(A) Adhesion to renal epithelial cells A498 was measured after 30 min. Curliated strains adhered significantly better to renal epithelial cells than strains lacking curli, independent of the expression of cellulose (### P<0.0001, t-test). Cellulose expression decreased the number of cell-associated bacteria in curliated (*** P<0.0001, t-test) and non-curliated strains (** P = 0.001, t-test). Results from three independent experiments in quadruplicates are shown as mean and standard deviation. Similar results were obtained for bladder epithelial cells (data not shown). (B) Induction of IL-8 was measured in culture supernatants of renal epithelial cells A498 stimulated with E. coli for 24 h. Curliated bacteria induced a significantly stronger IL-8 response than the mutants lacking curli in the presence (## P = 0.001, t-test) and absence (### P<0.0001, t-test) of cellulose. In curliated bacteria, the expression of cellulose reduced IL-8 induction (** P = 0.001, t-test). Results from three independent experiments in quadruplicate are shown as mean and standard deviation. Similar results were obtained for bladder epithelial cells (data not shown). (C+D) The phenotype of E. coli No. 12 could be restored by complementation of its mutants. Cellulose expression in strain B23 is inducible by aTc (left panels) and reduces adherence and IL-8 induction (*** P<0.0001 and ** P = 0.007, respectively, t-test). The curli subunits CsgA and CsgB are expressed from pWSK29-csgBA in strain WE11_1 (right panels) and increases adherence and IL-8 induction compared to WE11_1 carrying the vector pWSK29 only (* P = 0.048 and ** P = 0.003, respectively, t-test). Results in A498 cells are shown as mean and standard deviation. Data from three experiments in quadruplicate for adherence and from two experiments in triplicate for IL-8 induction are presented. (E) Mice were infected with the isogenic E. coli strains for 1 h. The curliated mutant were isolated from kidneys in significantly higher numbers than the double knockout (# P = 0.026, Mann-Whitney U test). Individual values from n = 8–10 mice/group and medians are shown. (F) Levels of MIP-2 were measured in kidney tissue of infected mice after 16 h. In the absence of cellulose, the curliated mutant strain induced higher levels of MIP-2 compared to the non-curliated strain (## P = 0.001, Mann-Whitney U test). Expression of cellulose reduced the induction of MIP-2 in the presence (*** P<0.0001, Mann-Whitney U test) and absence of curli (** P = 0.001, Mann-Whitney U test). Individual values from n = 5–10 mice/group and medians are shown.