Despite the importance of Neospora caninum in veterinary medicine, knowledge of distribution of neosporosis in dog populations in some countries is still poor. The aims of the present study were to determine the occurrence of anti-N. caninum antibodies in one-hundred dogs living in cattle farms or dog breedings in central Italy and to evaluate the risk factors associated with seropositivity. The incidence of reproductive system disorders (e.g. infertility after first pregnancy) was also evaluated. Serum from breeding and farm dogs was tested to an indirect immunofluorescent antibody test (IFAT) to assess the occurrence of seropositivity. Management and individual data were collected and analysed both by linear and logistic multiple-regression models to find reliable predictors of seroprevalence and anti-N. caninum antibody level. The seropositivity for N. caninum was 32%. Dogs reared for breeding and presence of cattle on the farm were associated with seropositivity for N. caninum. Dogs living in the cattle farms showed a higher seropositivity for N. caninum (46%) compared with those living in dogs breeding (18%) (P < 0.05). The high presence of seropositive dogs in cattle farms of the study region demonstrates the potential risk of horizontal transmission of N. caninum between dogs and cattle, regardless the occurrence of reproductive system disorders or with infectious bovine tissues contact. Although the Neospora seropositivity in dog breedings may appear relatively low if compared with that found in dogs living with livestock, this infection, apparently underestimated, should be considered as a potential serious problem in canine medicine.