Fig 6

(A-C) Treatment with estradiol (E2) by water-borne exposure. The mutant fish (30/group) were treated from 40 to 60 dpf with E2 (10 nM). E2 treatment resulted in all-female phenotype in contrast to untreated group which showed a male-biased sex ratio (A). Electrophoresis of the sera (1 μl/lane) from treated mutant females showed mass production of vitellogenin protein (arrow); in contrast, there was no vitellogenin in the sera of untreated mutant fish (-/-). The control fish (+/-) without E2 treatment showed a weak band at the location of vitellogenin. A similar band could also be seen in the liver extract (10 μg/lane) in response to E2 treatment. Each lane represents one representative individual, and the serum and liver samples were obtained from the same individual for each genotype and treatment combination (B). Histology of the ovary from E2-treated fish showed arrest of follicles at PG stage with a few follicles (circled) showing rudiment cortical alveoli in some individuals (C). (D) Treatment with E2 by oral administration (feeding). The mutant fish were fed with E2-containing diet (20 μg/g) for 17 days (45–62 dpf) followed by examination for secondary sex characteristics (genital papilla in females, arrow; breeding tubercles in males, arrowhead) and gonadal histology. The treated group showed higher female ratio (44% females, 8/18) compared to the control group (21% females, 3/14). Females from both groups showed normal genital papilla with no breeding tubercles and arrested PG follicles in the ovary. The number in each histological photo indicates the number of fish examined with the same phenotype. PG, primary growth.