Fig. 3: Identification of a fimE mutant reverting biofilm formation despite high MEcPP levels.
From: An evolutionarily conserved metabolite inhibits biofilm formation in Escherichia coli K-12
a Biofilm production assessed in the high-MEcPP accumulating strain ispg-2, fimE transposon-insertion revertant ispg-2 fimE::Tn5 and knockout mutants ispg-2 ∆fimE::kan, fimE overexpression strains ispg-2 fimE::Tn5 pTrc-fimE-sfGFP and ispg-2 fimE::Tn5 pTrc-fimE-FLAG. The upper panel displays crystal violet-visualized biofilm in PVC tubes, while the lower panel shows relative biofilm mass calculated as the ratio between the OD570nm of dissolved crystal violet-stained biofilm and the OD600nm of planktonic cells at the time of staining. The data are obtained from three biological replicates per genotype, with each biological replicate containing one or two technical replicates. Bars represent mean values, and error bars represent standard deviations. b–e AFM imaging and statistical analyses of fimbriae production examined in CRISPRi strains and the ispg-2 fimE::Tn5 revertant (d), as well as in the MG1655 strain grown in increasing concentrations of benzyl viologen (e). Panels (b and c) display box and whisker plots of the apparent number of fimbriae outside of the aforementioned E. coli cells. The box represents the interquartile range from the 25th to 75th percentiles, with a horizontal line inside indicating the median value; the whiskers extend to the minimum and maximum values in the data, and all individual data points are depicted as dots. The apparent number of fimbriae is counted across all cells within a minimum of six 50 × 50 µm imaging fields, using slides from two biological replicates per genotype. Statistical tests used are Brown-Forsythe and Welch ANOVA and two-sided Dunnett’s multiple comparisons tests (a) and Kruskal-Wallis ANOVA and two-sided Dunn’s multiple comparisons tests (b, c). Panels (d and e) show representative AFM images of the analyzed cells, with white arrowheads pointing at clusters of fimbriae and white asterisks denoting flagella, with scale bars indicating 2 µm. Source data are provided in a Source Data file.
