Background: This study aimed to investigate the bacterial colonization and antibiotic sensitivity profile of endotracheal tubes (ETTs) in mechanically ventilated patients, identifying key pathogens and resistance patterns to inform treatment strategies.Materials and Methods: A prospective observational study was conducted in a tertiary care hospital, enrolling 125 mechanically ventilated patients. Inclusion criteria were adult patients requiring mechanical ventilation for more than 48 hours with ETTs in place. Endotracheal tube samples were collected during extubation and processed using standard microbiological techniques for bacterial isolation and antibiotic sensitivity testing, following CLSI guidelines. Descriptive statistics and chi-square tests were used for data analysis.Results: Among the 125 patients, 60% were male, and the mean age was 55.20 years. Staphylococcus aureus (38.4%) and Pseudomonas aeruginosa (25.6%) were the most prevalent bacterial species identified. High resistance was observed against Ampicillin (60.8%), while Ciprofloxacin and Meropenem exhibited high sensitivity (87.2% and 93.6%, respectively). The colonization rate increased with the duration of mechanical ventilation, with 35.2% of patients ventilated for more than 96 hours showing colonization. Diabetes (50.4%) was the most prevalent comorbidity associated with increased colonization.Conclusion: This study highlights the importance of age, duration of mechanical ventilation, and comorbidities in bacterial colonization and infection risk. Prolonged mechanical ventilation and conditions like diabetes significantly increase colonization rates, emphasizing the need for effective infection control and targeted antibiotic therapies in critically ill patients. High resistance rates to commonly used antibiotics underscore the importance of careful antibiotic stewardship.