Cavity-roosting species of bats generally require patches of forest containing older timber, but such habitats have been reduced in the southeastern United States. Rafinesque's big-eared bat, Corynorhinus rafinesquii, is a rare species for which data on natural roosts in the Gulf Coastal Plain (where caves are largely absent) are scant. We used radiotelemetry to locate tree roosts of C. rafinesquii in southeastern Mississippi from August 2001 until August 2004. We captured and radiotagged 25 bats that led us to 14 trees. Corynorhinus rafinesquii used hollow Nyssa spp. and Magnolia grandiflora. Most of these trees were still alive, relatively large (mean DBH = 79.4 cm, mean height = 18.5 m), possessed cavities with openings not located at their bases and were located adjacent to streams or other bodies of water. When bats shifted roosts among different trees, these sites were usually located nearby (mean distance ≅ 360 m). Tree roosts were apparently rare within the study area, and C. rafinesquii showed roost fidelity to particular areas of forest. Six tree roosts were used by multiple individuals and several trees were reused within tracking sessions and among years.