Environmental hormones and their impacts on sex differentiation in fathead minnows

Runoff from lands fertilized with animal manure from concentrated animal feeding operations (CAFOs) is a source of hormones to surface water. In this study we tested the hypothesis that larval fathead minnows exposed to sex steroids singly or in a "typical" CAFO mixture during sex differentiation would respond with changes in the expression of a set of target genes, leading to gonadal abnormalities later in life. In the first experiment, a static daily-renewal system was used to expose larvae during the period of 10-20 days post-hatch (dph) to either 5 ng/L 17β-trenbolone (17β-TRB) or 5 ng/L 17α-ethinylestradiol (EE2). In a second experiment, fish were exposed from 0 to 45 dph in a flow-through system to a CAFO mixture composed of steroids and degradates (2-16 ng/L), atrazine and degradates (15-250 ng/L), and nitrate (3-11 mg/L). In the single hormone experiment, expression of genes involved in steroidogenesis (cyp19a, cyp17, and star) was decreased in females. In contrast, no differences in gene expression were observed in fish exposed to the CAFO mixture. However, the majority (84%) of treated males had testes containing an ovarian cavity, indicative of feminization, compared to 0% in the control males. Overall, our results show that: (1) changes in gene expression after single hormone exposures are sex-specific, with females more responsive than males; and (2) phenotypic alterations in testicular development can be elicited by a simulated "CAFO" mixture when fathead minnows are exposed during the first 45 days of development. More research is needed to further discern the complex response of fish to steroid mixtures, especially those associated with runoff from land-applied CAFO waste.