BIOCOMPLEXITY (IDEA): In Situ Measurement of Marine Microbes to Investigate Mechanisms of Community Structure Regulation

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This project is supported by the program Biocomplexity in the Environment, subprogram Instrumentation Development for Environmental Activities (BE-IDEA). The objective is to develop instrumentation and software for automated in situ monitoring of marine microbial communities. Microbes account for almost all the primary productivity and a majority of the biomass in the ocean, and the structure of the microbial community affects higher trophic levels, including species consumed by humans. A fundamental understanding of the complex interaction between physical and biological factors that regulate community structure requires more detailed and sustained observations than are possible with current equipment. This project will develop and deploy in situ instrumentation to monitor individual cells of microscopic plankton. The planned approach will combine in situ flow cytometry and imaging-in-flow techniques to measure the abundance and properties of cells (and other particles) ranging from the smallest phytoplankton (less than 1 um) to large chain diatoms (100 um). An automated image analysis system will be used to determine taxonomic affiliation of each cell for the larger size classes. Smaller cells (which often lack distinctive morphological features) will be evaluated by pulse-shape analysis of individual cell light scattering and fluorescence. For more detailed investigation of particular groups or species, a module capable of applying group-specific rRNA probes (via Peptide Nucleic Acid in situ hybridization) will be developed. The probed samples will be analyzed in real time by the in situ flow cytometer. The same module will also be used to apply fluorescent probes for other properties such as DNA content and cell viability. Software to analyze and integrate the flow cytometric and image data in near-real time will also be developed. The instrument will be based on an in situ flow cytometer developed at WHOI (plus hybridization technology currently under development) and integrated with a commercially available imaging-in-flow system. When completed, the instrument package will be field tested near the Marthas Vineyard Coastal Observatory (MVCO).In combination with physical oceanographic, meteorological, and bulk fluorescence data, individual particle data will facilitate an understanding of processes affecting coastal ecosystems, including the mechanisms regulating phytoplankton blooms, species succession, and trophic interactions. The planned work will form the foundation for an unprecedented long term high resolution time series of natural microbial populations, whose members vary in size by many orders of magnitude. These kinds of observations are critical for evaluating and developing models of long-term ecological change.