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Most cultured Japanese eels (Anguilla japonica) show male sex differentiation; however, natural gonadal sex differentiation has not been evaluated. In this study, this process was characterized in wild eels. Differentiated ovaries and testes were observed after the eels grew to 320 and 300 mm in total length, respectively. The youngest ovary and testis appeared at 3 and 4 years old, respectively; however, undifferentiated gonads were found up to 7 years, suggesting that sex differentiation was triggered by growth rather than aging. gsdf, amh, foxl2b and foxl3b were highly expressed in the testes, whereas figla, sox3, foxn5, zar1, and zp3 were highly expressed in the ovaries. The expression of cyp19a1a and foxl2a did not differ significantly between the testis and ovary. In the ovaries, the cyp19a1a and foxl2a levels were highest in the early stages, suggesting that their function is limited to early ovarian differentiation. The foxn5, zar1 and zp3 levels tended to increase in the later stages, suggesting that they function after the initiation of ovarian differentiation. In undifferentiated gonads, dimorphic gene expression was not observed, suggesting that the molecular sex differentiation phase is short and difficult to detect. These findings provide the first demonstration of the whole course of natural gonadal sex differentiation in eels at molecular and morphological levels.

Estuaries — one of the most vulnerable ecosystems globally — face anthropogenic threats, including biodiversity loss and the collapse of sustainable fisheries. Determining the factors contributing to the maintenance of estuarine biodiversity, especially that of fish, is vital for promoting estuarine conservation and sustainability. We used environmental DNA metabarcoding analysis to determine fish species composition in 22 estuaries around Japan and measured watershed-scale land-use factors (e.g., population size, urban area percentage, and forest area percentage). We sought to test the hypothesis that the richness of the most vulnerable estuarine fish species (i.e., registered by the Japanese Ministry of the Environment in the national species red-list) is determined by watershed-scale land-use factors. The richness of such species was greater where forest cover was highest; thus, forest cover contributes to their conservation. The proportion of agriculture cover was associated with low species richness of red-listed fishes (redundancy analysis, adjusted R2 = 43.9% of total variance, df = 5, F = 5.3843, p = 0.0001). The number of red-listed species increased from 3 to 12 along a watershed land-use gradient ranging from a high proportion of agriculture cover to a large proportion of forest cover. Furthermore, the results showed that throughout Japan all the examined watersheds that were covered by >74.8% forest had more than the average (6.7 species per site) richness of red-listed fish species. This result can be attributed to the already high average forest cover in Japan of 67.2%. Our results demonstrate how the land use of watersheds can affect the coastal sea environment and its biodiversity and suggest that proper forest management in conjunction with land-use management may be of prime importance for threatened fish species and coastal ecosystems in general.

Biodiversity is an important parameter for the evaluation of the extant environmental conditions. Here, we used environmental DNA (eDNA) metabarcoding to investigate fish biodiversity in five different estuaries in Japan. Water samples for eDNA were collected from river mouths and adjacent coastal areas of two estuaries with high degrees of development (the Tama and Miya Rivers) and three estuaries with relatively low degrees of development (the Aka, Takatsu, and Sendai Rivers). A total of 182 fish species across 67 families were detected. Among them, 11 species occurred in all the rivers studied. Rare fishes including endangered species were successfully detected in rich natural rivers. Biodiversity was the highest in the Sendai River and lowest in the Tama River, reflecting the degree of human development along each river. Even though nutrient concentration was low in both the Aka and Sendai Rivers, the latter exhibited greater diversity, including many tropical or subtropical species, owing to its more southern location. Species composition detected by eDNA varied among rivers, reflecting the distribution and migration of fishes. Our results are in accordance with the ecology of each fish species and environmental conditions of each river.

Colonization of new ecological niches has triggered large adaptive radiations. Although some lineages have made use of such opportunities, not all do so. The factors causing this variation among lineages are largely unknown. Here, we show that deficiency in docosahexaenoic acid (DHA), an essential w-3 fatty acid, can constrain freshwater colonization by marine fishes. Our genomic analyses revealed multiple independent duplications of the fatty acid desaturase gene Fads2 in stickleback lineages that subsequently colonized and radiated in freshwater habitats, but not in close relatives that failed to colonize. Transgenic manipulation of Fads2 in marine stickleback increased their ability to synthesize DHA and survive on DHA-deficient diets. Multiple freshwater ray-finned fishes also show a convergent increase in Fads2 copies, indicating its key role in freshwater colonization.