Acinetobacter baumannii is a significant pathogen of bloodstream infections in hospital patients that frequently causes single clone outbreaks. We aimed to evaluate the genetic relatedness and antimicrobial susceptibility of Acinetobacter spp. bloodstream isolates, in order to obtain insight into their cross-transmission. This prospective study was conducted at the Erciyes University Hospital. During a 1-y period, all patients with nosocomial BSI caused by Acinetobacter spp. were included in the study. All data with regard to the patients, underlying diseases and risk factors for BSI and the severity of disease were collected. Blood culture isolates of Acinetobacter spp. were identified according to their morphology and biochemical reactions. The antimicrobial susceptibility was determined using the Kirby-Bauer disk diffusion test according to the NCCLS; the genetic relatedness of isolates was determined by RAPD-PCR analysis and pulsed-field gel electrophoresis (PFGE). 41 patients acquired a nosocomial bloodstream infection caused by A. baumanii during this period. 88% of these infections ( 36 of 41) occurred while the patients were treated in the intensive care unit. Nearly 80% of the isolates belonged to 3 genotypes, suggesting cross-transmission in ICU settings where infection control practices are poor. All Acinetobacter isolates were multidrug-resistant and the crude mortality of patients infected with A. baumanii was 80.5%. We concluded that the genetic relatedness of Acinetobacter spp. causing BSI was very high, indicating cross-transmission within the ICU setting. Essential components of an infection control programme to prevent nosocomial transmission of A. baumannii are early detection of colonized patients, followed by strict attention to standard precautions and contact isolation.