The effects of wastewater effluent on a fish and amphibian species

108 Pages·2016·4.09 MB·English

Clemson University
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All Dissertations Dissertations
5-2009
The effects of wastewater effluent on a fish and
amphibian species
Anthony Sowers
Clemson University, [email protected]
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THE EFFECTS OF WASTEWATER EFFLUENT ON
A FISH AND AMPHIBIAN
SPECIES

A Dissertation
Presented to
the Graduate School of
Clemson University

In Partial Fulfillment
of the Requirements for the Degree
Doctor of Philosophy
Environmental Toxicology

by
Anthony D. Sowers
May 2009

Accepted by:
Dr. Stephen Klaine, Committee Chair
Dr. Cindy Lee
Dr. Aaron Roberts
Dr. William Baldwin
Dr. Michelle Boone
ABSTRACT

Wastewater effluents have been shown to contain a variety of anthropogenic
compounds, many of which have endocrine-disrupting properties. While multiple
laboratory studies have shown the effects of such compounds on an individual basis at
elevated concentrations, little research has attempted to characterize the effects of
exposure to environmentally relevant mixtures of endocrine disruptors. The current study
examined the effects of long-term exposure to graded concentrations of wastewater
effluent on the fathead minnow, Pimephales promelas, and the northern leopard frog,
Rana pipiens. Fathead minnows were exposed from the larval stage through sexual
maturity, while northern leopard frogs were exposed as eggs and grown through
metamorphosis. An F1 generation of fathead minnows was cultured in control water to
test for transgenerational effects of the parental exposure to wastewater effluent. A third
experiment examined the effect of triclosan, an antimicrobial compound commonly
found in wastewater effluent, on the growth and development of the pickerel frog, Rana
palustris. A decrease in the expression of secondary sex characteristics and an increase
in the gonadosomatic index of male fathead minnows exposed to wastewater effluent in
the parent generation were observed. Accelerated reproductive activity and an increase
in the expression of male secondary sex characteristics were observed in the F1
generation whose parent generation was cultured in wastewater treatments. Juvenile
male northern leopard frogs exhibited an increased incidence of individuals with
testicular oocytes. Leopard frogs cultured in the two highest concentrations of
wastewater effluent took significantly longer to reach metamorphosis than both the
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control and the lowest wastewater concentration treatment. Exposure to triclosan did not
significantly affect the growth or development of the pickerel frog. The results of this
study suggest that long-term exposure to wastewater effluent can interfere with the sexual
development of the fathead minnow and can elicit responses in subsequent generations.
Sexual development of male leopard frogs and the metamorphic process in leopard frogs
is particularly sensitive to wastewater exposure.

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ACKNOWLEDGEMENTS
I would like to take this opportunity to thank everyone who has contributed to the
completion of this dissertation. My committee members as well as my labmates were all
instrumental in my success as a PhD candidate. My achievements would not have been
possible without the continued support of my family. I would also like to acknowledge
the United States Environmental Protection Agency for funding me as a research
assistant.

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TABLE OF CONTENTS

Page
TITLE PAGE ..................................................................................................................... i

ABSTRACT ....................................................................................................................... ii

ACKNOWLEDGEMENTS ............................................................................................... iv

LIST OF TABLES ............................................................................................................ vii

LIST OF FIGURES .........................................................................................................viii

CHAPTER 1: Literature review of endocrine disruptors in wastewater effluent and their
biological effects on fish and amphibians ...................................... 1

I. Endocrine disruptors in wastewater effluents ..................................................... 1

II. Fish reproductive endocrinology ....................................................................... 4

III. Effects of estrogenic EDCs on fish .................................................................. 6

IV. Amphibian developmental endocrinology ....................................................... 8

V. Effects of estrogenic EDCs on amphibians ...................................................... 11

VI. References ....................................................................................................... 14

CHAPTER 2: The effects of wastewater effluent on the fathead minnow, Pimephales
promelas........................................................................................ 19

I. Abstract .............................................................................................................. 19

II. Introduction ...................................................................................................... 20

III. Materials & methods ....................................................................................... 22

IV. Results............................................................................................................. 28

V. Discussion ........................................................................................................ 32

VI. References ....................................................................................................... 40

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CHAPTER 3: The effects of wastewater effluent on the northern leopard frog, Rana
pipiens…………………………………………………………… 53

I. Abstract .............................................................................................................. 53

II. Introduction ...................................................................................................... 54

III. Materials & methods ....................................................................................... 56

IV.Results.............................................................................................................. 61

V. Discussion ........................................................................................................ 66

VI. References ....................................................................................................... 73

CHAPTER 4: Long-term exposure of the pickerel frog, Rana palustris, to triclosan...... 82

I. Abstract .............................................................................................................. 82

II. Introduction ...................................................................................................... 82

III. Material & methods ........................................................................................ 84

IV. Results............................................................................................................. 87

V. Discussion ........................................................................................................ 88

VI. References ....................................................................................................... 92

CONCLUSION ................................................................................................................. 96

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LIST OF TABLES

Table Page

2.1 Endocrine-disrupting compounds measured in Mauldin Road Wastewater
Treatment Plant effluent………………………………………… 43

2.2 Estradiol equivalency factors in wastewater effluent .......................................... 44

2.3 Length, weight, condition factor, GSI, and HSI of fathead minnows exposed to
wastewater effluent……………………………………………... 45

2.4 Cumulative egg production of fathead minnows exposed to effluent .................. 46

3.1 Endocrine-disrupting compounds measured in Mauldin Road Wastewater
Treatment Plant effluent………………………………………… 77

3.2 Estadiol equivalency factors in wastewater effluent ............................................. 77

3.3 Survival, HSI, length, weight, and swim speed of northern leopard frogs exposed
to wastewater effluent…………………………………………... 78

4.1 Survival, HSI, length, hind limb length, and weight of pickerel frogs exposed to
triclosan…………………………………………………………..94

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LIST OF FIGURES

Figure Page

2.1 Outline of isolated breeding experiment ............................................................... 47

2.2 Percent survival of fathead minnows exposed to wastewater effluent ................. 48

2.3 Secondary sex characteristics of male fathead minnows ...................................... 49

2.4 Histological sections of fathead minnow gonads.................................................. 50

2.5 Secondary sex characteristics of male fathead minnows in an F1 generation ...... 51

2.6 Testis from fathead minnows displaying gonadal abnormalities .......................... 52

3.1 Time to metamorphosis for the northern leopard frog and the percentage of
individuals reaching metamorphosis by the end of the
experiment………………………………………………………. 79

3.2 Histological sections of northern leopard frog gonads ......................................... 80

3.3 Histological sections of northern leopard frog thyroid glands .............................. 81

4.1 Time to forelimb emergence for the pickerel frog and the percentage of
individuals at forelimb emergence by the end of the
experiment………………………………………………………..95

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Chapter 1: Literature review of endocrine disruptors in wastewater effluent and
their biological effects on fish and amphibians
I. Endocrine disruptors in wastewater effluents:
Municipal wastewater effluents around the world have been shown to contain
measurable concentrations of pharmaceuticals and personal care products (PPCPs) and
natural and synthetic hormones (Nakada et al. 2004, Nakada et al. 2006, Snyder 2008). A
large number of these compounds act as endocrine disruptors (EDC) and interfere with
endocrine system-mediated functions acting as either agonists or antagonists of
endogenous hormones. A survey conducted throughout the United States between 1999
and 2000 by the US Geological Survey found the presence of organic wastewater-derived
contaminants, many of which are known EDCs, in 80% of the 139 surface waters
sampled (Kolpin et al. 2002).
Aquatic systems in arid regions that rely on wastewater effluents as a dominant
portion of their flow and/or volume are vulnerable to elevated EDC concentrations when
compared to systems with significant dilution effects. During dry periods, the flow in the
Trinity River below the Dallas/Ft. Worth metroplex in Texas is comprised of >90%
wastewater effluent (Fono et al. 2006). Boulder Creek, a tributary of the South Platte
River, receives wastewater effluent discharge from the city of Boulder. During low-flow
conditions, >75% of Boulder Creek’s flow is comprised of effluent. A large increase in
the estrogenic activity of Boulder Creek is positively correlated with increased effluent
concentrations (Vajda et al. 2008).
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