We conducted a plume-scale study of the microbial ecology in the anaerobic portion of an aquifer contaminated by crude-oil compounds. The data provide insight into the patterns of ecological succession, microbial nutrient demands, and the relative importance of free-living versus attached microbial populations. The most probable number (MPN) method was used to characterize the spatial distribution of six physiologic types: aerobes, denitrifiers, iron-reducers, heterotrophic fermenters, sulfate-reducers, and methanogens. Both free-living and attached numbers were determined over a broad cross-section of the aquifer extending horizontally from the source of the plume at a nonaqueous oil body to 66 m downgradient, and vertically from above the water table to the base of the plume below the water table. Point samples from widely spaced locations were combined with three closely spaced vertical profiles to create a map of physiologic zones for a cross-section of the plume. Although some estimates suggest that less than 1% of the subsurface microbial population can be grown in laboratory cultures, the MPN results presented here provide a comprehensive qualitative picture of the microbial ecology at the plume scale. Areas in the plume that are evolving from iron-reducing to methanogenic conditions are clearly delineated and generally occupy 25-50% of the plume thickness. Lower microbial numbers below the water table compared to the unsaturated zone suggest that nutrient limitations may be important in limiting growth in the saturated zone. Finally, the data indicate that an average of 15% of the total population is suspended.