Half-smooth tongue sole (Cynoglossus semilaevis) is an important cultured marine fish and shows large sexual dimorphism in growth. In general, the females grow about four to five times faster than males. However, the percentage of males in cultured group is higher than that of females. Is the sex ratio deviation from expected value(1∶1) associated with natural sex reversal？ In the present study, the natural sex reversal phenomenon in halfsmooth tongue sole was studied by means of female specific marker sex identification technique. We determined the phenotypic sex of 71 four-year-old halfsmooth tongue sole by means of abdominal compression during reproduction season. The individuals not producing sperm or eggs were injected with (0.4-2.0) μg/kg LRHA₃ and fed in concrete tanks until sexual maturation. In total, there were 39 phenotypic males and 32 females, and the mean body weight of phenotypic males was (0.339?0.078) kg and (1.471?0.214) kg of females. However, the body weight of one phenotypic male was 0.585kg and significantly higher than other males. The female-specific marker analysis also showed that the phenotypic male tongue sole produced female-specific band, but not other 38 phenotypic males; 32 phenotypic females all produced female-specific fragment of about 205 bp. As a result, the phenotypic male half-smooth tongue sole producing female-specific fragment could be natural sex reversal male(neomale). Large-scale genetically sexual identification of phenotypic males in Laizhou Mingbo Co. Ltd showed the rate of natural sex reversal was 1.66%. It also proved that phenomenon of natural sex reversal in tongue sole was universal. Histological analysis of the sex-reversal male gonads showed that they contained functional testicular tissue with sprmatocytes. However, the amounts of spermatocytes in sex reversal individuals were fewer than that in control males. The phenomenon of natural sex reversal of female half-smooth tongue sole is helpful to explain the high male percentage in cultivating group and provides a new technique to produce all-female half-smooth tongue soles.