The reintroduction of beaver (Castor canadensis) into arid and semi-arid rivers is receiving increasing management and conservation attention in recent years, yet very little is known about native versus non-native fish occupancy in beaver pond habitats. Streams of the American Southwest support a highly endemic, highly endangered native fish fauna and abundant non-native fishes, and here we investigated the hypothesis that beaver ponds in this region may lead to fish assemblages dominated by non-native species that favour slower-water habitat. We sampled fish assemblages within beaver ponds and within unimpounded lotic stream reaches in the mainstem and in tributaries of the free-flowing upper Verde River, Arizona, USA. Non-native fishes consistently outnumbered native species, and this dominance was greater in pond than in lotic assemblages. Few native species were recorded within ponds. Multivariate analysis indicated that fish assemblages in beaver ponds were distinct from those in lotic reaches, in both mainstem and tributary locations. Individual species driving this distinction included abundant nonnative green sunfish (Lepomis cyanellus) and western mosquitofish (Gambusia affinis) in pond sites, and native desert sucker (Catostomus clarkii) in lotic sites. Overall, this study provides the first evidence that, relative to unimpounded lotic habitat, beaver ponds in arid and semi-arid rivers support abundant non-native fishes; these ponds could thus serve as important non-native source areas and negatively impact co-occurring native fish populations.