Natterer's bat Myotis nattereri is one of the least known European bats, using roosts inaccessible to humans and shifting to alternative roosts every few days. The characteristics of day and night roosts used by maternity colonies during summer was assessed. Roosts were located by radio-tracking. Roost sites (which may contain more than one roost) were concentrated mainly in core areas of up to 2 km2, within which roost site density ranged from 7 to 15 per km2. Day roosts included tree cavities, apices of attics, mortises in attics and in barns, soffit boxes at eaves, tops of gable walls, crevices in stone walls and a modern cavity wall. Trees comprised 67% of roost sites. Two maternity colonies each used between 21 and 31 roosts distributed across 15 to 25 roost sites. During late summer, the temperature of roosts adjacent to a roof covering averaged 22.5 °C during the day and were significantly warmer than roosts in trees, which averaged 17.3 °C. Attic mortise roosts were intermediate at 20.0 °C. The maximum ambient roost temperature in which bats remained was 32 °C and the maximum body surface temperature recorded while roosting was 37.6 °C. Thus, bats seem to require access to a large number of roosts with a range of temperatures that they can use depending on their reproductive state and energy requirements. The study colonies also exhibited high inter-annual fidelity to roosts. Attic maternity roosts used during late gestation and early lactation were reused for the same function in successive years, as were heavily timbered barns during late lactation and post-lactation. This continuing dependence upon established roosts indicates that these types of maternity roost should be given high conservation priority. Field boundaries should also be managed to maintain mature trees with cavities for potential day roosts and hedges suitable as night perches.