The Ecological Effects of Endocrine Disruption Dr. David Walker University of Arizona David Walker 1, Nick Paretti 2, Gail Cordy 2, Timothy S. Gross 3, Edward T. Furlong 4, Dana W. Kolpin 5, and Dennis McIntosh 6 1 University of Arizona, Environmental Research Laboratory, 2601 E. Airport Dr., Tucson, AZ USGS., WRD, 520 N. Park Ave, Suite 221, Tucson, AZ USGS-Florida Caribbean Science Center, 7920 NW 71 st St., Gainesville Florida, USGS, National Water Quality Laboratory, Denver Federal Center, P.O. Box 25046, MS 407, Lakewood, CO USGS, WRD, P.O. Box 1230, Iowa City, IA Delaware State University, 1200 N. DuPont Highway Dover, DE

For Our Purposes… An endocrine disruptor is a synthetic (anthropogenic) chemical that when absorbed into the body mimics, blocks, or otherwise alters hormone level, function, or binding and subsequently disrupts normal bodily functions including behavioral and/or strictly physiologic responses.

Aquatic Ecology and Endocrine Disruption An individual organisms ability to better- exploit a resource (or group of resources) in the face of environmental stress and inter-specific competition, coupled with conservation of the genetic material enabling this exploitation, is what drives speciation.

Genetic conservation of traits is initiated, and sustained by, subtle behavioral cues for mating, spawning, aggression, territoriality, avoidance, etc. Any impairment of these behavioral cues or manifestation into physiological or morphological changes has the capability to stunt speciation by lowering fertility and fecundity.

Endocrine Disrupting Compounds FAR more than what can be included in this presentation. By the time breakdown products and metabolites are added to the mix, iterations become astronomical.

Just a Few Examples by Use Category Detergent Metabolites Fire/Flame Retardants Fragrances/Flavors Fuels/PAH’s Herbicides/Insecticides Household Wastewater Compounds Non-Prescription Drugs Plasticizers/Antioxidants Prescription Drugs Steroids

At the landscape scale, those compounds known to be “powerful” EDC’s, but are not environmentally-persistent, exert less of an effect than those persistent, but relatively weaker, compounds.

Quantification and Research Design Issues; One Size Does Not Fit All Mechanistic Understanding Ecological Significance Genetic Biochemical Physiological Behavioral Reproductive Assemblages Histopathological Immunological Bioenergetic Populations

Observational versus Controlled Studies True control and replication is not possible in the field. Laboratory studies with control and replication give up some ecological significance.

Exposure and Causation Several studies have examined the effect of one or a very few EDC’s on the physiological response of an organism. The vast magnitude of compounds in a matrix makes assumptions about individual compounds difficult to ascertain. Non-monotonic dose-response curves

With new and emerging contaminants found almost on a daily basis, making assumptions about exposure and physiologic response must always carry the caveat “of the compounds we analyzed for”…

Grab, Composite, or Integrated Samples? Problems associated with not knowing long-term exposure to fish or other organisms.

Passive Organic Chemical Integrative Sampler (POCIS) and/or Semi-Permeable Membrane Devices (SPMD’s aka “fatbags”)

Of all the tools at our disposal to study complex environmental issues in aquatic ecosystems, a sound understanding of ecological principles as they pertain to these ecosystems is the most essential.

Quantifying Endocrine Disruption in a Threatened and Endangered Fish Species

Unlike semi-arid or north-temperate regions, effluent-dependent water’s (EDW’s) in arid regions usually contain 100% effluent year-round.

The Santa Cruz River Near Tucson, Arizona Flows from Mexico near Nogales, Sonora northward to Tucson, Arizona. The only sections with flowing water are those due to discharge from WWTP’s.

Roger Road WWTP Built in Treated effluent is discharged into the Santa Cruz River or diverted into the city’s reclaimed water system. Treats the wastewater generated by a population of about 419,000. A capacity of 41 mgd and treated an average of 38 mgd from 2004 to Produces secondarily-treated wastewater

Roger Road WWTP Tucson Santa Cruz River

This Study Laboratory study with controls, replicates, and randomization. Use fish native to the region (largely pollution-tolerant). Framework or foundation for refinement of future studies. Varying doses of effluent (“treatments”). Concentrate on long-term, persistent compounds.

Bonytail Chub (Gila elegans)

Treatment Control

Water temperature maintained between o C. Photoperiod was maintained at 12 hours of light and dark

Treatment/Dosages Fish in raceways exposed for 3 months per treatment 1 st treatment = 1/3 by volume treated ww and 2/3 water treated by RO 2 nd treatment = 2/3 by volume treated ww and 1/3 water treated by RO 3 rd treatment = full strength treated ww

Results

Males - Overall 17β-Estradiol Control (n = 6): Treatment (n = 13): = ketotestosterone Control (n = 6): = Treatment (n = 13): = Vitellogenin Control (n = 6): = 0.09 Treatment (n = 13): = 0.32

Females - Overall 17β-Estradiol Control (n = 54): Treatment (n = 47): ketotestosterone Control (n = 54): Treatment (n = 47): Vitellogenin Control (n = 54): 0.18 Treatment (n = 47): 0.18

Synergistic Effects Ratios of primary male and female sex hormones, in “undisturbed” populations would be expected to have an inverse relationship i.e. as one increased, the other would decrease. We could therefore assume that major deviations from this inverse relationship between male and female primary sex hormones, could be attributed to impairment.

ln 17βln 11- keto ln Vtg ln 17β ln 11- keto ln Vtg Control MalesTreatment Males ln 17βln 11- keto ln Vtg ln 17β ln 11- keto ln Vtg

ln 17βln 11- keto ln Vtg ln 17β ln 11- keto ln Vtg Control Females Treatment Females ln 17βln 11- keto ln Vtg ln 17β ln 11- keto ln Vtg

Synergism, feedback mechanisms, and non-linearity of bio-markers makes data reduction necessary to determine trends. Ordination is a good statistical tool but still assumes some degree of linear correlation as would occur with a typical dose-response curve.

Eigenvalue Percent Eigenvectors Impairment Det. Met Fire Ret Fragrances Herbicides HH Waste Plasticizers Males

Eigenvalue Percent Eigenvectors Impairment Det. Met Fire Ret Fragrances Herbicides HH Waste Plasticizers Females

Summary Significant hormonal impairment of both sexes, as compared to controls, at very low concentration of compounds. This impairment could never have been determined in a field study. Commonly-used parametric analyses are often inadequate in determining impairment.

Summary (cont) Determination of either hormonal impairment or endocrine disruption requires using phased biomarkers. –Phase 1: Aromatase/GnRH –Phase 2: GtH I, GtH II –Phase 3: Sex hormones –Phase 4: Protein development (vtg, oocyte, spermiation) –Phase 5: Intersex

Current and Future Research Fertility/fecundity and sex ratio/development of F2 generation. Behavior. Treatments using streambed sediment from affected EDW’s.

This study is highly representative of the biological effect of endocrine-disrupting compounds at the landscape scale.

Questions?