Genetic Basis of Gap Formation Between Migrating Helicobacter pylori Colonies in Soft Agar Assays

<i>Helicobacter pylori</i> is a motile bacterial pathogen that causes severe gastric diseases. <i>H. pylori</i> motility and chemotaxis are key colonization factors. Motility and chemotaxis are studied in many microbes, including <i>H. pylori</i>, using soft agar assays. In these assays, bacteria are inoculated into low-percentage agar and expand in a motility- and chemotaxis-dependent manner. <i>H. pylori</i> similarly expands in soft agar, but, if a plate was inoculated at multiple points, the expanded <i>H. pylori</i> colonies did not merge and left gaps. The basis of these gaps was unknown. We report here that gap formation was not affected by media components such as nutrient and agar concentrations, nor did it require chemotaxis, but it did rely on quorum sensing. To broaden our understanding of this <i>H. pylori</i> property, an <i>H. pylori</i> Tn7 transposon library was screened for mutants that lost gap formation. Fourteen mutants were identified, with transposon sites mapped to genes encoding outer membrane proteins, cysteine-rich proteins, phosphatidyl glycerophosphate synthase, an endorestriction nuclease, and several hypothetical proteins. Our results suggest that <i>H. pylori</i> may use specific proteins to avoid contact with other <i>H. pylori</i>, a behavior that may relate to previous observations that different <i>H. pylori</i> strains do not mix populations in stomach glands.