| We are living in a microbial world, surrounded by uncountable numbers of bacteria that are in a constant state of warfare over limited nutritional resources. The gram-positive soil- and plant-associated bacterium Bacillus subtilis is an adept competitor, growing rapidly when nutrients are plentiful, but acting aggressively once nutrients become scarce. During stationary phase, wild strains (3610) of B. subtilis can sporulate, produce antimicrobials and cannibalism factors, take up DNA from the environment, swarm, and form multicellular biofilms. These behaviors are coordinated by the stationary phase master regulator Spo0A, which is activated by phosphorylation through a complex phospho-relay. In turn, Spo0A∼P regulates the expression of over 121 stationary phase genes.;Here, we demonstrate that four of the five phospho-relay histidine kinases, KinA, KinB, KinC, and KinD are partially redundant but each contribute to proper biofilm formation. Furthermore, these four kinases fall into two classes with different spatiotemporal patterns of activity within colony biofilms; KinA and KinB are active in the older, inner region of the colony biofilm, whereas KinC and KinD are active in the younger, outer regions.;Next, we demonstrate that domesticated strains of B. subtilis are unable to form proper biofilms because they have accumulated five biofilm altering mutations. These include mutations in sfp, required for surfactin production, epsC, required for matrix exopolysaccharide production, and regulatory genes swrA and degQ. Finally, domesticated strains of B. subtilis are missing the plasmid encoded locus rapP, a putative response regulator aspartate phosphatase. Together, these five mutations explain the biofilm defects seen in domesticated strains of B. subtilis.;Finally, we explore the killing of domesticated strain PY79 by 3610, due to production of the antimicrobial sublancin. Sublancin production is controlled in parallel to biofilm formation, requiring Spo0A activation by KinC and KinD, but not KinA and KinB. In addition, in strain 3610, sublancin production is decreased in biofilm matrix overproducing strains. These findings support a model in which antimicrobial production occurs at low Spo0A∼P levels in the outer regions of the biofilm, while the high levels of activated Spo0A∼P in the inner regions of the biofilm lead to early sporulation. |
