Activity and Characteristics of the Dissolved Organic Carbon-Dependent Picoplankton over the Annual Cycle in a Subarctic Lake

首席研究员将在北极附近的一个湖泊进行全年周期的现场测量，由于太阳角度和积雪的巨大变化，光合作用速率从0到500 mg C/m2每天变化。这项工作将重点关注浮游细菌在冬季/夏季过渡期间的变化。细菌的活性将由底物摄取来决定，通常是氨基酸混合物。测量将包括人工增温和降温，以确定活化能，从而评估温度偏好。孵育将包括辅助底物，如葡萄糖，以帮助确定在所收集的样品中生物体的通电状态。这些活化能是通过流式细胞术结合单个细胞的膜电位测量来确定的。伴随的测量将是通过流式细胞术计算特定亲和力的细胞群和质量。细胞质量和DNA/细胞的分布被用作生物体在季节中活动的进一步指标。在不同季节消耗的基质的细胞产量将用于帮助确定生产力和内源性需求，特别是在冬季。这些与溶解的游离氨基酸浓度有关，作为不稳定有机底物的指示。进一步的鉴定将通过DNA片段和序列分析。将进一步开发现有的分离物培养集合，以寻找和表征典型的低DNA，小型，典型的淡水寡菌进行生理研究。培养物用于确定特定亲和力理论中缺失项的值，例如底物摄取阈值的大小和细胞质池中运输底物的浓度。扩展的统计理论用于测试细胞-细胞相互作用将应用于提高目前达到的小（1%）培养率的目标。这项工作涉及应用新的理论和方法来理解北极的微生物过程，特别是水温的季节性变化。这将有助于提高对细菌活性和生产力的估计。新的灭绝培养程序与新的统计和动力学理论相结合，可以帮助巩固对我们最大的活性生物物质形式之一的稀疏理解。发展中的动力学理论开始改变范式，定量流式细胞术和营养通量计算也开始改变范式，从而改变了对寡菌生理学的认识。这项新技术将通过研究生和本科生的课程和研讨会推广到广泛的学生群体中。这项工作的广泛影响包括新概念的发展，例如将碳和营养循环的各个方面与生物体活动、生理学、营养浓度和温度联系起来的方法。组分技术的发展，如定量流式细胞术和灭活培养，一般适用于微生物学。最重要的是，这项工作将为Michaelis Menten范式提供一个机械上有效的替代方案，以改进微生物活动和动力学的描述和比较。

ABSTRACT ButtonOPP-0243913The Principal Investigator will make in situ measurements made throughout the annual cycle in a near arctic lake where photosynthesis rates vary from zero to 500 mg C/m2 per day due to large changes in sun angle and snow cover. The work will focus on changes in the bacterioplankton over the winter/summer transition. Bacterial activity will be determined by substrate uptake, usually an amino acid mixture. Measurements will include artificial warming and cooling to determine activation energies from which temperature preferences are evaluated. Incubations will include an auxiliary substrate such as glucose to help determine the energization state of organisms in the collected sample. These activation energies are determined in conjunction with membrane potential measurements of the individual cells by flow cytometry. Companion measurements will be cell population and mass by flow cytometry for calculating specific affinities. The distribution of cell mass and DNA/cell is used as a further index of organism activity over the seasons. Cell yields from substrate consumed over the seasons will be used to help determine productivity and endogenous requirements, particularly during winter. These are linked to dissolved free amino acid concentrations as an indication of labile organic substrates. Further characterization will be by DNA fragment and sequence analyses. Existing culture collections of isolates will be further developed with the objective of finding and characterizing a typical low DNA, small, typical freshwater oligobacterium for physiological study. Cultures are used to determine values for missing terms in specific affinity theory such as the size of thresholds for substrate uptake and the concentration of transported substrate in cytoplasmic pools. Expanded statistical theory for testing cell-cell interactions will be applied with the objective of improving the small (1%) culturability presently attained. This work involves the application of novel theory and methods to the understanding of microbial processes in the Arctic with particular reference to seasonal changes in water temperature. These will serve to improve estimates of bacterial activity and productivity. Novel extinction culture procedures coupled with new statistical and kinetic theory can help solidify the sparse understanding of one of our largest forms of active living material. Developing kinetic theory is beginning to alter paradigms, as is quantitative flow cytometry together with nutrient flux calculations, and is thereby altering perceptions of oligobacterial physiology. The novel technology will be extended to a wide cross section of students through graduate and undergraduate courses and seminars. The broader impacts of the work include the development of novel concepts, such as ways of linking aspects of the carbon and nutrient cycle to organism activity, physiology, nutrient concentrations, and temperature. Developments in component technologies, such as quantitative flow cytometry and extinction culturing, are applicable to microbiology in general. Most importantly the work will provide a mechanistically competent alternative to the Michaelis Menten paradigm to improve description and comparison of microbial activity and dynamics.