Real-time, label-free detection and identification of bacteria through non-invasive optical imaging

Abstract

Currently, traditional and newer molecular and mass spectrometry techniques of identifying bacteria from biological samples requires lengthy sample preparation, growth and labelling/staining assays. Thus, there is a pressing clinical need for an adjunct method that accurately identifies bacteria in real time. Here we report on the evaluation of confocal microscopy for the identification of clinically important and multi-drug resistant (MDR) bacteria in real time, using their intrinsic fluorescence features, i.e., emission spectra and fluorescence lifetime. The results demonstrate that difference in emission spectra and fluorescence lifetimes can be used as a fingerprint for identification of 12 bacterial species and MDR strains in real-time. Photostability or time-traces of bacteria demonstrated that these parameters could be used for tracking and recording without a need for labelling. Further, dilution experiments demonstrated that using intrinsic fluorescence S. aureus, Klebsiella pneumoniae and Escherichia coli bacteria can be detected and identified at clinically relevant concentrations as low as 2 × 102 CFU/mL. This non-invasive, non-labelling optical methodology may serve as the basis for development of a device that would quickly and accurately identify bacteria in biological samples. Thus, this intrinsic fluorescence technique would provide clinicians information, within minutes from sampling, to base accurate and specific treatments for patients.