﻿AbstractCurrently, the genusArctiaSchrank, 1802 includes approximately 16 species in the Palaearctic region, depending on the taxonomic interpretation. Here, populations of theArctiavillica(Linnaeus, 1758) morphospecies complex were studied from Europe to the Middle East (Turkey, northern Iran) by molecular methods. Morphological treatment has traditionally revealed the presence of five nominal taxa:A.villica(Linnaeus, 1758),A.angelica(Boisduval, 1829),A.konewkaii(Freyer, 1831),A.marchandide Freina, 1983, andA.confluensRomanoff, 1884. The molecular approach tests whether they represent well-delimited species. Subsequently, this study corroborates the suitability of the mitochondrial cytochrome c oxidase subunit 1 (COI) marker sequence for species delimitation. In total, 55 barcodes of theArctiavillicacomplex were compared, and two molecular species delimitation algorithms were applied to reveal the potential Molecular Operational Taxonomic Units (MOTUs), namely the distance-based Barcode Index Number (BIN) System, and the hierarchical clustering algorithm based on a pairwise genetic distances approach using the Assemble Species by Automatic Partitioning (ASAP). The appliedASAPdistance-based species delimitation method for the analysed dataset revealed an interspecific threshold of 2.0–3.5%K2Pdistance as suitable for species identification purposes of the IberianA.angelicaand the SicilianA.konewkaiiand less than 2% for the three taxa of theA.villicaclade:A.villica,A.confluens, andA.marchandi. This study contributes to a better understanding of the taxonomy of the genusArctiaand challenges future revision of this genus in Turkey, the Caucasus, Transcaucasia as well as northern Iran using standard molecular markers.