Canines are definitive hosts of Neospora caninum (Apicomplexa). For horizontal transmission from canines to occur, viable oocysts of N. caninum must occur in the environment of susceptible intermediate hosts. Canids in Australia include wild dogs and Aboriginal community dogs. Wild dogs are those dogs that are not dependent on humans for survival and consist of the dingo, feral domestic dog and their hybrid genotypes. Aboriginal community dogs are dependent on humans, domesticated and owned by a family, but are free-roaming and have free access throughout the community. In this study the extent of N. caninum infection was determined in a total of 374 dogs (75 wild dogs and 299 Aboriginal community dogs) using a combination of microscopic, molecular and serological techniques. Oocysts of N. caninum were observed in the faeces of two juvenile Aboriginal community dogs (2/132; 1.5%). To estimate N. caninum prevalence, a new optimised cut-off of 18.5% inhibition for a commercial competitive ELISA was calculated using a two-graph receiver-operating characteristic (TG-ROC) analysis and IFAT as the gold standard resulting in equal sensitivity and specificity of 67.8%. Of the 263 dog sera tested the true prevalence of N. caninum antibodies was 27.0% (95% confidence limit: 10.3'44.1%). The association between the competitive ELISA results in dogs less than 12 month old and older dogs was significant (P = 0.042). To our knowledge this is the first large scale parasitological survey of the Aboriginal community dogs and wild dogs from Australia. The high prevalence of N. caninum infection in Aboriginal community dogs illustrates that horizontal transmission of N. caninum is occurring in Australia. These results demonstrated that N. caninum in dogs is widespread, including the semi-arid to arid regions of north-western New South Wales and the Northern Territory. The populationsof free-ranging dogs are likely to be important contributors to the sylvatic life cycle of N. caninum.