Aims To determine the accuracy of diagnosing microbial keratitis by masked medical and non-medical observers using the Heidelberg Retina Tomograph II/Rostock Cornea Module in vivo confocal microscope.

Methods Confocal images were selected for 62 eyes with culture- or biopsy-proven infections. The cases comprised 26 Acanthamoeba, 12 fungus, three Microsporidia, two Nocardia and 19 bacterial infections (controls). The reference standard for comparison was a positive tissue diagnosis. These images were assessed on two separate occasions by four observers who were masked to the tissue diagnosis. Diagnostic accuracy indices, κ statistic and percentage agreement values were calculated. The Spearman correlation coefficient (r_s) was calculated for the number of correct diagnoses versus duration of disease.

Results The highest sensitivity and specificity values were 55.8% and 84.2%, respectively, and the lowest sensitivity and specificity values were 27.9% and 42.1%, respectively. The highest positive and lowest negative likelihood ratios were 2.94 and 0.59, respectively. Agreement values were: fair to moderate (κ 0.22–0.44) for reference standard versus observer diagnosis, moderate to good in intraobserver variability (repeatability, κ 0.56–0.88) and poor to moderate in interobserver variability (reproducibility, κ 0.15–0.47). The correct diagnosis was associated with duration of disease for Acanthamoeba keratitis (r_s=0.60, p=0.001).

Conclusions The diagnostic accuracy of microbial keratitis by confocal microscopy is dependent on observer experience. Intraobserver repeatability was better than interobserver reproducibility. Difficulty in distinguishing host cells from pathogenic organisms limits the value of confocal microscopy as a stand-alone tool in diagnosing microbial keratitis.