We used a DNA barcoding approach to identify specimens of the Daphnia pulex complex occurring in New Zealand lakes, documenting the establishment of non-indigenous North American Daphnia 'pulex'. Morphological delineation of species in this complex is problematic due to a lack of good morphological traits to distinguish the species, as there is a relatively high degree of morphological stasis within the group through evolutionary time. Accordingly, genetic analyses were used to determine the specific identity and likely geographic origin of this species. Morphologically, individuals most closely resembled Daphnia pulicaria or Daphnia pulex sensu lato, which cannot be separated morphologically. Furthermore, each of these taxa comprises separate species in North America and Europe, despite carrying the same names. We identified individuals using a 658 bp nucleotide portion of the mitochondrial cytochrome c oxidase subunit 1 gene (COI) as North American Daphnia 'pulex', being distinct from European Daphnia pulex sensu stricto and D. pulicaria from Europe or North America. Cellulose allozyme electrophoresis was used to confirm that individuals were not hybrids with D. pulicaria. North American Daphnia 'pulex' in New Zealand were first recorded in New Zealand from South Island lakes that are popular for overseas recreational fishers, indicating a possible source of introduction for this species (e.g. on/in fishing gear). Our study provides an additional example of how genetic techniques can be used for the accurate identification of non-indigenous taxa, particularly when morphological species determination is not possible. The growth of global databases such as GenBank and Barcode of Life Datasystems (BOLD) will further enhance this identification capacity.