Reproductive interference due to interspecific hybridization can lead to character displacement among related species with overlapping ranges. However, no studies have examined which reproductive traits are most important in reducing reproductive interference. We conducted molecular analyses of two nuclear genes (28S and Wingless) and a mitochondrial gene (COI) from two closely related ground beetle species, Pterostichus thunbergi and P. habui (Coleoptera: Carabidae), with overlapping distributions. In addition, we examined four reproductive traits (body size, organ morphologies of intromittent and non-intromittent male genital organs, and female reproductive period) in sympatric and allopatric habitats. We compared male genital morphology using geometric morphometric analysis. The species determined by morphology were classified into separate groups based on the phylogenetic tree constructed by the nuclear gene (Wingless). However, according to the mitochondrial genes examined, P. thunbergi was not monophyletic, while at the sympatric sites these species formed a monophyletic clade. This incongruence suggests that interspecific hybridization and subsequent mitochondrial introgression from P. habui to P. thunbergi have occurred. Concerning genital morphology, both of the intromittent and non-intromittent organs of P. thunbergi differed more from P. habui at the sympatric sites than between allopatric sites, suggesting directional reproductive character displacement. Pterostichus thunbergi, which likely arrived in P. habui habitat in small numbers, would have experienced stronger selection pressures than P. habui.