Fig 3.

DipA plays a minor role in biofilm formation by P. aeruginosa. (A) Biofilms of strains PA14 and PAO1 inactivated in or (over)expressing dipA, grown for 144 h, were visualized by CLSM. P. aeruginosa biofilms in the absence/presence of the empty plasmid (pMJT1) were used as controls. Biofilms were stained with the LIVE/DEAD BacLight viability stain (Invitrogen Corp.). White size bars = 100 μm. (B) c-di-GMP levels of P. aeruginosa PA14 biofilms inactivated in or (over)expressing dipA. “c-di-GMP/mg” refers to c-di-GMP levels per total cell pellet protein (in mg) used. *, significantly different from PA14 biofilms (P < 0.05), as determined by ANOVA and SigmaStat. (C) The dispersion-deficient phenotype of ΔdipA biofilms is restored upon complementation with dipA in trans. Biofilms were grown for 5 days before dispersion was induced by a sudden increase in the medium glutamate concentration. Biofilm effluents were collected for a period of 18 min, and the absorbance was determined at 600 nm. Determination of c-di-GMP levels was repeated at least 6 times, while all other experiments were carried out in triplicate. Error bars indicate standard deviations.