Portable device for rapid assessment of pathogens using autofluorescence lifetime characteristics

Pathogens such as bacteria and fungi express various autofluorescence markers such as NAD(P)H, flavins, porphyrins etc. In contrast to steady state fluorescence, wherein the amplitude variations are more sensitive to bacterial growth state, time dependant fluorescence characteristics of these autofluorescence biomarkers are more robust and can potentially provide higher specificity of detection especially in various clinical and industrial settings. However, fluorescence lifetime measurements are typically carried out using bulky and expensive instruments. Herein, we demonstrate an extremely portable device 10 cm*10 cm*5 cm for fluorescence lifetime measurement comprising of pulsed LED sources, optical filters and single photon avalanche photodetectors (SPADs). The entire instrument is driven by an embedded hardware board and the photon arrival times from the SPAD are measured using a Texas instrument TDC counter interfaced with the hardware. First, the instrument response function of the device is measured and is found to be <1 ns suitable for measuring the fluorescence lifetime of various pathogens. Subsequently, fluorescence lifetime measurements of pathogens such as Staphylococcus Aureus, Pseudomonas Aeruginosa and E.coli is done under 365 nm light excitation. Using the autofluorecence lifetime characteristics and classification algorithms, we demonstrate bioburden detection in water with sensitivity exceeding 90% with a minimum detection limit of 10³ CFU/mL. Furthermore, a handheld version of the device is also developed for assessment of the pathogens directly on the clinical samples such as ulcers, burns, surgical site infections. Notably, the handheld device is able to track bacterial cell growth, differentiate cancer tissues from noncancerous samples and is also able to detect microbial growth in flesh samples demonstrating its applicability in a range of potential applications.