Hybrids of Rhinogobius biwaensis and Rhinogobius sp. BF were identified in the ponds of Sento Imperial Palace, Kyoto City, Japan, from microsatellite and mitochondrial DNA markers. Mitochondrial DNA analysis indicated that five of seven Rhinogobius specimens had R. biwaensis haplotypes, the remaining specimens having those of Rhinogobius sp. BF. In DAPC and STRUCTURE analyses based on microsatelli