Table 1.
Antimicrobial susceptibility (Minimal Inhibitory Concentration (MIC)) with B. subtilis strains.
Figure 1.
Effect of ndoA deficiency on bacterial survival after antimicrobial treatment.
Wild-type strain (BD630, filled circles) and its ΔndoA mutant (3169, open circles) were treated with the indicated concentrations of kanamycin (Kan) for 1 hr (panel A), 10-times MIC (4 µg/ml) kanamycin for the indicated times (panel B), the indicated concentrations of moxifloxacin (Mox) for 2 hr (panel C), and 16-times MIC (0.4 µg/ml) moxifloxacin for the indicated times (panel D). Results with a complemented ΔndoA mutant strain (3322) containing a single copy of the intact ndoAI-ndoA operon inserted at the chromosomal amyE locus are shown in panels A and C (open squares). Percent survival was determined as in Methods. Error bars indicate standard deviation; similar results were obtained in replicate experiments.
Figure 2.
Effect of ndoA deficiency on bacterial survival after treatment with hydrogen peroxide.
Wild-type strain (BD630, filled circles) and its ΔndoA mutant (3169, open circles) were treated with the indicated concentrations of hydrogen peroxide for 15 min (panel A) or for the indicated times at 250 mM (panel B). Error bars indicate standard deviation; similar results were obtained in replicate experiments.
Figure 3.
Effect of ndoA deficiency on bacterial survival after treatment with UV irradiation.
Overnight cultures of B. subtilis were diluted with LB medium, 10-µl aliquots were spotted on LB agar plates, and spots were allowed to dry. The plates were exposed to UV irradiation (254 nm) at various intensities for the indicated times, and they were incubated overnight at 37°C. Wild-type strain (BD630, filled circles) and its ΔndoA mutant (3169, open circles) received low (0.14 mW/cm2, panel A), high (0.4 mW/cm2, panel B), or moderate exposure (0.2 mW/cm2, panel C). Error bars indicate standard deviation; similar results were obtained in replicate experiments.
Figure 4.
Effect of ndoA deficiency on bacterial survival during high-temperature treatment.
Overnight cultures of wild-type B. subtilis strain (BD630, filled circles) and its ΔndoA mutant (3169, open circles) were concentrated by centrifugation, washed in saline (0.9% NaCl), resuspended and serially diluted in saline, and incubated at the indicated temperatures for 20 min (panel A) or for the indicated times at 52°C (panel B). After incubation, the cultures were diluted and applied to LB agar plates for colony determination. Error bars indicate standard deviation. Similar results were obtained in replicate experiments.
Figure 5.
Effect of ndoA deficiency on bacterial sporulation and survival.
B. subtilis wild-type strain (BD630, circles) and its ΔndoA mutant (3169, squares) were incubated in Difco sporulation medium. At the indicated times, aliquots were removed and incubated at 90°C for 20 min to kill vegetative cells. Total bacterial cell (before heat, open symbols) and spore numbers (after heat, filled symbols) were plotted as a function of time. Dotted lines indicate extrapolation to detection limit (100 cfu/ml, dashed line). Error bars indicate standard deviation; similar results were obtained in replicate experiments.
Figure 6.
Effect of ndoA deficiency on peroxide accumulation following kanamycin treatment.
Wild-type B. subtilis strain BD630 (filled circles) and its ΔndoA mutant 3169 (open circles) were grown overnight, diluted 50-fold in LB medium, and treated with kanamycin (4 µg/ml). At the indicated times aliquots were removed for determination of hydrogen peroxide as in Methods. Error bars indicate standard deviation; Similar results were obtained in replicate experiments.