As significant nocturnal insectivores, bats are an integral part of many ecosystems. Determining bat species composition in an area is a critical first step in managing for this important resource. Little information exists concerning the bat species composition of Maryland's Coastal Plain, which is located on the northern periphery of the geographic ranges of five bat species occurring in the southeastern United States. We conducted mist net surveys for bats at Assateague Island National Seashore, a barrier island on Maryland's coast, in summer 2005 and summer and autumn 2006. In 2005 we captured 133 bats representing three species, including two big brown bats (Eptesicus fuscus), 129 eastern red bats (Lasiurus borealis) and two purported Seminole bats (L. seminolus). In 2006 we captured 60 eastern red bats in summer and autumn combined. We used Anabat II bat detectors to conduct long-term acoustic monitoring on the island year round and documented three additional bat species, including silver-haired bats (Lasionycteris noctivagans), hoary bats (L. cinereus) and eastern pipistrelles (Pipistrellus subflavus). We documented silver-haired bats during spring and autumn, probably as they were migrating through the area. We used Anabat II bat detectors to conduct short-term monitoring of bat activity at five habitat types during summer 2005 and 2006 and found that total bat activity and eastern red bat activity were similar among forested areas, freshwater pools and bayside marshes. In shrublands, total bat activity and eastern red bat activity was higher than at beach areas, lower than in forested areas and similar at freshwater pools and bayside marshes. The loblolly pine (Pinus taeda) – dominated forests at ASIS provide roosting and foraging habitat mostly for eastern red bats, but also for other migratory bat species.