The aim of this study was to evaluate the prevalence (seroprevalence) of antibodies against Borrelia burgdorferi and Ehrlichia phagocytophila among patients resident in Lazio, a region of central Italy. Of a sample of 1,050 patients, which presented clinical manifestations related to Lyme disease, 34 (3.2%) were Borrelia-seropositive (Lyme index value >/= 1.2). The sera of 25 out of the 34 patients that were Borrelia-positive were also analysed for the presence of antibodies against E. phagocytophila and 3 (12%) were found Ehrlichia-positive (titres >1:64). No Ehrlichia positive samples were found among sera of 250 Borrelia-negative patients. Since both B. burgdorferi s.l. and Ehrlichia species share the same tick vector (Ixodes ricinus), our results indicate that concurrent transmission of these microbial pathogens might have been occurred among the patients included in this study.