Application of DNA barcoding and morphometric analysis in differentiating cystacanths of Polymorphus species (Acanthocephala: Polymorphidae) from central Alberta, Canada.

Acanthocephalans are multi-host endoparasites, many of which use freshwater amphipods as intermediate hosts for their larval stages (e.g., cystacanths) while adults live in the intestines of vertebrates, including waterfowl. In central Alberta, Canada, several co-occurring species of the acanthocephalan genus Polymorphus use the amphipod Gammarus lacustris Sars, 1863 as an intermediate host. We applied DNA barcoding and morphometric analysis to differentiate cystacanth larvae from G. lacustris sampled from 17 Albertan water bodies. We slide-mounted specimens and measured morphological traits relating to proboscis hooks. We sequenced the standard DNA barcoding region of the mitochondrial cytochrome c oxidase subunit I gene (COI). Morphometric analysis suggested that the acanthocephalans we collected belonged to four morphologically different groups that keyed to Polymorphus contortus (Bremser, 1821) Travassos, 1926; P. marilis Van Cleave, 1939; P. paradoxus Connel et Corner, 1957; and P. strumosoides (Lundström, 1942) Amin, 2013. Our Bayesian tree based on COI sequences generally corroborated the morphological results and supported that the specimens assigned to P. cf. contortus and P. cf. strumosoides belong to two distinct species. In contrast, the Bayesian tree showed that specimens of P. cf. marilis were nested as a cluster within the P. cf. paradoxus clade. Similarly, small pairwise genetic distance (< 2%) between specimens identified as P. cf. contortus and P. cf. strumosoides suggests that they are conspecific. Future studies should use morphology and sequence data from adult acanthocephalans to assess the taxonomic identity of the cystacanth-based Polymorphus taxa. Our study is the first to provide genetic information for the four Polymorphus taxa and emphasizes the importance of applying multiple approaches to differentiate parasite species.