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. 2013 Aug 8;8(8):e71097. doi: 10.1371/journal.pone.0071097

Figure 3. Impact of over-expression of colanic acid related genes identified in the screen on the histone resistance.

Figure 3

We determined the minimal bactericidal concentration of histones to kill 90% (MBC90, left panels) or 99% (MBC99, right panels) of the initial bacterial inoculum after 1 h of incubation. (A–B) The simultaneous over-expression of wcaI, cpsB, cpsG genes in MACH1 carrying the plasmids pBAD33cpsG pBAD18wcaI/cpsB or pBAD33wcaI pBAD18cpsB/cpsG led to a high histone resistance in comparison with the strain carrying the empty vectors. (C–D) To refine the genes contributing the resistance, we tested the over-expression of wcaI/cpsB, cpsB/cpsG, or single genes, wcaI, cpsB, and cpsG, carried by pBAD18. (E–F) The other set of genes related to the colanic acid, wzc/wcaA, did not show any significant histone resistance in term of MBC90 and MBC99 in comparison with the bacteria carrying the empty vector, while the genes were over-expressed together (MACH1 pBAD18wzc/wcaA) or individually (MACH1 pBAD18wzc and MACH1 pBAD18wcaA). (G) The E. coli over-expressing this different combination of wzc and wcaA genes showed a higher number of survivors after 1 h treatment with histones at 100 µg/ml. To be able to compare the different clones, we standardized the values by dividing the CFU/ml after 1 h treatment by the CFU/ml of the inoculum. Each dot represent the result of one experiment and at least three independent experiments have been done. The differences were considered significant by the Mann-Whitney test with p≤0.05 (*) and p≤0.01 (**).