Abstract: The vitellogenin (VTGa) and estrogen/androgen receptor (ERa/ARa) gene mRNA transcriptional levels in mosquitofish (Gambusia affinis) were detected to monitor the pollution status of environmental estrogen/androgen substances in Hanxi River, Dongguan City. The results were that compared with the control site of Liuxi River (LX), the VTGa mRNA transcription levels in liver of male mosquitofish from sampling sites of Songshan Lake (SM), Yangwu Village (YW), Hengli Town (HL), Upstream of Zhangcun sewage treatment plant（ZU） and Downstream of Zhangcun sewage treatment plant (ZC) were significantly increased (P0.05 and P0.001) up to 7.67, 169.43, 148.69, 152.01 and 98.12 fold, respectively; the ER mRNA transcript levels in anal fin of male mosquitofish were significantly increased (P0.05) only in HL compared with the control point of LX; the variation of transcript levels in the rest of the sites were not significant (P0.05). The ARa mRNA transcript levels in anal fin of female mosquitofish in sampling sites of SM, YW, HL, ZU and ZC from Hanxi River were 1.21, 0.82, 0.34, 0.41 and 0.89 fold of the control point of LX, respectively. There was significant resistance decreases (P0.05) in the HL and ZU point; and the rest of the variations were not significant (P0.05). The results indicated that mosquitofish in Hanxi River was significantly influenced by estrogen/androgen material interference, and the estrogen effect was more significant. As biomarkers, fish liver VTG gene mRNA transcription level relative to the anal fin ER/AR gene mRNA transcription level was more suitable for monitoring the aquatic environment of estrogen/androgen substances.