High-throughput techniques for analyzing complex bacterial communities.

A more complete understanding of microbial diversity and the environmental processes they control will require much more than a biotic inventory. It will require a deeper understanding of the basic features of systems organization and inter-population interactions. Communities, not total biomass, control net process rates driving the biogeochemical cycles sustaining the biosphere. Although the general patterns of macroorganismal diversity are relatively well known, spatial and temporal patterns of microorganismal diversity are essentially unknown. Having tools capable of resolving these patterns is a prerequisite to developing an understanding of the relationship between community structure and function. This talk discusses conceptual and technical developments that now provide the framework for systematically resolving temporal and spatial patterns of microorganisms and relating those patterns to processes at local and system levels. Of particular emphasis will be ongoing studies using highly parallel analyses with DNA microarrays for intensive monitoring of microbial populations in environmental systems. Although microarray technology is reasonably well established for studies of model organisms in well-defined laboratory settings, the application of this technology to environmental systems of uncharacterized diversity imposes additional demands on implementation; in particular, the requirement for optimized discrimination between target and non-target nucleic acids in complex, and undefined, mixtures. To increase the resolving power (information content) of our DNA microarray format, we are investigating the use of thermal dissociation of hybrids immobilized on individual array elements to resolve target and non-target sequences that differ by a single nucleotide. These studies, combined with specialized algorithms for optimizing the readout of the microarray should serve for informed environmental application. Initial studies have validated the general approach for analyses of sediment systems.