Phylogenetic relationships within the European Monochamus (Coleoptera: Cerambycidae) remain understudied despite their increasing importance in the Pine Wood Nematode spread in Europe. To clarify the delimitation and the evolutionary history of the two main European Monochamus species, Monochamus galloprovincialis and Monochamus sutor, as well as their sub-species, a comparative study using morphological, molecular, and biogeographical criterions was conducted. Four morphological characters, including a newly-described morphological character on the male genitalia, separated the two species. Additionally, molecular data revealed twelve and two single nucleotide polymorphisms in cytochrome oxidase c subunit I and 28S, respectively, supporting species segregation. By contrast, incongruence between morphological and genetic results did not allow discriminating the sub-species of M. galloprovincialis and M. sutor, even though mitochondrial DNA revealed intraspecific differentiation, mostly consenting to a multiple refugia origin. Within-species variability was explained to a large extent by biogeography (i.e. altitude, climate). These different ecological adaptations within beetle species, together with potential climate change impact, increase the risk of spreading the nematode across Europe to novel conifer hosts and challenge the European biosecurity.