Although several studies have described roost use by Rafinesque's big-eared bats (Corynorhinus rafinesquii), few studies have examined roost selection. We examined roost use and selection by Rafinesque's big-eared bat at the tree, stand, and landscape scales during the maternity season in pristine old-growth habitat in the Coastal Plain of South Carolina. We located 43 roosts (14 maternity, 29 solitary) through cavity searches and radio-telemetry. Maternity colonies and solitary individuals selected roosts based on similar characteristics. The best model explaining roost selection by all bats included tree and stand characteristics; landscape factors had little influence on roost use. Bats selected large diameter trees in areas with a high density of trees with cavities in the surrounding area. Most roosts (67.4%) were in water tupelo (Nyssa aquatica) in semi-permanently flooded and saturated areas. Half of maternity roost cavities had upper bole openings whereas only 25.8% of solitary roosts had upper bole openings. Bats that roosted with maternity groups stayed in roosts for significantly shorter periods of time (1.3 ± 0.1 days) and used significantly more roost trees (5.0 ± 0.6 roosts) than adult males (3.8 ± 1.10 days, 2.3 ± 0.4 roosts, respectively). Maternity colony use of cavities with upper bole openings and shorter residency times suggest that predator avoidance may have been an important factor governing roosting behavior of maternity colonies in this area. Our results suggest that retention of large diameter, hollow trees in wetland areas will benefit Rafinesque's big-eared bat individuals and maternity colonies in this area.