Bacteria in nature are usually found in complex multicellular structures, termed biofilms, which protect them from numerous environmental threats. The biofilm structure is the end result of collective behavior of different cell types in a coordinated fashion, achieved by the accurate regulation of dedicated genetic programs. So far, it was well established that in the single-cell level once a bacterial cell commits to the biofilm state it represses motility both transcriptionally and post-translationally. However, at the population level, we found a motile cell sub-population that is tightly retained inside the biofilm of Bacillus subtilis, enabling fast occupation of new niches, as well as invasion of neighboring colonies of foreign bacterial species. This phenomenon depends on a unique signal originating in the extracellular matrix (ECM) that surrounds the cells in the biofilm, maintaining a motile cell sub-population. This ECM-based signal acts to induce motility, as mutants deficient in ECM production show lower numbers of motile cells. In particular, the proteinaceous component of B. subtilis ECM appears to be a specific chemical signal that strongly induces bacterial motility. Another line of evidence for the importance of maintaining motility in the biofilm emerges from the highly frequent appearance of hyper-motile suppressors in mutant colonies lacking the proteinaceous component. Using genomic analysis, we found that these suppressors acquired mutations in key regulators of the motility to biofilm switch. We suggest that in the absence of ECM-based induction of motility, a strong selective pressure acts to reinstate motility in the biofilm.
