Objectives: The aim of this study is to identify pathogenic microorganisms in tracheal aspiration samples and examine their sensitivity to antibiotics. Furthermore, the aim is to compare therapy administered based on results of tracheal aspirate cultures to adequate empirical treatment strategies. Study design: cross-sectional study Materials and Methods: 209 patients admitted to the ICU from October 2016 to September 2017 were included in the study. Disease histories, therapeutic list and in total 376 tracheal aspirates analyzed. Pathogenic microorganisms isolated in tracheal aspirates were identified and antibiotic sensitivity of the most commonly isolated bacteria was described. Antibiotic therapy on the day of tracheal aspirate sampling was compared to the therapy administered when results of tracheal aspirate cultures came in. Statistical analysis was performed using t-test, χ2 test, Fisher's exact test and Pearson's correlation. Results: Arithmetic mean age of patients was 64.6 years. The most common admission diagnosis was acute respiratory failure. The most frequent pathogenic microorganisms isolated in tracheal aspirates were gram negative bacteria, and the most frequently isolated one was multiresistant Acinetobacter baumanii. In 32% of cases antibiotic therapy on the day of tracheal aspirate sampling was not adequate and there was a change of antibiotics. On the day of tracheal aspirate sampling the most common antibiotics used in patients’ therapy were metronidazole, amoxicillin + clavulanic acid and gentamicin, while most frequent antibiotics administered on the day tracheal aspirate cultures results came in were metronidazole, amoxicillin + clavulanic acid and colistin. The increase in the use of colistin after the tracheal aspirates results came in is statistically significant. Conclusion: Multiresistant gram negative bacteria are most commonly isolated microorganisms in tracheal aspirate in patients admitted to the ICU, with a high share of Acinetobacter baumanii, which is sensitive to colistin therapy.