Retrospective Observational Cohort Study of Conjunctival Scraping Samples Analyzed by Light Microscopy and Scanning Electron Microscopy: Coinfections and Inflammation

The relationship between microbial presence and host inflammatory response on the ocular surface is poorly defined, and conventional culture and PCR often fail to identify the pathogen in therapy-refractory disease. This retrospective study analyzed 300 conjunctival scraping samples from 150 therapy-refractory patients referred between January 2022 and July 2026. Samples were examined by scanning electron microscopy (SEM) cytology for the morphotype-level identification of pathogens and coinfections, and by light-microscopy cytology for the presence of inflammatory cells. Yeasts morphologically consistent with Candida (135 detections, 45.0%) and Acanthamoeba (93 detections, 31.0%) predominated, followed by Mycoplasma and Pseudomonas; polymicrobial associations were frequent, the most common being Acanthamoeba–Candida. Cytological inflammation was present in 73–77% of the samples positive for each main pathogen, whereas in 53 of 300 samples (17.7%) a pathogen was detected without cytological inflammation. In this cohort of therapy-refractory patients, SEM cytology revealed single and polymicrobial infections, including in the absence of inflammation. These findings support the microbiological investigation of the inflamed refractory patient and of apparently healthy patients before ocular surgery.