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The aim of the present study was to assess the responses of the gonopodium morphology and the gonadal and liver histology of adult male Cnesterodon decemmaculatus to sublethal long-term exposure concentrations of 17α-ethinylestradiol (EE2). Two experiments were conducted exposing the fish to waterborne concentrations of EE2 ranging from 20 ng/L to 200 ng/L for 8 wk, 12 wk, and 16 wk. Intersex gonads were observed after 8 wk and 16 wk in fish exposed to 200 ng EE2/L and 100 ng EE2/L, respectively. Oocytes' development from testis germ cells and replacement of the efferent duct periodic acid-Schiff-positive secretion surrounding spermatozeugmata by parenchymal tissue and duct structure alterations were the major observed changes in the gonads. In contrast, no response was observed in the gonopodium morphology. Liver histology was also altered, showing increasing steatosis, single-cell necrosis to generalized necrosis, and disruption of acinar organization from 100 ng EE2/L to 200 ng EE2/L. In summary, the present results showed that although EE2 was not able to alter the morphology of a developed gonopodium, it was capable of inducing development of testicular oocytes in adult male C. decemmaculatus at environmentally relevant concentrations. Thus, externally normal but intersex C. decemmaculatus males would be expected in the wastewater-receiving streams that the species inhabits. According to the literature, the present study would be the first indicating estrogen-induced intersex in adult male poeciliid. Environ Toxicol Chem 2017;36:1738-1745. © 2016 SETAC.

1. Natural grassland ecosystems are increasingly threatened by excessive loadings of nutrients and by the presence of species bred for high productivity. By manipulating grazing regimes and nutrient availability, agricultural practices facilitate the establishment and spread of certain forage plant species outside managed landscapes, challenging local biodiversity. The ecological success of some species in the invaded range sometimes seems to be associated with the symbiosis with foliar fungal endophytes. Symbiotic fungi may increase the competitiveness of host species, but also the resistance to herbivory through the production of toxic secondary compounds such as alkaloids. While progress has been made in understanding how soil nutrients modulate other benefits offered by fungal endophytes to plants (e.g. stress tolerance, competitive ability, etc), the consequences for a higher trophic level (i.e. herbivores) and the potential feedbacks on plant invasion have not been explored yet. 2. We explored the relative and interactive importance of soil nitrogen (N) and phosphorus (P) in modulating the interaction of the invasive grass tall fescueassociated with fungal endophytes-and native herbivores in a natural grassland. We hypothesized that N and P nutrients modulate differentially leaf quality traits, namely nutritional value and fungal alkaloid contents, determining the level of damage by native insect herbivores on the exotic tall fescue. 3. We found that only the addition of P significantly increased native caterpillar density in the field, which corresponded to a concomitant increase in leaf damage. In contrast to expectations, the concentration of the alkaloid ergovaline in leaves was not strongly related to N. It was the level of soil P which dictated the concentration of the element (P) in the leaves and reduced the level of defence against herbivores in this endophyte-symbiotic species. Then, herbivore performance increased, and plants were more prone to be attacked. 4. Synthesis. Our study indicates a strong control of soil P fertility on the triple interaction among plants, fungal endophytes and native herbivores. This highlights the potential role of increased soil nutrients on the invasion spread of endophytesymbiotic forage plants in natural grasslands.

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