Steinernema ashiuense sp. n. was collected by a Galleria baiting method from grassland along the riverbank of the Yura river, near the Ashiu Research Forest Station of Kyoto University. The new species is characterised by an infective juvenile body length of 768 (720-800) μm, lateral field with five equally developed ridges (i.e., six lines or incisures), head smooth lacking horn-like structures, excretory pore located at level of middle of pharynx, hyaline portion short (ca one-third of tail length). Males have 50-65 μm long, broad, slightly yellowish spicules; spicule length / spicule width = 4 (3.5-4.4), and two subventral and one subdorsal pair of genital papillae in the postcloacal region. The analysis of ITS-rDNA sequence placed S. ashiuense sp. n. in the 'feltiae-kraussei-oregonense' group in the clade containing S. robustispiculum and S. monticolum. It can be distinguished from these species by morphological characters of the infective juveniles and males.