The highest incidence fungal and protozoan eye infections in the United States occur in South Florida. Fungal and protozoan eye infections present clinicians with a considerable diagnostic and therapeutic challenge. The initial clinical presentation resembles a microbial keratoconjunctivitis, which is presumed to be of a bacterial etiology. It is only after antibiotic therapy has failed to resolve the condition that the notion of fungal or protozoan involvement is considered. The incidence of fungal eye infections has increased in recent decades. Most fungal eye infections occur secondary to corneal trauma. The therapeutic challenge is to rapidly diagnose the presence of a fungal or protozoan pathogen because ocular morbidity is reduced by early initiation of antifungal or antiprotozoan treatment. We plan to utilize and test a high throughput, rapid diagnostic method (Luminex xMAP) to assess fungal and protozoan diversity in the healthy ocular tear film and in infected ocular tissues by focusing on yeast, filamentous fungal and protozoan populations. Combining efforts to identify microbial populations with ongoing studies funded by NIH (Drs. Fell and Diaz) presents a unique opportunity to create a rapid method to detect and identify infecting fungal and protozoan organisms in ocular tissues to reduce ocular morbidity and mortality. The initial emphasis of the project will be to sample, isolate and grow fungal and protozoan pathogens from ocular tissues. DNA samples of each isolate will be obtained, amplified, sequenced and the species will be identified via a blast search (Univ. Florida). Molecular probes for rapid detection and identification will then be developed for the more common microbial isolates. The epidemiology of fungal and protozoan ocular disease in South Florida will be assessed as a component of this project.