Ajo-Franklin, Caroline M2023-01-032022-122022-11-30December 2Tolar, Joe G. "Expanding the Understanding of Extracellular Electron Transfer Mechanisms and the Physiological Impacts in Gram-Positive Bacteria." (2022) Diss., Rice University. <a href="https://hdl.handle.net/1911/114197">https://hdl.handle.net/1911/114197</a>.https://hdl.handle.net/1911/114197EMBARGO NOTE: This item is embargoed until 2024-12-01Lactic acid bacteria are named for their nearly exclusive fermentative metabolism. Thus, the recent observation of extracellular electron transfer (EET) activity - typically associated with anaerobic respiration in Gram-negative bacteria - in this class of organisms has forced researchers to rethink the rules governing microbial metabolic strategies (Tejedor-Sanz et al., 2022). Lactic acid bacteria also differ from model EET microorganisms in their cell envelope structure and the niches they inhabit. These differences open important questions: What mechanisms do lactic acid bacteria use to accomplish EET? What are the physiological effects of EET on lactic acid bacteria? Here I address these questions by exploring the role of quinones and flavins in EET by Lactiplantibacillus plantarum (Chapter 2), by developing methods to monitor gene expression in L. plantarum under EET conditions (Chapter 3), and by characterizing the physiological implications of EET in Enterococcus faecalis (Chapter 4). I discovered that either the quinone, 1,4-dihydroxy-2-naphthoic acid (DHNA), or riboflavin support EET via similar, but unique pathways. Using genetic knockouts of candidate genes and electrochemical techniques, I constructed a working model of the EET mechanism for DHNA and riboflavin. Next, I developed a methodology to screen and validate fluorescent reporters of promoter activity under EET conditions to provide insight into the mechanism of the EET-induced fermentative metabolism changes. Additionally, I found that growth conditions and carbon sources can dramatically influence EET activity in E. faecalis. Lastly, I developed a method to quantify interspecies competition between E. faecalis and Staphylococcus aureus and simultaneously measure EET activity. Understanding the role of EET in lactic acid bacteria is of great importance due to the significance of lactic acid bacteria in agriculture, bioremediation, food production, and gut health. This work expands our molecular-level understanding of EET in Gram-positive microbes and provides additional opportunities to manipulate EET for biotechnology.application/pdfengCopyright is held by the author, unless otherwise indicated. Permission to reuse, publish, or reproduce the work beyond the bounds of fair use or other exemptions to copyright law must be obtained from the copyright holder.lactic acid bacteriaquinonesflavinsextracellular electron transfer mechanismmetabolismmicrobiologyThesis2023-01-03