FIG. 1.

Citrate stimulates cell-to-surface and cell-to-cell interactions. (A) The number of cells per microscopic field was determined at 45 min postinoculation by phase-contrast microscopy in the absence (no add'n) and presence of 0.2% citrate. *, P < 0.01 compared to no addition. (B) Visual aggregation assay with the WT and the fnbB mutant grown overnight with 0% or 0.2% citrate. The white arrow indicates the typical WT aggregate, and the black arrow indicates the altered aggregate formed by the fnbB mutant. (C) PIA/PNAG levels measured by dot blot assay. Extracts of the WT strain grown in the absence or presence of citrate were prepared as described in Materials and Methods and spotted in a series of fivefold dilutions. (D) Biofilm formation of the WT and fnb mutants was assessed in a 96-well plate assay in the absence or in the presence of 0.2% citrate (+Citrate). (E and F) Biofilm formation (E) and aggregation (F) phenotypes were assessed for the WT strain grown in the presence and absence of citrate and/or heparin. Biofilm formation and aggregation assays were performed as described in Materials and Methods.