Between 2007 and 2010, a total of 268 Croatian patients with lymphadenopathy were tested for IgM/IgG antibodies to Bartonella (B.) henselae and B. quintana. Samples from 44.4% patients showed positive IgG antibodies: 35.8% to B. henselae, 6.7% to B. quintana and 1.9% to both Bartonella species. There was no difference in seropositivity between males and females (47.4% vs. 41.5%). Seroprevalence was high in all age groups (40.4-60.9%). Patients from urban and rural areas showed a similar seroprevalence rate (44.1% vs. 44.8%). Positive IgM antibodies were found in 28.3% patients varying from 17.5% and 37.5% among age groups. Most cases were reported from August to March.