Figure 1.
Relative amount of biofilm formation in the various strains compared to WT (average±standard deviation).
Biofilm were formed for 24 h in the absence (CTRL) and presence of signalling molecules (AHL, BDSF or both; 5 µM). N = 60 for all experiments. Black bars: relative amount of metabolically active cells as quantified with CellTiter Blue. Grey bars: relative amount of total biomass as quantified with crystal violet.
Figure 2.
Representative confocal images of 24-h-old biofilms formed by various B. cenocepacia QS mutants.
Table 1.
MIC (µg/ml) of various antibiotics towards B. cenocepacia J2315 WT and mutant strains.
Figure 3.
Effect of treatment of mature biofilms with tobramycin (A) or meropenem (B).
Antibiotics were added in a concentration equal to 4×MIC for each strain to 24 old biofilms, grown in the absence (CTRL) or presence of signalling molecules (AHL, BDSF or both; 5 µM). Data are expressed as average log (CFU/biofilm) (±standard deviation). N ≥3 for all experiments. Treatment of the mutants resulted in significantly higher reductions than treatment of the WT (p<0.05). In addition, supplementation resulted in a significant decrease in susceptibility compared to the unsupplemented mutant (p<0.05).
Figure 4.
Percent survival of C. elegans (average ± standard deviation) infected with various B. cenocepacia strains in the absence (CTRL) or presence of signalling molecules (AHL, BDSF or both; 5 µM).
The results are expressed as the percent survival after 24 h (black bars) or 48 h (grey bars) of infection and treatment. *: significantly different survival compared to uninfected control (p<0.0001); **: significantly different survival compared to infection with WT (p<0.0001).
Figure 5.
Production of AHL molecules by various B. cenocepacia strains A) in the absence and B) presence of BDSF (5 µM).
*: significantly different from no AHL production (p<0.001); **: significantly different from control receiving no BDSF (p<0.001).
Figure 6.
Production of BDSF by the ΔcepIΔcciI double mutant.
Biofilm formation of the ΔBCAM0581 without supplementation, supplemented with supernatant of the ΔcepIΔcciI mutant or ΔBCAM0581 mutant or supplemented with BDSF (5 µM). The relative amount of metabolically active cells was quantified with CellTiter Blue (black bars), while the relative amount of total biomass was quantified with crystal violet (grey bars).
Figure 7.
Protease production in various B. cenocepacia strains in the abscence (CTRL) and presence of AHL (5 µM), BDSF (5 µM) or both AHL/BDSF (5 µM).
*: significantly different protease production compared to no signal (p<0.05).
Table 2.
Expression of the QS-genes cepI/R and cciI/R and virulence genes zmpA, lipA, lipB and orbI in the ΔBCAM0581, ΔcepIΔcciI and triple mutant in the absence (CTRL) and presence of signal molecules (5 µM).
Table 3.
Strains and plasmids used.
Table 4.
Primers used in this work.