This research was supported by the Deep Microbiology Subprogram of the Subsurface Science Program, Office of Health and Environmental Research, U.S. Department of Energy (DOE). The continued support of Dr. F. J. Wobber is greatly appreciated. Pacific Northwest National Laboratory is operated for the DOE by Battelle Memorial Institute under contract DE-AC06-76RLO 1830To investigate the distribution of microbial biomass and activities to gain insights into the physical controls on microbial activity and potential long-term survival in the subsurface, 24 shale and sandstone cores were collected from a site in northwestern New Mexico. Bacterial biomass in the core samples ranged from below detection to 31.9 pmol total phospholipid fatty acid (PLFA) g-1 of rock with no apparent relationship between lithology and PLFA abundance. No metabolic activities, as determined by anaerobic mineralization of [14C]acetate and [14C]glucose and35SO42 reduction, were detected in core samples with pore throats <0.2 µmn in diameter, smaller than the size of known bacteria. However, enrichments revealed the presence of sulfate-reducing bacteria, and35SO42 reduction was detected upon extended (14 days) incubation in some small-pore-throat samples. In contrast, relatively rapid rates of metabolic activity were more common in core samples containing a significant fraction of pore throats >0.2 fan in diameter. These results suggest that subsurface bacteria require interconnected pore throats greater than 0.2 μm diameter for sustained activity but that viable bacteria can be maintained and stimulated in poorly permeable rocks, such as shales, with restrictive pore throat diameters. In addition, the detrital organic matter in the small-pore-diameter shales is not subject to direct microbial attack. Rather, bacteria in adjacent sandstones with a more open pore structure are probably sustained by endogenous nutrients that are slowly released from the shale. These results have implications for the long-term maintenance of anoxia and the impact of anaerobic biogeochemical processes on groundwater chemistry. © 1997 Taylor & Francis.