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. Author manuscript; available in PMC: 2021 Sep 9.
Published in final edited form as: Cell Host Microbe. 2020 Jun 10;28(3):411–421.e6. doi: 10.1016/j.chom.2020.05.015

Figure 4. HsmA reduces oxidative stress.

Figure 4.

(A) C. difficile (CD) WT and hsmA::CT strains were grown for 6 hours followed by treatment with or without heme (25 μM) for 30 min. Samples were exposed to atmospheric oxygen and oxidative stress generation was determined by measuring fluorescence of dihydrorhodamine 123 (DHR123; ex. 507 nm, em. 529). (B) Growth of S. aureus (SA) ΔΔsod pOS1 and pOS1_hsmA strains in the presence or absence of paraquat (2 mM) and dihydrorhodamine 123. (C) Oxidative stress generation was quantified by measuring fluorescence of DHR123 from SA ΔΔsod pOS1 (empty vector), SA ΔΔsod pOS1_hsmA, or SA ΔΔsod pOS1_hsmA_5His-Ala (point mutant defective for heme binding) in the presence or absence of paraquat. The data are a representative from three independent experiments with standard error of the mean. (D) S. aureus ΔΔsod strains carrying the empty pOS1 vector or pOS1_hsmA were co-incubated with neutrophils derived from murine bone marrow (MOI of 1) for 12 hours. S. aureus CFUs were enumerated and compared to untreated controls. Statistical significance was determined using the multiple comparison two-way ANOVA test with the Sidak correction for multiple comparisons comparing the means of each group to one another. * denotes p-value < 0.05. See also Figure S4 and Table S2.