A sequence analysis of the mitochondrial cytochrome oxidase subunit II (COII) gene in Russian and Turkish maize-associated populations of Ostrinia nubilalis and a Slovenian population of O. nubilalis probably infesting maize revealed little diversity. This lack of diversity may have resulted from bottleneck event(s) when the maize-associated population of O. nubilalis expanded from small population(s) in association with the cultivation of maize in Europe ca. 500 years ago. In the genealogy of COII genes obtained in the present and previous studies, Eurasian samples were substantially differentiated from North American samples. Since the North American population of O. nubilalis came from Europe, our finding suggests that there is geographical differentiation in European maize-associated O. nubilalis, and that maize-associated populations of O. nubilalis expanded multiple times in Europe. Finally, a phylogenetic analysis of the COII gene did not support that O. nubilalis and O. furnacalis are the closest relatives within the O. furnacalis species group.