As both companion animals and opportunistic predators, dogs (Canis lupus spp.) have had a long and complex relationship with humans. In Australia, the dingo (C. l. dingo) was introduced 4,000 years ago and, other than humans, is now the continent's top mammalian predator. Domestic dogs (C. l. familiaris) were introduced by Europeans more recently and they interbreed with dingoes. This hybridization has caused growing concern about the roles that domestic dogs and dingoes play in shaping ecosystem processes. There is also considerable debate about whether anthropogenic environmental changes can alter the ecological roles of dingoes. We used scat analysis to test whether the dingo, as the longer-established predator, occupies a different dietary niche from that of free-roaming domestic dogs, irrespective of human influence. Our results demonstrate considerable dietary overlap between dingoes and domestic dogs in areas where humans provide supplementary food, providing evidence against our hypothesis. However, the consumption by dingoes of a greater diversity of prey, in association with historical differences in the interactions between dingoes and humans, suggests a partial separation of their dietary niche from that of domestic dogs. We conclude that anthropogenic changes in resource availability could prevent dingoes from fulfilling their trophic regulatory or pre-European roles. Effective management of human-provided food is therefore required urgently to minimize the potential for subsidized populations of dingoes and domestic dogs to negatively affect co-occurring prey.