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. 2020 Oct 12;6:37. doi: 10.1038/s41522-020-00148-6

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — a Representative 48 h 3D biofilm images of GFP-tagged PssUMAF0158 wild-type and mutants are shown. The obtained images were analysed with the Leica Application Suite (Mannheim, Germany) and the IMARIS software package (Bitplane, Switzerland). Scale bar 20 µm. b Area in the field of view covered by 48 h biofilms of the GFP-tagged PssUMAF0158 wild-type, extracellular matrix mutants and complemented strains. c Volume in the field of view occupied by 48 h biofilms of the GFP-tagged PssUMAF0158 wild-type, extracellular matrix mutants and complemented strains. The area and volume values were calculated with the IMARIS software package (Bitplane, Switzerland). The following GFP-tagged strains were tested: PssUMAF0158 wild-type (PssUMAF0158 wt), PssUMAF0158 alginate mutant (Δalg8), PssUMAF0158 cellulose mutant (ΔwssE), PssUMAF0158 Psl-like polysaccharide mutant (ΔpslE), PssUMAF0158 Δalg8,wssE double mutant (Δalg8,wssE), PssUMAF0158 Δalg8,pslE double mutant (Δalg8,pslE), PssUMAF0158 ΔwssE,pslE double mutant (ΔwssE,pslE), PssUMAF0158 Δalg8,wssE,pslE triple mutant (Δalg8,wssE,pslE), alginate complemented strain (Δalg8 + pBBRalg8), cellulose complemented strain (ΔwssE + pBBRwssE) and Psl-like complemented strain (ΔpslE + pBBRpslE). A minimal of three replicates, and three independent experiments were performed. Statistical significance was assessed by two-tailed Mann–Whitney test (*p < 0.05, **p < 0.01, ***p < 0.001). Error bars show the standard error of the mean (s.e.m.).