The tree-dwelling giant noctule Nyctalus lasiopterus, a partially carnivorous aerial-hawking bat, is one of the least known European bats, and more information is needed to evaluate its conservation status. Using radiotracking, we obtained the first data on spatial requirements and habitat preferences for the species in an area critically affected by deforestation. Two breeding populations separated by 60 km, one roosting in a city and the other in a nature reserve encircling a vast coastal marshland, showed marked differences in the size of their home ranges, but both used selectively the marshlands for foraging. Urban bats remained in the city for foraging during pregnancy in spring, but increased largely their home ranges towards the marshland during lactation in early summer. The nature reserve, with few roosting opportunities, was only a marginal roosting site for a population which switched between roosts located up to 90 km apart. Giant noctules had to travel long distances (up to 130 km recorded) to meet both foraging and roosting requirements within the fragmented landscape, exhibiting among the largest home ranges ever reported in Microchiroptera. To promote the protection of this threatened species, a network of artificial roosting options should be provided in good foraging habitats until long-term forest restoration is achieved.