PURPOSE: To review bacterial culture results in infectious keratitis at a tertiary referral center. DESIGN: Retrospective case series. METHODS: Analysis of Gram stain, bacterial culture, and targeted antibiotic sensitivities of all cases of presumed infectious keratitis at Duke University Eye Center from 1997 to 2004 (n = 453). RESULTS: Cultures were positive in 307 cases (68%); 21% demonstrated polymicrobial growth. Among 388 isolates, 81% were Gram-positive. Gram stain results were available in 334 cases, demonstrating 18% sensitivity and 94% specificity. Antibiotic sensitivities revealed 24% resistance of tested S. aureus isolates to cefazolin, and <5% resistance of tested gram-negative isolates to various antibiotics, with no statistically significant increase from 1997 to 2004. CONCLUSIONS: In this study, the microbial profile has remained stable from 1997 to 2004. Gram stain studies were of poor sensitivity, while culture yields were significantly higher and demonstrated a relatively high polymicrobial infection rate. Increasing antibiotic resistance was not clearly demonstrated.