Evidence of a possible multicellular life cycle in Escherichia coli

Biofilms are surface-attached multicellular microbial communities. Their genetics have been extensively studied, but the cell-scale morphogenetic events of their formation are largely unknown. Here, we recorded the entirety of morphogenesis in Escherichia coli, and discovered a previously unknown multicellular self-assembly process. Unattached, single-cells formed 4-cell rosettes which grew into constant-width chains. After ∼10 cell generations, these multicellular chains attached to surfaces and stopped growing. Chains remained clonal throughout morphogenesis. We showed that this process generates biofilms, which we found are composed of attached clonal chains, aligned in parallel. We investigated genetics of chain morphogenesis: Ag43 facilitates rosette formation and clonality; type-1 fimbriae and curli promote stability and configuration; and extracellular polysaccharide production facilitates attachment. Our study establishes that E. coli, a unicellular organism, can follow a multistage, clonal, genetically-regulated, rosette-initiated multicellular life cycle. These findings have implications for synthetic biology, multicellular development, and the treatment and prevention of bacterial diseases.