In human sera collected in three regions of Slovakia during an epidemic of respiratory infections, both genus-specific chlamydial and species-specific anti-Chlamydia pneumoniae antibodies, as detected by enzyme-linked immunosorbent assay and microimmunofluorescence test, respectively, were found. Based on seroconversion or significant rise of antibody titers and detection of antibodies of IgM class, an acute C. pneumoniae infection was indicated in 21 of 298 (7.0%) patients tested. The results obtained bring the first evidence on the role of C. pneumoniae as respiratory pathogen in Slovakia.