Small Scale Spatial and Temporal Variations in Bacterial Activity in Marine Waters

海洋中的细菌通过呼吸作用将溶解的有机物转化为二氧化碳，并转化为可供微生物食物网中更高营养水平使用的生物量，在这一过程中发挥着关键作用。它们是海洋中生物量的主要形式。从生产力高的沿海和上升流地区到营养极少的水域，细菌丰度分别仅在约10至10毫升之间。然而，这些细菌的活性差异要大得多。通常检查的米到公里的空间尺度可能不适合小于10-6英寸的细胞，并且可能出现在10-5到10-3M的尺度上的斑块。微尺度的斑块可以为细胞提供比散装方法测量的更高的有机营养浓度，导致对细菌对物理和生物地球化学扰动的响应的非常不同的预测。在这个项目中，我们建议使用自动化的流式细胞术和成像细胞术与敏感的荧光方法相结合来在小的空间和时间尺度上测量海洋细菌的单细胞组成和活性。将使用各种单细胞荧光探针来指示细胞组成，如蛋白质和核酸含量，以及生理过程，如呼吸、膜电位和酶活性。这些单细胞反应将使用自动化的流式和成像细胞术方法进行量化，以提高灵敏度和计数精度、每细胞荧光定量测量和客观检测-所有这些都是相对于目视显微镜方法的改进。我们的实验室在这些方法方面拥有独特的设备和经验。我们将对散装和单电池方法进行比较。细菌活动的微尺度斑块(以及丰度)将在一系列空间尺度和自然湍流混合条件下进行检查。将在季节周期和营养梯度上测量活跃细菌种群和总细菌种群的大小分布，以提供浮游生物群落结构的各种制度。还将研究细菌活性的短期变化，以确定细菌活性在周期内如何变化。实验将在季节性时间尺度上进行，并在从多产的沿海水域到贫营养的蓝色水域地点的游轮上进行。通过在适当的小空间和短时间尺度上测量单细胞活动，但跨越温带海洋的主要生态梯度，我们将对控制细菌生物地球化学过程的机制有重要的了解。

Bacteria in the ocean play a key role in transforming dissolved organic matter into C02 through respiration and into biomass that is available to higher trophic levels within the microbial food web. They are the dominant form of biomass in the ocean. From highly productive coastal and upwelling regions to extremely oligotrophic waters, bacteria abundances range only between about 10 to 10 ml, respectively. The activity of these bacteria, however, varies much more widely. Typically examined spatial scales of meters to kilometers may not be appropriate for cells that are smaller than 10-6 in and may occur in patches on the scale of 10-5 to 10-3 M. Microscale patchiness can provide concentrations of organic nutrients to cells much higher than those measured by bulk methods, leading to very different predictions of bacterial responses to physical and biogeochemical perturbations. In this project we propose to use automated flow and imaging cytometry combined with sensitive -fluorescence methods to measure the single-cell composition and activity of marine bacteria at small spatial and diel temporal scales. A variety of single-cell fluorescent probes will be used to indicate cell composition such as protein and nucleic acid content, and physiological processes such as respiration, membrane potential, and enzyme activity. These single-cell responses will be quantified using automated flow and imaging cytometric methods for increased sensitivity and count precision, quantitative fluorescence per cell measurement, and objective detection - all improvements over visual microscope methods. Our laboratory is uniquely equipped and experienced in these approaches. Bulk and single- cell methods will be compared. Microscale patchiness of bacterial activity (as ell as abundance) will be examined over a range of spatial scales and natural turbulent mixing conditions. The size distributions of active and total bacterial populations will be measured over seasonal cycles and across trophic gradients providing a variety of regimes of plankton community structure. Short time variations in bacterial activity will also be studied to determine how bacterial activity varies over diel cycles. Experiments will be conducted over seasonal time scales and on cruises from productive, coastal waters to oligotrophic, blue water sites. By measuring single-cell activities at appropriately small spatial and short temporal scales, but across the dominant ecological gradients of the temperate ocean we will gain significant insight into the mechanisms controlling bacterial biogeochemical processes.