While the impacts of black (Rattus rattus) and brown (Rattus norvegicus) rats on human society are well documented—including the spread of disease, broad-scale environmental destruction, and billions spent annually on animal control—little is known about their ecology and behavior in urban areas due to the challenges of studying animals in city environments. We use isotopic and ZooMS analysis of archaeological (1550s–1900 CE) rat remains from eastern North America to provide a large-scale framework for species arrival, interspecific competition, and dietary ecology. Brown rats arrived earlier than expected and rapidly outcompeted black rats in coastal urban areas. This replacement happened despite evidence that the two species occupy different trophic positions. Findings include the earliest molecularly confirmed brown rat in the Americas and show a deep ecological structure to how rats exploit human-structured areas, with implications for understanding urban zoonosis, rat management, and ecosystem planning as well as broader themes of rat dispersal, phylogeny, evolutionary ecology, and climate impacts.