Difenoconazole (DFZ) is a triazole fungicide which has been detected in the aquatic environment,including estuaries and embayments.However,few studies addressing the reproductive toxicity and transgenerational effects of DFZ on marine fishes are available.The present study was conducted to investigate the effects of DFZ on male marine medaka (Oryzias melastigma).After exposure of the embryo to 1,10,100 and 1000 ng/L DFZ for 180 days,the gonadosomatic index was significantly decreased in the 1000 ng/L treatment.The number of sperm was reduced while the abundances of spermatocytes and spermatogonia in the testes were increased in all the treatments.The mRNA levels of salmontype gnrh (sgnrh),the luteinizing hormone (lhβ) and the follicle-stimulating hormone (fshβ)genes in the brain all exhibited a significant down-regulation,the expression of androgen receptors (arα and arβ) was decreased and that of estrogen receptor β and cytochrome P450 aromatase (cyp19B) was increased in the testes.The expression levels of cyp19A and cyp19B were increased in the liver.The decrease of ars mRNA levels might be one of the reasons causing the reduction of sperm.The down-regulation of sgnrh,lhβ and fshβ mRNA levels suggested that DFZ might impact the spermatogenesis via the brain-pituitary-gonad pathway.The decrease of the fertilization success,the hatch ability and the swim-up success in the F1 generation indicated that DFZ pollution at environmental levels might cause a decrease of wild fish populations.