AbstractTaxonomic ambiguities of the Asplenium scolopendrium complex arise from multiple synonyms and reclassifications, subtle phenotypic variations, and a lack of information on phylogenetic relationships. This study thus aimed to resolve this taxonomic uncertainty and provide insight into identifying evolutionarily significant units (ESUs). We first utilized genomic approaches to resolve the phylogenetic position of the East Asian taxon, A. komarovii, which is currently treated as a separate species. Phylogenetic trees based on whole plastomes suggested that A. komarovii is a variant of A. scolopendrium most closely related to A. scolopendrium var. americanum, with A. scolopendrium var. scolopendrium as the sister to these in the clade. This three‐lineage relationship was also validated with the nuclear marker gapCp and newly developed infraspecific plastid markers. Asplenium komarovii should therefore be subsumed into A. scolopendrium, rather than remaining a distinct species. In addition, our phylogenetic analyses further revealed that A. scolopendrium var. americanum consisted of subclades with potential to be treated as distinct ESUs. Our results also grouped a newly discovered population from New Mexico (U.S.A.) as a member of A. scolopendrium var. americanum and identified a genetically admixed population in New York (U.S.A.) containing putative hybrids. Well‐defined taxonomy and ESUs can greatly improve the implementation of tailored conservation actions by adequately reflecting the underlying evolutionary potentials of the A. scolopendrium complex.