A serological survey was conducted to determine the occurrence of Leptospira spp. and Bartonella henselae in stray cats circulating in Palermo (Sicily, Italy). Samples from 721 cats were tested for Leptospira spp. antibodies by serological Microagglutination Test (MAT) and for B. henselae antibodies by Indirect Immunofluorescence Antibody assay (IFA). We found only one sample (0.1%) seropositive for antibodies to Leptospira spp. with a title of 1: 100 for Leptospira australis. Concerning B. henselae, 513 samples analyzed by IFA were positive (71%). This is the first screening in Sicily for these two pathogens and the results here obtained will be useful to take preventive measures to reduce the prevalence of such diseases in pets