Acinetobacter spp. is a known nosocomial pathogen causing a wide range of clinical diseases mainly pneumonia, wound infections and blood stream infections (BSI). A cross sectional descriptive study was performed to determine the prevalence of Acinetobacter infection in Hospital Universiti Sains Malaysia, Kelantan (HUSM). The risk factors of Acinetobacter BSI were determined by 1:1 case control analytical study, involving fifty-eight confirmed cases of Acinetobacter BSI patients compared to the cases caused by Gram-negative bacteria. The prevalence of Acinetobacter BSI in the HUSM was 6.11% (95% CI 4.88-7.53%). The attack rate of Acinetobacter BSI was 2.77 episodes per 1000 hospital admissions. Acinetobacter BSI patients were mostly located in intensive care unit and had a longer intensive care unit stay. In univariate analysis, the risk factors for Acinetobacter BSI include prior exposure to antimicrobial agents such as penicillins, aminoglycosides and cephalosporins, mechanical ventilation, presence of nasogastric tube, arterial catheter and urinary catheter. In multivariate analysis, the independent risk factors for Acinetobacter BSI were prior treatment with cephalosporins (OR 3.836 95% CI 1.657-8.881 p=0.002) and mechanical ventilation (OR 3.164 95% CI 1.353-7.397 p=0.008). This study revealed that rational use of antimicrobial agents is of paramount importance to control Acinetobacter BSI.