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Yinhong Qu, Karin E. Thomas, Alexander K. Schuster, Yi-Kai Wu, Tracy L. Purcell, II, Josef F. Bille, David J. Schanzlin; Identification of Microorganisms Utilizing the Two-Photon Ophthalmoscope. Invest. Ophthalmol. Vis. Sci. 2011;52(14):5823.
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To identify different microorganisms that cause infectious keratitis by using a prototype of two-photon ophthalmoscope. Previously, we found several microorganisms that are difficult to identify because of similarities in shape and size. Various laser power strengths for acquiring autofluorescence signals from different microorganisms can be used for narrowing diagnosis. We believe that this method will help ophthalmologists more quickly and precisely identify potentially dangerous microorganisms of the cornea.
The device is a nonlinear scanning laser ophthalmoscope which includes a two-photon excited fluorescence/second harmonic generation imaging modality. The infected donor corneal buttons were viewed under the two-photon ophthalmoscope with different laser sources (10mW/20mW/50mW). Infections that we have studied include Pseudomonas, Acanthamoeba and Microsporidium. We have used laser power strength, intensity of autofluorescence and morphological characteristics to identify these microorganisms.
All three microorganisms in our study, Pseudomonas, Acanthamoeba and Microsporidium, have been successfully viewed under the two-photon ophthalmoscope by using different laser powers. Autofluorescence signal from Pseudomonas can be acquired by using 25mW-30mW laser source while Acanthamoeba‘s autofluorescence could be viewed by using 15mW laser source. Microsporidium has a stronger autofluorescence so it may be visualized under 10mW. Due to the lack of fluorophors in most corneal structures, the autofluorescence of donor corneal buttons does not interfere with the autofluorescence imaging of microorganisms we studied.
Quick identification of corneal infection is crucial to saving vision. Pseudomonas, Acanthamoeba, and Microsporidium cause devastating loss of vision, which often result in full thickness corneal transplant. In this study, we found different microorganisms generate a range of autofluorescence signals, which can be viewed at various laser powers. Therefore, we could use this information to better identify different microorganisms. We have had initial success in identifying Pseudomonas, Acanthamoeba and Microsporidium by using this method. With greater laser power strength and better imaging resolution, we believe we will be able to identify a large variety of microorganisms in the future.
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