Fragipans are impermeable layers in soil that affect more than 950,000 ha of land in Kentucky and create seasonally perched water tables that may promote denitrifier growth and activity. We used denitrification enzyme activity assays and Most Probable Number (MPN) estimates of soil and water samples to investigate whether a fragipan soil in south-central Kentucky developed bacteria populations with increased denitrification capacity. Preliminary studies showed that up to 5.0 × 104 denitrifiers mL-1 were present in perched water above the fragipans. Nitrous oxide concentrations in peizometers were as high as 8.3 ppm and suggested that the denitrifiers were active throughout the year. Cover crops suppressed the denitrifier populations in perched water almost tenfold, and decreased N2O concentrations by 8%. Compared with unamended soil, denitrifiers were more numerous at all depths when manure was added. Subsequent experiments showed that there were up to 8.6 × 103 denitrifiers g-1 soil immediately above the fragipan and fewer than 4.6 × 102 within it. An analysis of denitrifier populations by depth indicated that their numbers increased in soil immediately above the fragipan. Denitrifier growth was greater than that of other heterotrophic bacteria. The ratio of MPN denitrifiers to total culturable heterotrophic bacteria plated at each depth increased 64% immediately above the fragipan. We conclude that the fragipan in this soil selected for denitrifiers, that the denitrifiers were affected by the management techniques applied at the soil surface, such as cover crop use and manure application, and that this environment has the potential to reduce NO-3 leaching beyond the plant rooting zone in fragipan soils.