Atomic force microscopy: A powerful tool for studying bacterial swarming motility

Swarmingmotility is a fascinating phenomenon by which some bacteria use flagella to move over solid surfaces. Understanding the molecular mechanisms underlying swarmingmotility requires studying the factors that induce and control flagella expression in swarming cells. Traditionally, flagella are observed by optical or electron microscopy, but none of these techniques combine versatility and easiness, with quantitative and high-resolution information. We report an atomicforce microscopy (AFM)-based approach for the fast imaging of bacterial phenotypes (cell shape, flagella expression) in swarmingmotility studies. Cells from the Gram-positive bacterium Bacillus thuringiensis sv. israelensis were inoculated on energy-rich media containing increasing agar concentrations. Following swarming assays (2 days), the cell morphology and the amount of flagella were directly observed by AFM imaging in air. Consistent with the macroscopic swarming behavior, cells harvested from the rim of colonies spreading on soft agar were hyperflagellated, elongated and arranged in chains. Increasing the agar concentration led to much lower amounts of flagella and to shorter rod-shaped cells, a finding consistent with the slower swarmingmotility of the cells. Cells taken from colony centers on soft and hard agar surfaces were generally non-flagellated, rod-shaped, rarely arranged in chains, and exhibited lysis and sporulation. This study shows that AFM imaging can readily discriminate between swarming and non-swarming cells, and quantify their morphological details, thus offering an important tool to study the dynamics of bacterial populations