Mechanical forces shape heterogeneous bacterial microcolonies as a precursor for multicellular differentiation

Microcolonies are aggregates of a few dozen to a few thousand cells exhibited by many bacteria. The formation of microcolonies is a crucial step towards the formation of more mature bacterial communities known as biofilms, but also mark a significant change in bacterial physiology. Within a microcolony, bacteria forgo a single cell lifestyle for a communal lifestyle hallmarked by high cell density and physical interactions between cells potentially leading to differentiation. It is thus crucial to understand how this transition occurs: how identical single cells assemble in these tight communities and might start to behave differently. Here we show that cells in the microcolonies formed by the human pathogen Neisseria gonorrhoeae (Ng) present differential motility behaviors within an hour upon colony formation. Observation of merging microcolonies and tracking of single cells within microcolonies reveal a heterogeneous motility behavior: cells close to the outside of the microcolony exhibit a much higher motility compared to cells towards the center. Extensive numerical simulations of a biophysical model for the microcolonies at the single cell level indicate that mechanical forces exerted by the bacterial cells are sufficient to generate the observed heterogeneous motility. Further corroborating this idea, experiment with cells lacking the ability to exert forces on their surroundings leads to their segregation on the outside of microcolonies as predicted by the biophysical model. This emergence of differential behavior within a multicellular microcolony is thus of mechanical origin and is likely the first step toward further bacterial differentiation and ultimately mature biofilms.