All ETDs from UAB

Advisory Committee Chair

Robert A Angus

Advisory Committee Members

W Mike Howell

Karolina M Mukhtar

R Douglas Watson

Document Type

Thesis

Date of Award

2014

Degree Name by School

Master of Science (MS) College of Arts and Sciences

Abstract

Environmental endocrine disrupting compounds (EDCs) have become a growing threat to the health of humans and wildlife. Numerous studies have demonstrated the presence of EDCs in freshwater aquatic environments. Environmental estrogens, both natural and synthetic, appear to be posing a threat to the reproductive fitness of aquatic organisms. Wastewater treatment plants (WWTPs) have been identified as significant routes by which the EDCs enter waterways. The egg yolk precursor protein vitellogenin (VTG) has proven to be a useful biomarker that can be used to identify organisms that have been exposed to environmental estrogens. In this study, a quantitative RT-PCR assay for expression of the VTG gene in the largescale stoneroller (Campostoma oligolepis) was developed. The dose-response pattern of VTG gene expression in largescale stonerollers exposed to various known concentrations of 17α-ethynylestradiol (EE2) was characterized. The newly developed assay was used to investigate the effects of the effluents of three WWTPs in the vicinity of Birmingham, Alabama on VTG gene expression in the largescale stoneroller. The largescale stoneroller has not previously been used as a biomonitor organism for the effects of environmental estrogens, and therefore one purpose of this study was to establish it as a potential model organism for future studies, particularly in the southeastern region of the United States, where this species is native. In order to design RT-PCR primers specific to largescale stoneroller VTG, a partial sequence of the VTG gene was obtained. This obtained sequence showed high homology to the VTG sequences of other fish in the Cyprinidae family. Next RT-PCR primers were designed for the obtained VTG sequence and for ribosomal protein L8 (RPL8), which was used as a normalizing gene. This RT-PCR assay was effective in detecting increased VTG gene expression in largescale stonerollers exposed to dietary EE2. It also showed that largescale stoneroller VTG expression was affected by EE2 in a dose-dependent manner. This assay was also effective in detecting increased VTG gene expression in stonerollers collected below the outfall of one of the three WWTPs included in this study.

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