The toxicity and estrogenicity of a final treated municipal effluent was examined while flowing through a constructed wetland in north-central Texas, USA. Fish data were collected, and a baseline wetland characterization was performed to assess wetland treatment potential for these effluent properties. Vitellogenin (VTG), gonadosomatic index (GSI), hepatosomatic index (HSI), and secondary sexual characteristics were biomarkers used in fish models to assess aqueous estrogenicity. Biological indicators used to assess overall fish health included hematocrit and condition factor. Estrogenic nature of final sewage treatment works effluent was screened, concurrent with a three-week fish exposure, via gas chromatography/mass spectrometry for target estrogenic compounds, including 17β-estradiol, ethynylestradiol, bisphenol A, nonylphenolic compounds, phthalates, and DDT. The VTG in Pimephales promelas was measured after exposure at four sites in a treatment wetland and was significantly elevated (p < 0.0001) in fish exposed at the inflow site. The GSIs were significantly less (α = 0.001) at the inflow site. At wetland sites closest to the inflow, secondary sexual characteristics, tubercle numbers, and fatpad thickness were less (α = 0.0001) than in laboratory controls. The HSIs and density of male breeding stripes were not significantly different from those of laboratory controls. However, elevated HSIs were found at the inflow site. Hematocrit and condition factors were both less (α = 0.001) in effluent-exposed fish at wetland sites closer to the inflow than in control fish or fish further downstream.