PURPOSE. To analyze the microbiologic spectrum and patterns of resistance of infectious keratitis in patients treated at a tertiary hospital in South Australia. METHODS. Retrospective review of microbiology laboratory records of all patients with infectious keratitis who had corneal scrapings, from 1998 to 2003. All records were subsequently reviewed for Gram staining and culture results, as well as antibiotic sensitivity and resistance. RESULTS. Positive corneal cultures were obtained in 134 out of 211 patients who had corneal scrapings (63.5%). Coagulase negative Staphylococcus was the commonest pathogen identified (29.8% of positive cultures), followed by Staphylococcus aureus (18.7%), Pseudomonas aeruginosa (12.7%), Moraxella (6.7%), Streptococcus pneumonia (6.0%), and fungal keratitis (5.2%). In 43.3% of culture positive cases, the organisms were also identified in Gram stain, and in all these cases there was a full correlation between the two methods. In vitro sensitivities were highest for gentamicin. Fourteen cases (35%) of coagulase negative Staphylococcus were found to be resistant to cephazolin. No ciprofloxacin resistance was identified in all Pseudomonas isolates tested. CONCLUSIONS. Staphylococcus species continue to be the commonest causative organism for infectious keratitis; however, there is an emerging resistance to cephazolin, which is commonly used as the first-line antibiotic for Gram-positive cocci.