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Background: One of the most common infections in neonatal period is ophthalmia neonatorum. In this study, the bacterial agents, drug resistance and susceptibility of bacterial agents were studied. Methods: In this study a total of 72 newborns with ophthalmia neonatorum admitted in Bahrami Hospital in Tehran during the years 2008-2011 were continuously enrolled in a case series, descriptive study. Demographic data, including age, sex, cause of admission and culture of discharge from the eyes and its antibiogram, as well as experimental treatments and treatment outcomes were collected. Results: Forty four infants (61.1%) were males and 28 (38.9%) were females and the mean age on admission was 11.6±7.7 days. In 51 patients (70.8%) the onset of ophthalmia neonatorum was prior to admission. More than 56% of cases with ophthalmia neonatorum were associated with sepsis. On the other hand, positive blood culture was detected in 15.3% of cases. Among 72 neonates with ophthalmia neonatorum, 26 (36.1%) had a positive culture of the eye discharge. The most common causes of bacterial agents were Staphylococcus aureus (46.1%) (12 of 26 cases). Other causes included streptococcus species (23%), Pseudomonas (15.3%), E-coli (11.5%) and Haemophilus influenza (3.8%). The most frequent causes of drug resistance were Ampicillin, Penici-llin, Cefixime, and Ceftazidime (100% resistance). The most sensitive antibiotics were vancomycin and imipenem (100% sensitivity). Based on the conventional treatment, clinical response to local gentamicin was approximately 60%. Sulfacetamide was associated with no clinical response in 40% of cases. Conclusion: The antibiogram and clinical response to empiric treatment showed that resistance to ampicillin and some third generation of cephalosporine was 100%. Aminoglycosides' sensitivity was more than 50% locally and systemically. Our recommendation is performing eye discharge culture before antibiotic treatment. More studies with numerous cases should be done for better definition of bacterial resistance. [ABSTRACT FROM AUTHOR]