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A cell-based infection assay identifies efflux pump modulators that reduce bacterial intracellular load

Fig 6

EPMs do not disrupt bacterial inner or outer membranes.

(A,B) Salmonella treated with DMSO or EPMs [100 μM] but not CCCP [1 mM] acquire TMRM staining within 30 minutes. (A) Representative data from one of three independent experiments. (B) Median fluorescence intensity from three experiments normalized to unstained control (0). (C) Disk diffusion assays; the radius of the zone of growth inhibition after 16 hours of exposure to compound across a dose range. Black lines, semilog fit for the combined antibiotic data; gray lines, semilog fit for CCCP and PAβN; dotted lines, limit of detection (disk radius). Average of two measurements from each image captured from one experiment representative of two independent experiments. (D,E) Nitrocefin access to the periplasm as monitored by nitrocefin [100 μM] hydrolysis in the presence of the indicated concentrations of compounds. (D) Absorbance 486 nm of bla+ Salmonella normalized to bla- Salmonella. Data is representative of at least two independent biological replicates. (E) Slope of the linear region of the A486 plot from at least three experiments. Data is normalized to A486/minute. * p < 0.05, *** p < 0.001, **** p < 0.0001 by one-way ANOVA with Dunnett’s post-test.

Fig 6