Development of Bioluminescent Bacterial Bioreporters to Quantify the Bioavailability of Fe in Seawater

虽然铁在海洋系统中作为初级生产力限制剂的作用已经被广泛研究，但海洋细菌对铁有效性变化的反应(S)仍然不太清楚。许多小组的研究表明，许多海洋原核生物保持产生铁载体(或至少铁载体类化合物)的能力，以响应铁限制的生长条件，因此在铁可获得性低的条件下可以有效地持续存在。虽然分析技术可以在极小的浓度下量化全铁，并提供有关铁的形态和铁与有机配体结合/解离的动力学的信息，但没有指标表明不同形式的铁对自然群落的生物有效性。为了了解铁在全球海洋生产力中的作用，需要一种基于细菌铁感应系统的工具。出于这个原因，田纳西大学诺克斯维尔分校的一名研究人员将开发、表征和校准一系列异养细菌生物发光生物记者，这些生物记者可以定量地响应环境铁生物有效性的变化。在构建了一系列新的报告系统，并用构成荧光蛋白标记(以便在自然海洋微生物群存在的情况下对生物报告细胞进行计数)后，将在新西兰东南部的亚热带辐合高营养-低叶绿素带进行田间试验，以验证该报告的野外使用，并表征天然溶解有机物对不同细菌的生物有效性的影响。与蓝藻中正在开发的类似工具进行并列比较，将有助于理解模型异养铁获取策略和光营养铁获取策略之间的差异。目标是开发和验证一套工具，它将直接代表海洋细菌的一个子集，并在实验室研究中与所有可培养的海洋细菌进行比较，并在野外研究中与在不同地点发现的变异种群进行比较。就更广泛的影响而言，这项研究将为海洋界提供一种生物传感器工具，能够确定异养细菌群落对铁的生物有效性，这是了解海洋中铁的生物地球化学循环的关键组成部分。预计一旦完成，生物报告菌株将通过环境生物技术培养收集中心提供给合格的研究人员。作为该项目的一部分，两名研究生和两名本科生将在这个促进竞争研究(EPSCoR)机构的实验计划中接受培训。

ABSTRACTOCE-0526159While the role of iron as a limiting agent of primary productivity in marine systems has been extensively studied, the response(s) of marine bacteria to changes in iron availability remain less clear. Studies from numerous groups suggest that many marine prokaryotes maintain the ability to produce siderophores (or at least siderophores-like compounds) in response to iron-limiting growth conditions and as such can effectively persist under conditions of low iron availability. While analytical techniques can quantify total iron at incredibly small concentrations and provide information on iron speciation and the kinetics of iron association/dissociation with organic ligands, there are no indicators of the bioavailability of iron in different forms to natural communities. To understand the role that iron plays in global marine productivity, a tool based on the iron-sensing systems of bacteria is needed. For this reason, a researcher from the University of Tennessee at Knoxville will develop, characterize, and calibrate a series of heterotrophic bacterial bioluminescent bioreporters that quantitatively respond to changes in ambient iron bioavailability. After the construction of a series of new reporter systems, and the tagging of these bioreporters with a constitutive fluorescent protein marker (so that bioreporter cells can be enumerated in the presence of the natural marine microbial population), field tests will be carried out in the subtropical convergence high nutrient-low chlorophyll zone southeast of New Zealand to validate the field use of this reporter and characterize the effects of natural dissolved organic matter of iron bioavailability to different bacteria. Side-by-side comparisons with similar tools being developed in cyanobacteria will provide an understanding of the differences between model heterotrophic and phototrophic iron acquisition strategies. The goal is to develop and validate a tool set that will directly represent a subset of marine bacteria and serve as a comparative in laboratory studies with all cultivable marine bacteria, and in field studies with the variant populations found in diverse locations.In terms of broader impacts, this study will provide the oceanographic community with a biosensor tool capable of determining iron bioavailability to heterotrophic bacterial communities, a key component towards understanding the biogeochemical cycle of iron in the ocean. It is anticipated that once completed, the bioreporter strains will be available to qualified researchers through the center for Environmental Biotechnology culture collection. Two graduate and two undergraduate student will be trained at this Experimental Program to Stimulate Competitive Research (EPSCoR) institution as part of this project.