Confocal autofluorescence identification of bacteria, fungi, and acanthamoeba in infected porcine cornea models

Current standard in the clinical diagnostic of corneal infection in vivo is reflectance confocal microscopy using photons in the wavelength range of 600 s nm. While pathogens with large spatial features (∼10 μm) such as acanthamoeba and fungal species can be identified, smaller pathogens such as bacteria (∼1 μm) are difficult to detect. Furthermore, reflected light produces images anywhere there are index of refractive index changes. The non-specificity of such images can contribute to confusion in pathogen identification. In this work, we used 488 nm to excite autofluorescence in nine common pathogens (4 bacteria, 3 fungi, and 2 acanthamoeba species) inoculated in ex vivo porcine corneas. We found that the detected green autofluroescence is sufficient to clearly image the pathogens. Histograms of the autofluorescence intensity can also act as a metric in pathogen classification.