A putative hybrid zone between flightless carabid beetles, Carabus (Ohomopterus) insulicola and C. (O.) arrowianus nakamurai in the Ina Valley central Honshu, japan, was studied using experimental hybridization and morphological analysis. The principal morphological character analysed was a functional part of the male genitalia (the copulatory piece), which is also the key diagnostic character in taxonomy. The habitat of the carabid beetles is fragmented hv the Tenryu River, which runs through the Ina Valley: and its tributaries. Ohomopterus insulicola and O. a. nakamurai occur in the upstream and downstream areas of the river basin, respectively, and a putative hybrid swarm (O. insulicola ssp. pseudinsulicola) is found in the boundary area on the east side of the Tenryu River. Experimental hybridization between O, insulicola and O. a. nakamurai resulted in F-1 offspring that were morphologically similar to O. i. pseudinsulicola. Pre-zygotic reproductive isolation was incomplete, although the F-1 males had a dysfunction in sperm production and were almost sterile. Only a single F-1 pair produced F-2. Backcrosses of F-1 females of males of the parental species resulted in offspring that were similar to the parental species in genital morphology. Based on the discriminant function for the shape of the copulatory piece resulting from the experimental hybridization, we demonstrate that similar hybrid swarms exist on both sides of the Tenryu River, but in locations 25 km apart. This demonstrates parallelism in hybridization events with similar consequences. The hybrid swarms consisted of beetles with intermediate morphological characters and did not contain obvious parental forms. The establishment of such intermediate populations may have been facilitated by selection for fertile hybrids in segregated local sites in the absence of frequent immigration of parental species. This study suggests that a segmented river basin provides an opportunity for establishing novel evolutionary lineages resulting from hybridization. (C) 2000 The Linnean Society of London.