Fig. 5

Intracellular pH measurement in WT and mutant L. reuteri strains during histamine production. Cells were grown for 24 h in MRS prior to washing and loading with pHrodo Red pH sensitive dye. a Representative images from an intracellular pH standard curve using pHrodo Red. Native stained cells exhibit slight intercellular variation (left panel), but intracellular pH can be equilibrated to the extracellular pH using ionophores (right panels). Image scale bar indicates 2 µm. Cells were grown for 24 h in MRS prior to washing and loading with pHrodo Red pH sensitive dye. For quantitative measures, fluorescence of cells was monitored in a plate reader at pH 7.3, pH 4.9 + d-histidine, and pH 4.9 + l-histidine. Graphs above depict fluorescence traces of the wild type strain compared to a HdcA and HdcP knockout strains, b EriC mutant strains, c EriC2 mutant strains, and d double EriC/EriC2 mutant strains. Error bars represent ± SEM. Indicated P-values are from a two-way repeated measures ANOVA with Dunnett’s multiple comparison test (*P < 0.05, **P < 0.01, ***P < 0.001, n.s. not significant)