FIG 5.

Bicarbonate selectively dissipates B. glumae PMF and reduces toxoflavin production and virulence without impacting growth. (A) Sodium bicarbonate (NaHCO₃) sensitivity of B. glumae strains. Serially log₁₀-diluted cells of B. glumae 336gr-1 and the ΔdbcA strain transformed with control vector (vec) and pSC301 (DbcA) were spotted and grown on MH2 agar medium containing 100 μg/ml trimethoprim and either 2.5, 5, or 7.5 mM sodium bicarbonate at 37°C for 48 h. (B) Growth curve of B. glumae 336gr-1 in LB broth buffered to pH 7.0 with 70 mM BTP with and without 5 mM NaHCO₃. (C) Measurement of membrane potential (ΔΨ). Fluorescence ratio in the graph represents the red (595 nm)/green (530) emission ratio of JC-1 dye. 5 mM NaHCO₃ was present when indicated. **, P < 0.01. (D) Production of toxoflavin by B. glumae 336gr-1 in growth medium containing sodium bicarbonate. LB broth medium was buffered with 70 mM BTP, pH 7.0. Equal numbers of bacterial cells (5 × 10⁷ CFU/ml) were transferred into 250-ml conical flasks containing 25 ml buffered LB broth and either 0, 5, or 7.5 mM NaHCO₃ and grown at 37°C with shaking for 30 h. (E) TLC analysis for toxoflavin produced by B. glumae 336gr-1 in growth medium containing indicated concentrations of NaHCO₃. (F) Onion scales were inoculated with B. glumae preincubated with or without 5 mM NaHCO₃, which were then incubated at 30°C for 6 days in a humid chamber. (G) Area of maceration (cm²) produced by indicated strains. **, P < 0.01; *, P < 0.05; NS, not significant.