UNLV Theses, Dissertations, Professional Papers, and Capstones December 2016 Occurrence and Proliferation of Antibiotics and Antibiotic Resistance in Wastewater Treatment Plants Majid Neyestani University of Nevada, Las Vegas Follow this and additional works at: https://digitalscholarship.unlv.edu/thesesdissertations Part of the Civil Engineering Commons, and the Environmental Engineering Commons Repository Citation Neyestani, Majid, "Occurrence and Proliferation of Antibiotics and Antibiotic Resistance in Wastewater Treatment Plants" (2016). UNLV Theses, Dissertations, Professional Papers, and Capstones. 2889. http://dx.doi.org/10.34917/10083194 This Dissertation is protected by copyright and/or related rights. It has been brought to you by Digital Scholarship@UNLV with permission from the rights-holder(s). You are free to use this Dissertation in any way that is permitted by the copyright and related rights legislation that applies to your use. For other uses you need to obtain permission from the rights-holder(s) directly, unless additional rights are indicated by a Creative Commons license in the record and/or on the work itself. This Dissertation has been accepted for inclusion in UNLV Theses, Dissertations, Professional Papers, and Capstones by an authorized administrator of Digital Scholarship@UNLV. For more information, please contact [email protected]. OCCURRENCE AND PROLIFERATION OF ANTIBIOTICS AND ANTIBIOTIC RESISTANCE IN WASTEWATER TREATMENT PLANTS By Majid Neyestani Bachelor of Science in Engineering – Civil Engineering University of Qom 2010 Master of Science in Engineering – Civil and Environmental Engineering Isfahan University of Technology 2012 A dissertation submitted in partial fulfillment of the requirements for the Doctor of Philosophy – Civil and Environmental Engineering Department of Civil and Environmental Engineering and Construction Howard R. Hughes College of Engineering The Graduate College University of Nevada, Las Vegas December 2016 0 Copyright 2016 Majid Neyestani All Rights Reserved 1 Dissertation Approval The Graduate College The University of Nevada, Las Vegas November 23, 2016 This dissertation prepared by Majid Neyestani entitled Occurrence and Proliferation of Antibiotics and Antibiotic Resistance in Wastewater Treatment Plans is approved in partial fulfillment of the requirements for the degree of Doctor of Philosophy – Civil and Environmental Department of Civil and Environmental Engineering and Construction Daniel Gerrity, Ph.D. Kathryn Hausbeck Korgan, Ph.D. Examination Committee Chair Graduate College Interim Dean Dale Devitt, Ph.D. Examination Committee Member Donald Hayes, Ph.D. Examination Committee Member Jacimaria Batista, Ph.D. Examination Committee Member Eduardo Robleto, Ph.D. Graduate College Faculty Representative ii Abstract Solids retention time (SRT) is one of the most important factors in designing and operating activated sludge systems for biological wastewater treatment. Longer SRTs have been shown to alter the structure and function of microbial communities, thereby leading to improved treatment efficacy with respect to bulk and trace organics, nutrient removal, and membrane fouling. However, research has also shown that longer SRTs lead to increased prevalence of antibiotic resistant bacteria, perhaps due to increased exposure to antibiotics present in influent wastewater. The purpose of this study was to characterize changes in microbial community structure in a laboratory-scale activated sludge system as a function of SRT (2-20 days) and influent concentrations (1x-100x ambient concentrations) of five antibiotics: ampicillin, sulfamethoxazole, tetracycline, trimethoprim, and vancomycin. Also, this research aimed to characterize the role of SRT and elevated antibiotic concentrations on AR proliferation in biological treatment processes. Changes in microbial community structure were evaluated based on traditional plating methods and 16s rDNA sequencing, and microbial community function was evaluated based on changes in effluent water quality, including bulk organic matter characterization and antibiotic concentrations. Spread plate technique was used to determine the number of Gram positive Staphylococcus/Streptococcus strains. The extent of AR was also determined based on minimum inhibitory concentrations (MICs) of resistant isolates. The results indicated that SRT—but not antibiotic loading—had a significant impact on microbial community structure (e.g., reduction in relative prevalence of Acinetobacter and Arcobacter) and effluent water quality. Therefore, spikes in influent antibiotics (at sub-therapeutic concentrations) are not expected to adversely impact biological wastewater treatment. The results revealed that longer SRTs and higher antibiotic concentrations select for antibiotic resistant bacteria (ARBs). iii The data obtained from this study suggests that longer SRTs may select for trimethoprim- resistant bacteria and/or result in false positives for trimethoprim resistance due to higher concentrations of free thymine or thymidine. iv Acknowledgements My most sincere thanks go to my advisor, Dr. Daniel Gerrity. I thank him for his guidance, encouragement and support. I would like to thank my committee members for their valuable advice and time. I would also like to thank the personnel at the study site, particularly Bruce Dacko, and Peter Faught at UNLV for their assistance in designing and setting up the experimental system. Finally, I would like to express my eternal gratitude to my wife and my family for their everlasting love and support. v Dedication I dedicate this dissertation to my wife, Elnaz, and to my parents, Abbas and Fatemeh, with my gratitude for their love, support, and inspiration throughout my life. vi Table of Contents 1.0 Introduction ...................................................................................................................................... 1 1.1 Environmental reservoirs of thymidine as a mechanism of trimethoprim resistance: ................. 2 Background .......................................................................................................................................... 2 Research question ................................................................................................................................. 3 Hypothesis ....................................................................................................................................... 3 Approach .............................................................................................................................................. 3 Outcome ............................................................................................................................................... 3 1.2 Microbial community structure and function: ............................................................................. 4 Background .......................................................................................................................................... 4 Research question 1 .............................................................................................................................. 4 Hypothesis 1 .................................................................................................................................... 4 Research question 2:............................................................................................................................. 5 Hypothesis 2 .................................................................................................................................... 5 Approach .............................................................................................................................................. 5 Outcome ............................................................................................................................................... 5 1.3 Relative abundance and extent of AR: ........................................................................................ 6 Background .......................................................................................................................................... 6 Research question 1 .............................................................................................................................. 6 Hypothesis 1 .................................................................................................................................... 7 Research question 2 .............................................................................................................................. 7 Hypothesis 2 .................................................................................................................................... 7 Approach .............................................................................................................................................. 7 Outcome ............................................................................................................................................... 7 2.0 State of knowledge ........................................................................................................................... 8 vii 2.1 Antibiotic Resistance as an Emerging Threat .............................................................................. 8 2.2 Drivers of Antibiotic Resistance ...............................................................................................