Figure 3. The PA4834 gene product plays a role in iron acquisition.

(A) Effect of ferric iron on growth of the ΔPA4834 mutant in AMS. Growth indices of PAO1 and the ΔPA4834 mutant were measured after growth in AMS without (grays bars) or with (black bars) 100 μM FeCl₃. Six independent experiments were performed, and mean values ± SD (error bars) are displayed in each bar. *p < 0.05 (students’ t-test) vs. growth of the ΔPA4834 mutant in AMS without FeCl₃. (B) Effect of an iron chelator on bacterial growth. Growth indices of PAO1 (gray bars) and the ΔPA4834 mutant (black bars) were calculated after growth in LB supplemented with increasing concentrations of 2, 2′-bipyridyl. Bacterial cells were grown for 12 h. Three independent experiments were performed, and mean values ± SD (error bars) are displayed in each bar. *p < 0.05 (students’ t-test) vs. PAO1 growth in the same media. (C) Intracellular iron concentration of PAO1 (gray bars) and the ΔPA4834 mutant (black bars) after growth in LB or in AMS. Inductively coupled plasma mass spectrometry (ICP-MS) was used for the measurement and iron concentrations are displayed in parts per billion (ppb). Three independent experiments were performed, and mean values ± SD (error bars) are displayed in each bar. *p < 0.05 (students’ t-test) vs. all other values. (D) Fluorescent microscope images of PAO1 and the ΔPA4834 mutant. After growth in LB or AMS, bacterial suspensions were stained with PhenGreen™ SK, diacetate. Experiments were repeated three times and representative images, processed at the same magnification, are shown. (E) Bacterial strains indicated at the bottom were grown in LB for 16 h and cell-free culture supernatants were subject to the iron sequestration assay. The iron sequestration activities of indicated species were normalized with that of PAO1 (black bar). SA, Staphylococcus aureus; BC, Bacillus cereus; EC, Escherichia coli; VC, Vibrio cholerae; ST, Salmonella enterica serovar Typhimurium; LM, Listeria monocytogenes. Three independent experiments were performed, and mean values ± SD (error bars) are displayed in each bar. *p < 0.05 (students’ t-test) vs. all other strains but ΔPA4834/pJN105::PA4834.