This paper presents data on the occurrence and breeding of Eurasian eagle owl (Bubo bubo) in south Moravia, Czech Republic (Břeclav and Znojmo districts), in the floodplains of the Morava and Dyje Rivers, since 1994, and the first cases of confirmed breeding from 2009 to 2011. In two cases, eagle owls bred successfully in large, wooden artificial nestboxes originally installed for saker falcon (Falco cherrug).