Host-seeking adult Dermacentor reticulatus ticks were examined for the prevalence of Francisella tularensis in an active natural focus of tularemia along the lower reaches of the Dyje (Thaya) river in South Moravia (Czech Republic) and adjacent Lower Austria, in four localities of the flood plain forest-meadow ecosystem during the spring of 1996. The ticks were pooled (10 male or female ticks per group) and inoculated subcutaneously in 4-week-old SPF mice. Dead mice were sectioned, the spleens were homogenized in PBS and passaged in mice, and impression smears from the spleens, liver and heart blood were stained by Giemsa. Twenty-five isolates of F. tularenis were recovered from 1098 pooled D. reticulatus: the minimum infection rate (MIR) is 2.3%. MIRs for 629 female and 469 male D. reticulatus were 2.4% and 2.1%, respectively. The prevalence varied according to locality, but did not significantly differ between the Moravian (2.2%) and Austrian (2.8%) sites. The monitoring of D. reticulatus for F. tularensis might be a valuable contribution to the surveillance of tularemia in Europe.