We collected 138 Kumamoto like oyster specimens in and around the Nakatsu tidal flat to monitor species diversity. The mitochondrial large subunit of ribosomal DNA (LSrDNA) and the nuclear ribosomal internally transcribed spacer-1 (ITS-1) region were amplified by polymerase chain reaction and restriction fragment length polymorphism analysis, leading to the discovery of the Kumamoto oyster Crassostrea sikamea in the Nakatsu tidal flat. A more accurate species identification using the nucleotide sequences for cytochrome c oxidase subunit I and ITS-1 confirmed the presence of the oysters. These results provide the first evidence of C. sikamea in the Seto Inland Sea of Japan.