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

摘要OPP-0243913首席调查员将在北极附近的一个湖泊进行整个年度周期的现场测量，由于太阳角和积雪的巨大变化，该湖泊的光合作用速率每天从零到500毫克C/m2。这项工作将集中在冬季/夏季过渡期间浮游细菌的变化。细菌的活性取决于底物的摄取，通常是氨基酸混合物。测量将包括人工升温和降温，以确定评估温度偏好的激活能。培养将包括一种辅助底物，如葡萄糖，以帮助确定收集的样本中有机体的通电状态。这些激活能是通过流式细胞仪结合单个细胞的膜电位测量来确定的。随之而来的是用流式细胞术测量细胞数量和质量，以计算特定的亲和力。细胞质量和DNA/细胞的分布被用来作为生物在季节中活动的进一步指标。四季消耗的底物的细胞产量将被用来帮助确定生产力和内源需求，特别是在冬季。这些与溶解的游离氨基酸浓度有关，作为不稳定的有机底物的指示。进一步的鉴定将通过DNA片段和序列分析来进行。将进一步开发现有的分离物培养库，目的是寻找和鉴定一种典型的低DNA、小、典型的淡水寡杆菌用于生理研究。培养用于确定特定亲和力理论中缺失的项的值，例如底物吸收阈值的大小和细胞质池中运输的底物的浓度。将应用扩展的统计学理论来测试细胞与细胞之间的相互作用，目标是提高目前达到的小(1%)可培养能力。这项工作涉及应用新的理论和方法来了解北极的微生物过程，特别是水温的季节变化。这些将有助于改进对细菌活性和生产力的估计。新的灭绝文化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形式的活跃生命材料之一的稀疏理解。发展的动力学理论开始改变范式，就像定量的流式细胞术和营养通量计算一样，从而改变了对寡菌生理学的看法。这项新技术将通过研究生和本科生课程和研讨会推广到更广泛的学生群体中。这项工作的更广泛影响包括新概念的发展，例如如何将碳和营养循环的各个方面与生物体活动、生理、营养浓度和温度联系起来。组分技术的发展，如定量流式细胞术和消光培养，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合格的替代方案，以改进对微生物活动和动态的描述和比较。

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.