Quantifying the effect of helical cell shape on Helicobacter pylori's motility and niche acquisition

Thesis (Ph.D.)--University of Washington, 2016-03Half of all humans harbor the extracellular pathogen Helicobacter pylori in their stomachs. Successful colonization by H. pylori requires flagellar-based motility for the bacterium to traverse the thick gastric mucus layer and reach its preferred niche, close to the gastric epithelium. Helical cell shape is thought to facilitate H. pylori’s ability to bore into the mucus layer in a corkscrew-like motion, thus enhancing colonization of the stomach. In a mouse model of infection, H. pylori cell shape mutants show impaired stomach colonization highlighting the importance of cell shape in infection and motility. To investigate how cell shape impacts H. pylori’s motility in vitro, I modeled the viscous environment of the human gastric mucosa with physiologic concentrations of purified porcine gastric mucin (PGM). Using single-cell microscopic tracking and quantitative morphology analysis, I document marked variation in both cell body helical parameters and flagellum number among H. pylori strains leading to distinct and broad swimming speed distributions that reflect both temporal variation in swimming speed and morphologic variation within the population. Isogenic mutants with straight rod morphology showed reduced swimming speeds (7-21%) and a higher fraction of immobilized bacteria. Mutational perturbation of flagellum number revealed a 19% increase in swimming speed for H. pylori with 4 vs. 3 median flagellum number. Resistive force theory (RFT)-modeling incorporating both cell morphology and flagellum number variation predicts quantitative speed differences of 10-30% among strains. However, quantitative comparisons suggest RFT underestimates the effect of helical shape on speed. To gain a deeper understanding of how helical cell morphology promotes host colonization by H. pylori, I then performed a time course of single-strain infections with wild-type bacteria, a curved rod mutant (Δcsd1), and a straight rod mutant (Δcsd6). Cell shape mutants show significant attenuation during initial colonization (1 day and 1 week). After persistent infection (1-4 months), a subset of mice infected with Δcsd1 or Δcsd6 mutant bacteria show enhanced infection, while others maintain low or no infection. I used confocal microscopy and 3-D reconstructions of thick tissue sections, and performed volumetric analysis to quantify the number of bacteria within different regions of the stomach. Localization of H. pylori reveals multiple gastric niches and specific deficits for cell shape mutants in colonization of the antral glands early after infection, and altered progression of inflammation and gland hyperplasia after chronic infection. Our studies are elucidating the mechanisms by which helical cell morphology promotes motility and sustains host colonization by H. pylori, which may impact human health and disease