We examined 444 dogs by visiting farms and peripheral districts on the Gargano promontory (Province of Foggia, Apulia, South Italy). Clinical examination and immunofluorescence antibody test for leishmaniasis were performed. Bone marrow and lymph node samples obtained from 25 dogs with positive serological test were cultured on Tobie-Evans medium. The results obtained show both a higher seropositivity rate for canine leishmaniasis (14.4%) and a higher percentage of asymptomatic dogs with positive serological test (53.1%) compared to previous research in the same area. Seven strains isolated from infected dogs belonged to Leishmania infantum species zymodeme Montpellier 1. None of 82 sera of humans living in close contact with infected dogs were positive for leishmaniasis.