Clonal microcolonies recruit individual planktonic colonizers using extracellular matrix factors to assemble Vibrio cholerae pellicles

Pellicles, expansive bacterial communities that form on liquid surfaces, contain residents that incorporate asynchronously and exist in distinct physiological states. How interactions between cell populations convert segregated founder pellicle microcolonies into a contiguous, macroscale structure is unknown. Using high resolution timelapse microscopy of Vibrio cholerae pellicle formation, we show that the confluent community forms through the recruitment of individual planktonic cells to regions between founder microcolonies. In addition to Type IV MSHA pili, individual planktonic V. cholerae cells attach to the air-liquid interface using their cell-surface-bound vibrio polysaccharide (VPS), which binds to the Bap1 and RbmC adhesins secreted by existing pellicle microcolonies. Planktonic V. cholerae cells readily attach to the interface by VPS-adhesin binding when they exist in the low-cell-density quorum-sensing state because this is the mode that promotes VPS production. Single-molecule FISH of V. cholerae pellicles reveals that regions near pellicle microcolonies, where secreted Bap1 and RbmC adhesin levels are the highest, recruit a higher proportion of VPS-producing planktonic cells than do more distant regions. Thus, existing pellicle microcolony inhabitants in an advanced phase of sessile growth prime the surface for new colonizer cell attachment, driving a spatial pattern of gene expression within the pellicle community.