FIG. 3.

The GraRS two-component regulatory system is required for citrate-stimulated biofilm formation and aggregation. (A) Biofilm formation in the absence and presence of 0.2% citrate for the indicated strain. *, P < 0.001 compared to WT with no citrate; **, P = 0.026 compared to WT with no citrate; #, P = 0.98 compared to the graR mutant without citrate. (B) Aggregation assay after overnight growth for the indicated strains without and with 0.2% citrate. The arrow indicates the aggregate formed by the WT grown with citrate. (C) Quantification of an aggregation assay for citrate-stimulated aggregation by the WT and the ΔgraS mutant. *, P < 0.001 compared to the WT with no citrate. (D) VISA isolate Mu50 requires a functional graR gene for citrate-stimulated biofilm formation in a 96-well plate assay. Biofilm and aggregation assays were performed as described in Materials and Methods.