Linking Resource and Stress Gradients to Microbial Community Composition and Function through the Soil Profile of a California Annual Grassland at the Sedgwick Reserve

尽管最近环境微生物学取得了进展，但我们对控制微生物群落组成和动态的因素的理解仍然有限，对群落组成和功能之间的联系也没有很好的理解。这一方向的发展将来自自上而下（过程）和自下而上（分子多样性）方法的融合，并与现有的生态学理论相结合。植物生态学的中间干扰和资源供应假说应通过土壤剖面适用于微生物群落，因为资源有效性和压力在土壤表面都很高，但随深度而下降。压力和干扰也可以部分解释为什么在很少取样于10厘米以下的研究中很难将土壤多样性与功能联系起来。土壤要深得多，在深度处有大量的微生物种群，占整个剖面活动的很大一部分。这个微生物观察站的中心假设是，资源和压力的梯度是整个土壤剖面中微生物群落的大小、多样性、动态、活性和生长策略（生物膜形成）的主要驱动因素。该项目将在塞奇威克保护区进行，这是一个积极合作的地点，研究加州一年生草地/橡树热带稀树草原生态系统的景观过程、土壤动力学、植物群落结构和土壤微生物过程。工作将把整个保护区的广泛采样与两个核心地点的密集操作研究结合起来，在那里监测土壤气候、化学和至少4米深的群落，并建立操作研究。将监测土壤湿度和温度，以评估应力梯度，而基材梯度将通过物理土壤有机质分异和生物测定方法的组合进行分析。将测量微生物生物量，以及细菌和真菌的总数和活跃数量以及生物体积。通过DGGE（变性梯度凝胶电泳）和t-RFLP（末端标记限制性片段长度多态性）配对分析细菌群落结构和组成。DGGE的优点是可以表征单个条带并开发特定的探针，而t-RFLPs可以更清晰地分析复杂的DNA混合物。这些指纹识别方法将通过开发针对特定细菌的探针，并通过剖面和实验操作来量化它们的种群动态。该项目还将通过检查生物膜形成和细胞外多糖生产的程度来研究微生物微栖息地如何沿着压力和资源梯度变化，这似乎是细菌应激反应的核心组成部分。生物膜程度将通过环境电子显微镜和多糖的化学分析来评估。微生物过程动力学将通过一套完整土壤样品上的C和N转换过程来测量。这些指标包括净氮周转率和总氮周转率、各种14c底物（一系列单体、纤维素、木材）的最大消耗率、对这些底物的利用效率、利用它们的微生物的生长速度以及能够利用每种底物的种群的生物量。这种对资源和压力梯度、群落组成、生物膜和群落功能的整合研究将极大地增加我们对微生物群落生态学及其与生态系统过程的联系的理解。

Despite recent advances in environmental microbiology, our understanding of the factors that control microbial community composition and dynamics is still limited, and of the links between community composition and functioning not well understood. Developments in this direction will come from blending top-down (process) and bottom-up (molecular diversity) approaches, and tying into existing ecological theory. The intermediate disturbance & resource supply hypotheses from plant ecology should apply to microbial communities through the soil profile, as both resource availability and stress are high at the soil surface but decline with depth. Stress and disturbance may also partially explain why linking soil diversity to function has been difficult in studies that rarely sample below 10 centimeters. Soils are much deeper, with significant microbial populations at depth accounting for a substantial amount of whole-profile activity. The central hypothesis of this Microbial Observatory is that the gradients of resources and stress are the primary drivers of the size, diversity, dynamics, activity, and growth strategies (biofilm formation) of the microbial community throughout the soil profile. This project will be carried out at the Sedgwick Reserve, a site of active collaborative work on landscape processes, soil dynamics, plant community structure, and soil microbial processes in the California annual grassland/oak savanna ecosystem. Work will couple extensive sampling across the reserve to intensive manipulative studies at two core sites, where monitoring of soil climate, chemistry, and communities down to at least 4 meters depth and manipulative studies will be established. Soil moisture and temperature will be monitored to evaluate stress gradients, while substrate gradients will be assayed through a combination of physical soil organic matter fractionation and bioassay approaches. Microbial biomass will be measured, as will total and active numbers and biovolumes of bacteria and fungi. Bacterial community structure and composition will be assayed by pairing DGGE (denaturing gradient gel electrophoresis) and t-RFLP (terminally labeled restriction fragment length polymorphism). DGGE has the advantage that individual bands may be characterized and specific probes developed, while t-RFLPs provides cleaner analysis of complex DNA mixes. These fingerprinting approaches will be followed up by developing probes for specific bacteria and quantifying their population dynamics through the profile and under experimental manipulations. This project will also examine how the microbial microhabitat changes along the stress and resource gradients by examining the extent of biofilm formation and extracellular polysaccharide production, which appear to be central components of bacterial stress responses. Biofilm extent will be evaluated both by environmental electron microscopy and chemical analysis of polysaccharides. Microbial process dynamics will be measured by a suite of C and N turnover processes on intact soil samples. These include net and gross N turnover rates, maximum consumption rates for a variety of 14 C substrates (a range of monomers, cellulose, wood), the substrate use efficiency on those substrates, the growth rate of microbes using them, and the biomass of the populations capable of using each substrate. This integration of studies on resource and stress gradients, community composition, biofilms, and community functioning will greatly increase our understanding of microbial community ecology and its links to ecosystem processes.