Bacterial cell walls are usually composed of an interconnected mesh made of peptidoglycan (PG). PG is unique to these microorganisms. It is essential for bacterial survival, and has a remarkably conserved structure throughout diverse bacterial groups. PG is not found in plants and animals, and given its essentialness and conservation in bacteria, it is an excellent target for antibiotics and innate immunity. Thus, PG is a focus of strategies for bacterial detection and control. Blocking the ability of bacteria to make a normal PG structure is generally lethal, yet in rare instances bacteria with novel PG have evolved naturally. This project uses the model animal-associated bacterium Vibrio fischeri to explore what happens when PG is experimentally forced to evolve. The results will shed light on the natural evolution of peptidoglycan, on the constraints of its function in bacteria, and on the limits of targeting PG as a means of controlling bacteria. Moreover, V. fischeri is a natural symbiotic bacterium that colonizes a Hawaiian squid, and PG is a key signaling molecule in this symbiosis. Thus, it offers the opportunity to examine how peptidoglycan structure affects the ability of host animal tissue to detect and respond to its resident bacteria (i.e., its microbiome). The broader impacts of this work for society include interdisciplinary graduate and undergraduate student training, outreach to community K-12 schools, and contributions to a book on bacterial symbionts geared for use in the classroom.
This project will elucidate how unusual changes to bacterial cell-wall peptidoglycan (PG) structure can evolve, and it will help define constraints on PG evolution. This project will examine how the tractable bacterium Vibrio fischeri can evolve new PG structure(s) using a strategy of iteratively blocking both normal and alternative pathways to PG biosynthesis. Viable mutants generating novel PG will be selected. Such mutants are likely to have distinctly non-wild-type properties of growth, cell shape, motility, etc.; however, rounds of growth in culture will lead to evolved strains that have accommodated the new PG. A combination of phenotypic testing and whole-genome resequencing will underpin a systems-type analysis of these evolved strains. The integrated analysis of genetic and phenotypic data will improve our understanding of how the cell wall is coordinated with other cellular components and processes, and it will lead to the discovery of new phenotypically important interconnections in bacterial cells. The results will inform our understanding of the natural evolution of PG and lead to a more predictive understanding of how PG might evolve in the future, e.g. under selective pressure from antibiotic use. V. fischeri is a symbiont in a model squid symbiosis, in which symbiont PG triggers developmental changes in the host. This study will provide insight into the relationship between PG structure and its function in this symbiosis, with broader implications for understanding innate immunity in higher organisms.
