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Long-term degeneration of the double mutant ovaries. a The ovaries in the double mutant (nobox−/−;dmrt1−/−) showed different degrees of degeneration at 120 dpf with loss of oocytes and decreased ovarian size (n = 5 independent fish). In addition, empty cavities or vacuoles left by degenerated oocytes were often observed (asterisks). The degenerated ovaries were gradually replaced by testicular tissues with meiotic cells (me); however, spermatogenesis could not proceed further due to the lack of dmrt1. The ovarian degeneration process was categorized into four stages based on gonadal size and morphological features. In stage I, the ovary was significantly smaller than the control (approximately half the size), containing both PG and PV follicles. Stage II was characterized by a dramatically reduced ovarian size, while still housing PG and PV follicles. In stage III, the ovary contained PG follicles only. In stage IV, the ovary was devoid of all follicles, featuring empty cavities or vacuoles left by degenerated oocytes. Additionally, testicular tissues began to emerge with meiotic germ cells. b Ovarian sizes of the control and double mutant ovaries undergoing degeneration. The area size of the largest section of each ovary was measured with ImageJ and the data are expressed as the ratios relative to the control. c Number of vacuoles resulting from oocyte loss. The vacuoles were counted on the largest section and classified based on their sizes compared to those of PG and PV follicles. d Composition of gonadal tissues in the control and double mutant ovaries. The areas of different gonadal tissues were measured using ImageJ.
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