Tehran University of Medical Sciences Iranian Journal of Microbiology 2008-3289 17 5 2025 10 14 725 733 Aya Maytah Department of Basic Sciences, Ma′an University College, Al-Balqa Applied University, Ma′an, Jordan Omar AlKofahi Department of Medical Laboratory Science, Faculty of Allied Medical Sciences, Zarqa University, Zarqa, Jordan Rania Al-Groom Department of Medical Laboratory Science, Faculty of Allied Medical Sciences, Zarqa University, Zarqa, Jordan; Department of Allied Medical Sciences, Zarqa University College, Al-Balqa Applied University, Zarqa, Jordan Mohd Sajjad Ahmad Khan Department of Basic Sciences, Imam Abdulrahman Bin Faisal University, Dammam, Saudi Arabia Basem Fouad Dababneh Department of Medical Laboratory Sciences, Faculty of Allied Medical Sciences, Al-Ahliyya Amman University, Amman, Jordan Anas Da'meh Department of Medical Laboratory Science, Faculty of Allied Medical Sciences, Zarqa University, Zarqa, Jordan Rahaf Alsarayereh Department of Medical Laboratory Science, Faculty of Allied Medical Sciences, Zarqa University, Zarqa, Jordan Fuad Alhawarat Department of Applied Medical Sciences, Al Hussein Bin Abdullah II Academy for Civil Protection, Al-Balqa Applied University, Salt, Jordan Heba Ahmad Al Shqairat Precision Medical Lab (PMLAB), AL-Karak, Jordan 2025 07 08 2025 08 22 Background and Objectives: Acinetobacter baumannii (A. baumannii) is an opportunistic bacterial pathogen principally related with hospital-acquired infections. This study aimed to isolate and identify A. baumannii strains, investigate their resistance to various antibiotics, and characterize A. baumannii at the molecular level. Materials and Methods: A total of 100 samples were obtained from various hospital departments, including the intensive care unit (ICU), emergency room, kidney dialysis and surgery units. The incidence of drug resistance was studied using the Vitek 2 Compact system and further using molecular techniques such as polymerase chain reaction to analyze the genes responsible for resistance. Results: The study exhibited a high prevalence of multidrug-resistant (MDR) A. baumannii isolates, especially in ICU patients. The males were the predominant group, accounting for 60% whereas females were 40%. The most frequent samples were from urine (43%) and skin (24%). Majority of samples were from the ICU (42%) and emergency departments (20%). The tested isolates exhibited the highest resistance (66%) to oxacillin, whereas the maximum sensitivity (52%) was recorded for Erythromycin. Molecular analysis revealed the occurrence of resistance genes blaOXA-23, blaOXA-24, blaOXA-51, and blaOXA-58, which contribute to carbapenem resistance. Conclusion: The findings emphasize that A. baumannii remains a formidable nosocomial pathogen, and there is pressing requirement for enhanced infection control procedures and antibiotic stewardship. Through improved molecular observation, judicious use of antibiotics and improved infection control practices, healthcare providers can alleviate the impact of MDR A. baumannii infections and improve the prognosis for affected patients in Jordan and beyond. https://ijm.tums.ac.ir/index.php/ijm/article/view/5606 https://ijm.tums.ac.ir/index.php/ijm/article/download/5606/1818