1. Effects of 17α -Ethinyl Estradiol on the Reproductive Success of Freshwater Fish ENSC 202 April 24, 2008 Katie Chang Lucas Chapman Jackie Travers Alea Tuttle

2. Overview Introduction to 17α-ethinyl estradiol (EE2) EE2 in wastewater treatment effluent Effects of EE2 on the sexual characteristics and development of individual fish Effects of EE2 on reproductive success of fish populations Case study Kidd et al. 2004

3. 17-ethynyl-13-methyl-7,8,9,11,12,13,14,15,16,17- decahydro-6H- cyclopenta[a] phenanthrene-3,17-diol (image source:wikipedia.org, 2008) WikimediaCommons.org

4. Problem Statement EE2, distributed in freshwater systems by wastewater effluent discharge, negatively impacts reproductive success in freshwater fish.

5. A Day in the life of EE2 Copyright © 2004 American Chemical Society

6. En route to treatment Image source: http://www.wplwloo.lib.ia.us/wfr/trng3.html All estrogens EE2

7. EE2 removal in Sewage Treatment http://www.waterencyclopedia.com/ Ineffective at breaking down E1 and EE2 Hydrophobicity of EE2 plays a role in Activated Sludge treatment Studies inconsistent due to detection limits and variability among treatment type, and loading. Alternative treatment: Physical:Sonolysis UV radiation with HOOH Biological: White rot fungi-laccase Rhodococus Bacteria

8. Surface Water and Effluent Concentrations Taipei Taiwan: Dan-Shui River surface water = 15 ng/L treatment effluent = 26 ng/L (Chen et al. 2007) 3 out of 7 British STW effluent had detectable concentrations between 0.2-7 ng/L (Desbrow et al. 1998) Vs. What is “ Environmentally Relevant ” ? Tricky to Answer. Depends on: % of waterbody discharge effluent strength detection limit plays a role

9. Effects of EE2 on Sexual Development of Individual Fish Feminization of Fish At lifetime exposure to 5ng/L EE2 VTG production in males phenotypic changes of males* no expressible sperm no normal testes development ovary-type tissue development in males *effect also seen at lifetime exposure to 0.96ng/L EE2 http://cobbers.com/wp-content/uploads/2006/09/lady-fish.jpg

10. Blood VTG concentrations for a zebrafish population exposed to EE2 for 40 days (A), and an offspring population with life-long exposure (B). (Nash et al., 2004)

11. Effects of EE2 on Sexual Development of Individual Fish Egg Production and Embryo Viability Number of eggs produced after exposure Lifetime exposure to 0.5ng/L and 5.0ng/L (Nash, et al., 2004) >3.5ng/L (Parrott, et al., 2004) 3 week exposure to 0.1ng/L and 1.0ng/L (Pawlowski, et al., 2004) embryo survival rates Lifetime exposure to 0.2ng/L and 1.0ng/L (Lange, et al., 2001) 0.5ng/L and 5.0ng/L (Nash, et al., 2004) 10ng/L (Balch et al., 2004) reduced gonadosomatic index Lifetime exposure to >3.5ng/L (Parrott, et al., 2004) Mortality 10 day exposure to 50ng/L (Nash, et al., 2004)

12. Breeding Behavioral Effects Little research done Nash, et al. study (zebrafish) –Increased aggression in males Wolfand, et al. study (fathead minnows) –Decreases competitiveness and aggression in males –Lowered male interest in cleaning nests

13. (Wolfand, et al., 2007)

14. Acute vs. Chronic Effects Duration and timing of exposure key Different for each species Studies: –Schafer, et al. –Nash, et al.

15. Schafer et al. (zebrafish) Three experiments –F1 - partial life cycle (0-75days post fertilization (dpf)) –F1 - full life cycle –F2 - full life cycle Four concentrations –(0.05, 0.28, 1.7, and 10ng/L) Zebrafish exposure to 9.3ng/L –Full life cycle: reproduction unsuccessful –Partial life cycle: able to recover

16. Schafers, et al., (2007)

17. Schafers, et al., 2007

18. LOAEL Values calculated by Schafer et al. For zebrafish* –First generation: 1.1ng/L –Second generation: 2.0ng/L Reduced fecundity and fertility –23% and 45% in F1 generation –83% and 98% in F2 generation *Will be different in other species, e.g. Chinese Rare Minnow (Zha et al., 2008)

19. Karen Kidd et al. Study What makes the Kidd et al. study so important? Where did it take place? What [EE2] was achieved during the experiment? Why? What type of fish was used? Why? Conclusions? What kind of species are at risk?

20. Kidd et al. results Both male and female fathead were found to have increased VTG production Male gonad development was effected: Sperm mobility and production decreased, ovatestes. Altered oocytes in females Population collapse due to loss of YOY Short lived species may be at the greatest risk of population collapse after chronic EE2 exposure.

21. Questions?