Here, we describe a new microsporidium Percutemincola moriokae gen. nov., sp. nov., which was discovered in the intestinal and hypodermal cells of a wild strain of the nematode Oscheius tipulae that inhabits in the soil of Morioka, Iwate Prefecture, Japan. The spores of Pe. moriokae had an average size of 1.0 × 3.8 µm and 1.3 × 3.2 µm in the intestine and hypodermis, respectively, and electron microscopy revealed that they exhibited distinguishing features with morphological diversity in the hypodermis. Isolated spores were able to infect a reference strain of O. tipulae (CEW1) through horizontal transmission but not the nematode Caenorhabditis elegans. Upon infection, the spores were first observed in the hypodermis and then in the intestine the following day, suggesting a unique infectious route among nematode-infective microsporidia. Molecular phylogenetic analysis grouped this new species with the recently identified nematode-infective parasites Enteropsectra and Pancytospora forming a monophyletic sister clade to Orthosomella in clade IV, which also includes human pathogens such as Enterocytozoon and Vittaforma. We believe that this newly discovered species and its host could have application as a new model in microsporidia–nematode association studies.