Rapid identification of bacteria by the pattern of redox reactions rate using 2′,7′-dichlorodihydrofluorescein diacetate

Abstract

Bacterial infection is a life-threatening situation, and its rapid diagnosis is essential for treatment. Apart from medical applications, rapid identification of bacteria is vital in the food industry or the public health system. There are various bacterial identification techniques, including molecular-based methods, immunological approaches, and biosensor-based procedures. The most commonly used methods are culture-based methods, which are time-consuming. The objective of this study is to find a fingerprint of bacteria to identify them.Three strains of bacteria were selected, and seven different concentrations of each bacterium were prepared. The bacteria were then treated with two different molar concentrations of the fluorescent fluorophore, dichlorodihydrofluorescein diacetate for 30 minutes. Then, using the fluorescence mode of a multimode reader, the fluorescence emission of each bacterium is scanned twice during 60 minutes. Plotting the difference between two scans versus the bacteria concentration results in a unique fluorescence pattern for each bacterium.Observation of the redox state of bacteria, during 90 minutes, results in a fluorescence pattern that is clearly a fingerprint of different bacteria. This pattern is independent of fluorophore concentration. Mean Squares Errors (MSE) between the fluorescence patterns of similar bacteria is less than that of different bacteria, which shows the method can properly identify the bacteria.In this study, a new label-free method is developed to detect and identify different species of bacteria by measuring the redox activity and using the fluorescence fluorophore, dichlorodihydrofluorescein diacetate. This robust and low-cost method can properly identify the bacteria, uses only one excitation and emission wavelength, and can be simply implemented with current multimode plate readers.