding life in the oceanic crust. The second objective is to (2) investigate which chemical attractants are most attractive to the cultures isolated in objective 1. Chemotactic response will be observed within gradient tubes filled with 0.15% low-melt agarose containing hard agar nutrient plugs. Chemotactic responses will be evaluated as growth towards or away from the hard chemoattractant plug. The final objectives of this study are to measure the rates of nutrient consumption by crustal fluid organisms, their average and maximum speeds, and their reaction time to nutrients under (3) optimal growth conditions and (4) in situ JFR nutrient conditions. Motility will be studied using a custom microfluidic microscope slide. Micro-channels of this slide will be filled with the various nutrient attractants determined from objectives 1 and 2. Cell behavior and motility towards attractants will be monitored using an inverted brightfield microscope and digitally recorded to calculate and model 2D velocities. Collectively, this suite of independent, yet complementary, laboratory and genomic analyses are aimed to better understand the role of motility in the oceanic crust, ultimately resolving unique adaptations for survival in this environment and advancing the knowledge of geochemical cycles in oceanic crust.