From autumn 2004 to spring 2005, 70 fallow deer (Dama dama), 27 female and 43 male, living in a natural preserve of central Italy were examined by indirect immunofluorescence assay (IFA) to detect specific antibodies to Anaplasma phagocytophilum. Thirty-one (44.28%) sera scored positive: in particular 10 fallow deer (8 male and 2 female) scored positive at 40 antibody titer, 21 deer (8 male and 13 female) at > or = 80 titer. EDTA anticoagulated blood samples collected from 29 of the 70 deer examined were tested by a nested-PCR assay to disclose a 546 bp fragment, specific of A. phagocytophilum 16S rRNA gene. Twenty (72.41%) blood samples (8 male and 13 female deer) resulted positive. Fifteen PCR-positive deer also resulted positive to IFA, whereas the remaining six did not show specific antibodies. Three serologically positive animals gave negative results at the nested PCR. Five deer scored negative both to serological tests and PCR.