biostudies-literatureYan JNSF | BIO | Division of Molecular and Cellular BiosciencesHoward Hughes Medical InstituteHHS | NIH | National Institute of General Medical SciencesNIGMS NIH HHSE5337-43https://www.ebi.ac.uk/biostudies/studies/S-EPMC5018804biostudies-literatureUnknown113(36)Biofilms are surface-associated bacterial communities that are crucial in nature and during infection. Despite extensive work to identify biofilm components and to discover how they are regulated, little is known about biofilm structure at the level of individual cells. Here, we use state-of-the-art microscopy techniques to enable live single-cell resolution imaging of a Vibrio cholerae biofilm as it develops from one single founder cell to a mature biofilm of 10,000 cells, and to discover the forces underpinning the architectural evolution. Mutagenesis, matrix labeling, and simulations demonstrate that surface adhesion-mediated compression causes V. cholerae biofilms to transition from a 2D branched morphology to a dense, ordered 3D cluster. We discover that directional proliferation of rProceedings of the National Academy of Sciences of the United States of AmericaVibrio cholerae biofilm growth program and architecture revealed by single-cell live imaging.PMC5018804R01 GM065859NAR37 GM0658595R01GM065859MCB-0948112MCB-1344191Yan JSharo AGWingreen NSStone HABassler BLfalseVibrio cholerae biofilm growth program and architecture revealed by single-cell live imaging.Biofilms are surface-associated bacterial communities that are crucial in nature and during infection. Despite extensive work to identify biofilm components and to discover how they are regulated, little is known about biofilm structure at the level of individual cells. Here, we use state-of-the-art microscopy techniques to enable live single-cell resolution imaging of a Vibrio cholerae biofilm as it develops from one single founder cell to a mature biofilm of 10,000 cells, and to discover the forces underpinning the architectural evolution. Mutagenesis, matrix labeling, and simulations demonstrate that surface adhesion-mediated compression causes V. cholerae biofilms to transition from a 2D branched morphology to a dense, ordered 3D cluster. We discover that directional proliferation of r2016-01-01T00:00:00Z2016 Sep2025-04-19T07:40:09.354Z2019-03-27T02:23:56ZS-EPMC50188042755559210.1073/pnas.1611494113