The synthetic estrogen 17α-ethinylestradiol (EE2) is an endocrine-disrupting chemical released into aquatic environments from sewage treatment facilities. We tested the effects of two environmentally relevant concentrations of waterborne EE2, 10 and 100 ng L−1, on reproductive endpoints in the teleost fish Betta splendens. In the first experiment, testes were removed from males and sperm were exposed to EE2 directly through the activation water. Direct exposure to EE2 had no effect on any measure of sperm swimming performance. In the second experiment, we exposed sexually mature male B. splendens to EE2 using a semi-static exposure protocol for 4 weeks. There were no significant treatment effects in the 10 ng L−1 treatment group, but at the 100 ng L−1 dose we found that fish had smaller gonads and reduced sperm swimming velocity. When allowed to interact freely with female conspecifics, males exposed to 100 ng L−1 EE2 built smaller nests and showed a nonsignificant decrease in fertilization success. To investigate further the potential mechanism underlying the decrease in sperm quality, we repeated the chronic exposure experiment and analyzed the ATP content of sperm from fish in each treatment group. We found that males exposed to 100 ng L−1 of EE2 had fewer moles of ATP per sperm than did fish in the other two treatment groups, suggesting that a decrease in intracellular ATP caused a reduction in sperm swimming velocity. The current study adds to the growing body of literature that indicates the risks to aquatic organisms of exposure to environmentally relevant concentrations of EE2. © 2012 Wiley Periodicals, Inc. Environ Toxicol 29: 243–252, 2014.