Novel Genetic Tools to Direct Biosynthesis in Intestinal Bacteria

Recent correlative studies have linked the metabolism of the microbiota to the onset or prevention of various disease states. Complementary advancements in ‘Omics approaches and anaerobic culture techniques have significantly de-risked harnessing diverse native microbiota members as chassis for implementing synthetic biology strategies. Of great import, gut symbionts, namely butyrate-producing Lachnospiraceae, are thought to play disease-protective roles and provide a unique access point for interventions that modulate host physiology. However, our explicit understanding of how these microbes affect host-facing protective mechanisms remains elusive. This knowledge gap is due to a continued dearth of genome manipulation technologies to control the metabolic activity of health-associated gut bacteria. Here, we propose a blueprint to enable genetic engineering in currently intractable symbionts of the Lachnospiraceae family. Establishing these genetic tools allows for direct genotype-to-phenotype analyses of important “beneficial” gut symbionts. Additionally, with new physiological insight of these microbes, we can genetically augment living therapeutics and uncover novel microbial metabolites to treat gut-associated disease. To this end, we co-opted plasmid-based systems to establish genetic engineering in Lachnospiraceae. These tools served as a platform to develop a robust genetic toolkit that enables precise manipulation of gene expression in Lachnospiraceae. Moving forward, we will employ these strategies to regulate the production of potentially beneficial metabolites. The development of these tools will aid precise genetic study of many key symbiotic microorganisms, provide a basis to genetically manipulate other intractable microbes, and allow for tuning microbial function in defined contexts to investigate hypotheses that have remained beyond our reach.