Bridging species boundaries: eDNA and genetic analysis reveal hybrid settlement at the very end of American eel distribution

Documenting species distributions and hybridization patterns is paramount for elucidating biogeography and understanding speciation processes. Here we combined genetic specimen analysis and environmental DNA (eDNA) to investigate the presence of American eel (Anguilla rotrata) and American x European eel (Anguilla anguilla) hybrids in Greenland freshwater. We further tested the use of eDNA to document hybridization by using European eel mtDNA as a proxy for hybrid occurrence. Overall, eDNA analysis detected mainly American eel but also European eel mtDNA. This finding was validated by DNA sequencing, which identified 3 out of 26 captured eels (14.3%) carrying European eel mtDNA. Five eels (19.2%), including all three with European mtDNA, were heterozygous for species-specific nuclear gene variants, supporting a hybrid ancestry. Further, eDNA successfully identified eels in lakes where they were caught by fyke net fishing, extending their confirmed northern range by 40 km, and indicated eel presence >200km further north. The study provides an empirical demonstration of the use of eDNA to document hybrid occurrence and extends the reported northern distribution of eels in Greenland significantly beyond previous observations. Further, the existence of hybrid eels in Greenland may be key for understanding the complex mechanisms of hybridization between American and European eels.