Acridine orange as an indicator of bacterial susceptibility to gentamicin

We have studied the response of Escherichia coli NCTC10418 to gentamicin with flow cytometry. The susceptibility of individual bacterial cells to the antibiotic was assessed by differential staining using the metachromatic dye, acridine orange. Exponential phase cultures were exposed to the minimum bactericidal concentration of gentamicin and analysed at regular intervals over 90 min. Within 60 min of exposure to the drug, two sub-populations of organisms could be distinguished in cultures by their different acridine orange-associated fluorescence emissions of < 550 nm and > 550 nm. The number of bacteria exhibiting acridine orange-associated fluorescence at > 550 nm corresponded to counts of colony forming units.