Figure 3. S. paratyphi B requires PmrD to express PmrA-dependent genes and to resist polymyxin B during growth in Fe³⁺.

(A) β-galactosidase activity (Miller units) produced from a pbgP-lac transcriptional fusion in the following strains: S. typhimurium (EG9241), S. paratyphi B (EG16652), S. paratyphi B (pmrA G211) (EG16275), S. paratyphi B ΔpmrD (EG16277), S. paratyphi B pmrA (G211) ΔpmrD (EG16276), and S. paratyphi B ΔpmrA (DC167). Bacteria were grown in N-minimal medium at pH 7.7 with low Mg²⁺, high Mg²⁺ or low Mg²⁺ and high Fe³⁺. Data correspond to the mean values of three independent experiments performed in duplicate, and error bars show standard deviation. Asterisks indicate statistically significant differences based on a two-tailed Student t-test (p<0.05). (B) β-galactosidase activity (Miller units) produced from a pbgP-lac transcriptional fusion in isogenic pmrD⁺ or pmrD⁻ S. typhimurium strains harboring either the pmrA (E211) or (G211) allele (EG9241, EG11775, EG14331 and DC300) were determined as described in (A). (C–D) Growth of S. enterica strains harboring either the pmrA (E211) or the pmrA (G211) allele in the presence of the antibiotic polymyxin B (2.5 µg/ml) on plates containing 10 µM Mg²⁺ and 100 µM Fe³⁺. S. paratyphi B growth was determined in the wild-type (SARA46), in a strain harboring either the pmrA (E211) (DC282) or pmrA (G211) (DC280) gene, in a ΔpmrD mutant expressing the pmrA (E211) (DC165 and DC287) or pmrA (G211) gene (DC285), and in a ΔpmrA mutant (DC167) (C). As a control, growth of S. typhimurium 14028s was monitored on the same plate (C). S. typhimurium growth was determined in the wild-type (14028s), in a strain harboring a 3′ FLAG-tagged S. typhimurium pmrD gene and the pmrA (E211) (EG16279) or pmrA (G211) (EG13404) allele, in a ΔpmrD mutant expressing the pmrA (E211) (DC46) or pmrA (G211) (EG14088) gene, and in a pmrA mutant (EG7139) (D).