FIG 5.

E. faecalis-mediated increased LEE expression requires cell contact. A schematic of the experiment performed is shown. EHEC was cultured alone or with ΔgelE E. faecalis for 16 h. Samples from these initial cultures were included on the final dot blot, designated original cultures, and also filtered to generate preconditioned (PC) medium for subsequent EHEC treatment. EHEC was then grown alone, in direct coculture with E. faecalis, or in the presence of preconditioned medium from EHEC alone (negative control) or EHEC and E. faecalis coculture for 6 h. Three different concentrations of preconditioned medium were tested (30%, 50%, and 70%). Sterile DMEM was mixed with preconditioned medium in the correct ratios to allow for subtraction of EspB signal originating from the preconditioned medium itself. A dot blot assay for EspB was performed, spot intensity from three replicate spots was quantified, and the average and standard deviation from one experiment are plotted. The experiment was repeated three times to ensure reproducibility.