The medaka, Oryzias latipes, has an XX/XY sex determination mechanism. A Y-linked DM domain gene, DMY, has been isolated by positional cloning as a prime candidate for the sex-determining gene. Furthermore, the crucial role of DMY during male development was established by studying two wild-derived XY female mutants. In this study, to find new DMY and sex-determination related gene mutations, we conducted a broad survey of the genotypic sex (DMY-negative or DMY-positive) of wild fish. We examined 2274 wild-caught fish from 40 localities throughout Japan, and 730 fish from 69 wild stocks from Japan, Korea, China, and Taiwan. The phenotypic sex type agreed with the genotypic sex of most fish, while 26 DMY-positive (XY) females and 15 DMY-negative (XX) males were found from 13 and 8 localities, respectively. Sixteen XY sex-reversals from 11 localities were mated with XY males of inbred strains, and the genotypic and phenotypic sexes of the F(1) progeny were analyzed. All these XY sex-reversals produced XY females in the F(1) generation, and all F(1) XY females had the maternal Y chromosome. These results show that DMY is a common sex-determining gene in wild populations of O. latipes and that all XY sex-reversals investigated had a DMY or DMY-linked gene mutation.