Cannibalism shapes biofilm structure and composition in Bacillus subtilis

Abstract

Programmed cell death (PCD) is a ubiquitous and crucial mechanism to structure eukaryotic multicellular tissues. PCD-like processes have also been described in bacteria, but their contribution to multicellular development is poorly understood. Cannibalism in Bacillus subtilis has been described as a sporulation delay strategy, in which one subpopulation produces antimicrobial peptides that kill susceptible nonproducing siblings. Their lysis is thought to release nutrients that delay the sporulation in the producing subpopulation. This study comprehensively analyses the role of the three cannibalism toxins in shaping colony biofilms. By combining MALDI-mass spectrometry imaging, colony biopsy, flow cytometry, and luminescence reporters, we demonstrate that cannibalism toxins are crucial for biofilm structure. They show a discrete and interdependent localization within the colonies. While cannibalism inhibits sporulation and causes severe envelope stress within biofilms, our data challenge the established role of cannibalism-dependent killing as the mechanism behind this sporulation delay.