The genus Mesobuthus belonging to family Buthidae is most diverse in Central Asia and Iran. The scorpion M.eupeus is one of the most naturally widespread members of the family Buthidae, being present in eastern Turkey, Caucasia, southern Russia, Middle East, Central Asia, southern Mongolia and northern China. Additionally, considerable morphological variation has been attributed to populations of M.eupeus within this extensive geographic distribution and 21 recognized valid subspecies have been described. A recent comprehensive update or revision of M.eupeus is not available. In this study, the first molecular phylogeny assessment of M.eupeus in Iran based on sequence data of a ~700bp fragment of Cytochrome C oxidase, subunit I is presented. Fifteen populations collected from different localities within Iran were included in the study. Phylogenetic relationships were inferred using neighbour-joining (p-distance) and maximum likelihood under GTR+Γ substitution model. The results support monophyly for M.eupeus, but they do show a clear deep split between northern and southern clades within the species. The northern clade included M.e.eupeus, M.e philippovitschi, M.e.afghanus, and M.e.thersites, while the southern clade comprised M.e.phillipsi and M.e.kirmanensis. In the topology observed, the \"southern clade\" has a basal position relative to the \"northern clade\". Accordingly, some scenarios for the evolution and phylogeographic structure of this species based on the paleogeography and geological history of Iranian plateau were proposed and tested. In addition to the intraspecific divergence, our topology showed a deeper phylogenetic split within the genus Mesobuthus. Here, a \"western clade\" consisting of M. gibbosus and M. cyprius is separated from an \"eastern clade\" containing M.eupeus and M.caucasicus. Finally, the molecular clock hypothesis using the likelihood ratio test was applied in order to test the constancy of the evolutionary rates across the tree.

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