Relative contributions of aquaculture-origin and naturally-reproduced grass carp (Ctenopharyngodon idella) in the Laurentian Great Lakes have been unknown. We assessed occurrence and distribution of aquaculture-origin and wild grass carp in the Great Lakes using ploidy and otolith stable oxygen isotope (δ¹⁸O) data. We inferred natal river and dispersal from natal location for wild grass carp using otolith microchemistry and estimated ages of wild and aquaculture-origin fish to infer years in which natural reproduction and introductions occurred. Otolith δ¹⁸O indicated that the Great Lakes contain a mixture of wild grass carp and both diploid and triploid, aquaculture-origin grass carp. Eighty-eight percent of wild fish (n = 49 of 56) were caught in the Lake Erie basin. Otolith microchemistry indicated that most wild grass carp likely originated in the Sandusky or Maumee rivers where spawning has previously been confirmed, but results suggested recruitment from at least one other Great Lakes tributary may have occurred. Three fish showed evidence of movement between their inferred natal river in western Lake Erie and capture locations in other lakes in the Great Lakes basin. Age estimates indicated that multiple year classes of wild grass carp are present in the Lake Erie basin, recruitment to adulthood has occurred, and introductions of aquaculture-origin fish have happened over multiple years. Knowledge of sources contributing to grass carp in the Great Lakes basin will be useful for informing efforts to prevent further introductions and spread and to develop strategies to contain and control natural recruitment.