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Bioaugmentation and Correlating Anaerobic Digester Microbial Community to Process Function

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Bioaugmentation and Correlating Anaerobic Digester Microbial Community to Process Function Marquette University e-Publications@Marquette Dissertations, Theses, and Professional Dissertations (1934 -) Projects Bioaugmentation and Correlating Anaerobic Digester Microbial Community to Process Function Kaushik Venkiteshwaran Marquette University Follow this and additional works at: https://epublications.marquette.edu/dissertations_mu Part of the Environmental Engineering Commons, and the Environmental Microbiology and Microbial Ecology Commons Recommended Citation Venkiteshwaran, Kaushik, "Bioaugmentation and Correlating Anaerobic Digester Microbial Community to Process Function" (2016). Dissertations (1934 -). 661. https://epublications.marquette.edu/dissertations_mu/661 BIOAUGMENTATION AND CORRELATING ANAEROBIC DIGESTER MICROBIAL COMMUNITY TO PROCESS FUNCTION by Kaushik Venkiteshwaran A Dissertation Submitted to the Faculty of the Graduate School, Marquette University, in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy Milwaukee, Wisconsin August 2016 ABSTRACT BIOAUGMENTATION AND CORRELATING ANAEROBIC DIGESTER MICROBIAL COMMUNITY TO PROCESS FUNCTION Kaushik Venkiteshwaran Marquette University, 2016 This dissertation describes two research projects on anaerobic digestion (AD) that investigated the relationship between microbial community structure and digester function. Both archaeal and bacterial communities were characterized using high- throughput (Illumina) sequencing technology with universal 16S rRNA gene primers. In the first project, bioaugmentation using a methanogenic, aerotolerant propionate enrichment culture was investigated as a possible method to increase digester methane production. Nine anaerobic digesters, seeded with different biomass, were operated identically and their quasi steady state function was compared. Before bioaugmentation, different seed biomass resulted in different quasi steady state function, with digesters clustering into high, medium or low methane (CH4) production groups. High CH4 production correlated with neutral pH and high Methanosarcina abundance, whereas low CH4 production correlated to low pH and high Methanobacterium and DHVEG-6 family abundance. After bioaugmentation, CH4 production from the high CH4- producing digesters transiently increased by 11±3% relative to non-bioaugmented controls (p <0.05, n=3), whereas no functional changes were observed for medium and low CH4 producing digesters. The CH4 production increase after bioaugmentation was correlated to increased relative abundance of Methanosaeta and Methanospirillum originating from the bioaugment culture. In conclusion, different anaerobic digester seed biomass can result in different quasi steady state function. The bioaugmentation employed can result in a period of increased methane production. In the second project, a quantitative structure activity relationship (QSAR) between anaerobic microorganism relative abundance values and digester methane production rate was developed using 150 lab-scale anaerobic digesters seeded with 50 biomass samples obtained from 49 US states. Although all digesters were operated identically for a minimum of 5 retention cycles, their quasi steady-state performance varied significantly, with the average daily methane production rate ranging from 0.09±0.004 to 0.98±0.05 L-CH4/LR-day (average ± standard deviation). Analysis of over 4.1 million-sequence reads revealed approximately 1300 operation taxonomical units (OTUs) at the genus level across all digesters, with each digester having 158±27 OTUs (mean ± standard deviation). Using Spearman’s rank correlation, 10 OTUs, which included one archaeal OTU, were found to correlate significantly to digester methane production rate. The relative abundance values of the 10 OTUs were used as descriptors to develop a multiple linear regression (MLR) equation, with good statistical prediction of the digester methane production rates. The results are encouraging and provide an initial step for further research to develop more robust QSAR models to predict the function of anaerobic and other bioprocesses using microbial community descriptors. i ACKNOWLEDGEMENTS Kaushik Venkiteshwaran I owe my deepest gratitude to my research advisor Dr. Daniel Zitomer for giving me an excellent opportunity to pursue my doctoral studies. My graduate experience has been one that I will cherish forever. I am greatly thankful for his guidance, care, patience, and providing me with an excellent atmosphere for doing research. He offered me a full- time research assistantship throughout my doctoral program that allowed me to focus on my research and not be concerned about my finance. His mentorship was a key in providing a well-rounded experience consistent with my long-term career goals. He encouraged me to not only grow as a scientist but also an independent thinker. I am confident that these professional qualities will help me in my future success. To my committee – Dr. Krassimira Hristova, Dr. James Maki, and Dr. Michael Switzenbaum– I thank you for your insights and willingness to participate in this project. I thank Mike Dollhopf, (Lab Manager, Water Quality Center, Marquette University) for assisting me in lab experimental set-up, and keeping track of lab supplies which helped a lot in completing the research tasks on time without any interruptions. I found him a great person on professional and personal level. I am also thankful to researchers Dr. Jean-Philippe Steyer, Dr. Kim Milferdest, Dr. Jerome Hamelin from the French National Agricultural Institute (INRA), Narbonne France, and Dr. Michael Johnson, Dr. Masanori Fujimoto from Marquette University for valuable inputs and expertise in the areas of bioinformatics and statistical analysis which were an essential part of this dissertation. ii I would also like to thank the undergraduate research assistants Michael Syverson, Andrew Suesse and Michael Billings for their great contribution in assisting, monitoring and operating of my research experiments. Without them, it would have been a challenge for me to collect the huge amounts of data required for this dissertation. I feel so fortunate to associate myself with all my past and present colleagues: Dr. Vaibhav Tale, Dr. Benjamin Bocher, Dr. Navaneethan Navaratnam, Dr. Ujjwal Bhattad, Dr. Rachel Morris, Dr. Daniel Carey, Dr. Matthew Seib, Dr. Prince Mathai, Dr. Zhongzhe Liu, Steve Graziano, Yiran Tong, Emily Gorsalitz, Carlan Johnson, Joseph Heffron, Nick Benn, Anna Avila, Saba Seyedi, Lee Kimbell and Dylan Friss. Thanks to all of them for being great friends and work colleagues. I am so grateful to all donors of Matthews, Joseph A. and Dorothy C. Rutkauskas, Arthur J.Schmitt and Albert and Cecil Lue-Hing Engineering Graduate Scholarships for supporting my education and research. I would like to express my special thanks to Marquette University for giving me a great platform for excellence, leadership and service. Last but not least, thank you as well to all my friends and family who have offered support, interest, and encouragement along the way. iii TABLE OF CONTENTS ACKNOWLEDGEMENTS .............................................................................................................................. i LIST OF TABLES ............................................................................................................................................. v LIST OF FIGURES .......................................................................................................................................... vi 1 Introduction ........................................................................................................................................... 1 2. Relating Anaerobic Digestion Microbial Community and Process Function .......... 5 2.1. Introduction ................................................................................................................................... 5 2.2 Phases in AD Process................................................................................................................. 6 2.2.1 Hydrolysis.............................................................................................................................. 6 2.2.2 Acidogenesis and Syntrophic Acetogenesis .......................................................... 7 2.2.3 Methanogenesis .................................................................................................................. 9 2.2.4 Syntrophic Acetate-Oxidizing Bacteria ................................................................ 11 2.3 Environmental Parameters Affecting Digester Microbial Community .......... 12 2.4 Relating Microbial Community Structure to Digester Stability and Function…. ................................................................................................................................................ 14 2.4.1 Qualitative Structure-Function Relationships .................................................. 14 2.4.2 Quantitative Structure Function Relation ........................................................... 16 2.4.3 Bioaugmentation as a Tool to Study Structure-Function Relationships….. ................................................................................................................................ 19 2.5 Conclusion ................................................................................................................................... 22 3. Anaerobic Digester Bioaugmentation Influences Quasi Steady State Performance and Microbial Community ........................................................................................
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