Utah State University DigitalCommons@USU All Graduate Theses and Dissertations Graduate Studies 8-2013 Utilizing Municipal and Industrial Wastes for the Production of Bioproducts: from Metagenomics to Bioproducts Joshua T. Ellis Utah State University Follow this and additional works at: https://digitalcommons.usu.edu/etd Part of the Biomedical Engineering and Bioengineering Commons Recommended Citation Ellis, Joshua T., "Utilizing Municipal and Industrial Wastes for the Production of Bioproducts: from Metagenomics to Bioproducts" (2013). All Graduate Theses and Dissertations. 1702. https://digitalcommons.usu.edu/etd/1702 This Dissertation is brought to you for free and open access by the Graduate Studies at DigitalCommons@USU. It has been accepted for inclusion in All Graduate Theses and Dissertations by an authorized administrator of DigitalCommons@USU. For more information, please contact [email protected]. UTILIZING MUNICIPAL AND INDUSTRIAL WASTES FOR THE PRODUCTION OF BIOPRODUCTS: FROM METAGENOMICS TO BIOPRODUCTS by Joshua Todd Ellis A dissertation submitted in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY in Biological Engineering Approved: Charles D. Miller, PhD Ronald C. Sims, PhD Committee Co-Chairman Committee Co-Chairman H. Scott Hinton, Dean of the Issa Hamud, MS, PE College of Engineering Committee Member Committee Member Jon Takemoto, PhD Mark McLellan, PhD Committee Member Vice President for Research and Dean of the School of Graduate Studies UTAH STATE UNIVERISTY Logan, Utah 2013 ii Copyright © Joshua Todd Ellis 2013 All Rights Reserved iii ABSTRACT Utilizing Municipal and Industrial Wastes for the Production of Bioproducts: from Metagenomics to Bioproducts by Joshua T. Ellis, Doctor of Philosophy Utah State University, 2013 Major Professors: Dr. Charles Miller and Dr. Ron Sims Department: Biological Engineering Global energy requirements are heavily dependent on fossil fuels such as oil, coal, and natural gas. With the expectation of fossil fuels being exhausted in the future, novel strategies need to be discovered for alternative energy generation. Biofuels such as acetone, butanol, ethanol, and hydrogen gas are gaining interest as high value energy sources. These fuels can be produced by anaerobic clostridia as metabolic byproducts of fermentation. The capability to produce these biofuels has been widely studied using glucose or other common feedstocks. Biofuels from renewable and industrial waste feedstocks such as algae and cheese whey may have significant implications on the efficiency of biofuel production, where the price associated with feedstocks is considered a major bottleneck in biotechnology processes. Algae and cheese whey are both rich in organic nutrients and can be utilized by clostridia to produce not only biofuels, but also bioacids, which are considered fuel intermediate compounds. Additionally, understanding microbial communities both in the biosphere and within bioreactors can provide iv knowledge on microbial relationships and novel microbes, and provide knowledge to optimize engineered systems for biofuels and bioremediation strategies. In this study, a comprehensive investigation of the Logan City Wastewater Lagoon System at the microbial level was executed. Microalgae were utilized for the production of acetone, butanol, and ethanol using Clostridium saccharoperbutylacetonicum. High-throughput 454 pyrosequencing technology was utilized to understand the biogas-producing microbial consortium within an algal-fed anaerobic digester inoculated with lagoon sludge. This technology platform was also utilized to study the microbial diversity of a municipal waste remediating community while probing for clostridia capable of producing biofuels. Bioproduct producing clostridia from this system were isolated and employed using cheese whey as feedstock for the production of hydrogen, ethanol, acetic acid, butyric acid, and lactic acid. Integrating fundamental science with engineering strategies was demonstrated using this lagoon system. To optimize and fully understand and manage anaerobic microbial systems, an understanding of their phylogeny and their capabilities are vital for success at the industrial level for the production of high value bioproducts. (262 pages) v PUBLIC ABSTRACT Utilizing Municipal and Industrial Wastes for the Production of Bioproducts: from Metagenomics to Bioproducts Developing renewable sources of energy is gaining interest due to limited supplies, rising costs, and environmental impacts of exploiting fossil fuels. Biosolvents such as acetone, butanol, and ethanol are attractive sources of fuel which can aid in replacing our dependence on foreign oil. Butanol is of particular interest due to its ability to directly replace gasoline, thus considered a drop-in-fuel. Biological hydrogen and methane gas are also attractive energy sources that can lower dependence on fossil energy. This research focused on incorporating DNA sequencing technologies in parallel with fermentation methods for understanding and producing energy. The natural environment provides diverse and uncultured communities of bacteria and microalgae that can be fingerprinted with DNA technologies, isolated and/or cultivated, and employed for energy production. Anaerobic bacteria were isolated from the environment and utilized to produce energy from both algae and cheese whey, which are abundant and energy rich feedstocks. Additionally, these DNA sequencing technologies were utilized to understand the biogas and bioremediating communities within the Logan City Wastewater Lagoon System. Comprehending microbial communities and their interactions is essential in optimizing industrial fermentations and isolating novel organisms of interest. This work demonstrates the capability to probe complex environments for community structures and vi functions using molecular techniques, while subsequently integrating this knowledge in the laboratory to produce high value bioproducts. Joshua T. Ellis vii VISUAL ABSTRACT viii DEDICATIONS This work is dedicated to my parents Mary K. and Stephen T. Ellis, and to Kristen A. Williams for their love and support. ix ACKNOWLEDGMENTS I would like to begin by thanking my major advisers, Dr. Charles Miller and Dr. Ronald Sims. I appreciate your camaraderie, enthusiasm, and support throughout the years. You both have been exceptional advisers and I thank you both very much for the opportunity. I would also like to thank my committee, Dr. Jon Takemoto, Mr. Issa Hamud, and the Dean of the College of Engineering, H. Scott Hinton. To those of you who share a passion for engineering microbial systems and developing alternative energy at Utah State University, I thank your friendship and hard work on these projects within the Department of Biological Engineering. I would like to recognize Asif Rahman, Cody Tramp, Ashik Sathish, and Reese Thompson for research support. Likewise, I would like to thank Neal Hengge for his dedication as an undergraduate researcher; you truly set the bar high amongst your peers and I wish you further success. I would like to thank the Department of Biological Engineering for the opportunity to complete my doctoral work here at Utah State University. And finally I thank the Utah Science Technology and Research (USTAR) initiative, the Utah Water Research Laboratory (UWRL), and the Sustainable Waste-to-Bioproducts Engineering Center (SWBEC) for funding these projects. Joshua Ellis x CONTENTS Page ABSTRACT .................................................................................................................................... iii PUBLIC ABSTRACT ..................................................................................................................... v VISUAL ABSTRACT ................................................................................................................... vii DEDICATIONS ............................................................................................................................ viii ACKNOWLEDGMENTS .............................................................................................................. ix LIST OF TABLES ........................................................................................................................ xiii LIST OF FIGURES ...................................................................................................................... xiv TERMINOLOGY ......................................................................................................................... xxi CHAPTER 1. INTRODUTION ...................................................................................................................... 1 1. Overview ................................................................................................................. 1 2. Engineering significance ......................................................................................... 4 3. Supporting patents .................................................................................................. 6 4. Bioremediation of domestic wastewater and production of bioproducts …… ….from microalgae using waste stabilization ponds ................................................... 6 5. Format of dissertation ............................................................................................. 9 6. References ............................................................................................................... 9 2. LITERATURE