The genus Acinetobacter is one of the main fears of hospital-acquired infection (HAI) wide-reaching mostly in ICUs. Objectives: This study aimed to identify the incidence of carbapenem-resistant Acinetobacter (CRA) HAI in Assiut University Hospitals ICU and determine their antimicrobial susceptibility profile. Methodology:The study included 1204 clinical samples from 928 patients with HAI and 625 environmental samples. Carbapenemase production was detected phenotypically by MIC determination using imipenem, meropenem and doripenem E-test, modified Hodge test, combined disc test and double disc synergy test (DDST). OXA genes (bla OXA-51 , bla OXA-23 , bla OXA-24 , and bla OXA-58 ) and MβL genes (bla SIM-1 , bla VIM , and bla IMP-1 ) were investigated genotypically by PCR. Results: Out of total 60 Acinetobacter isolates, 55 (91.7%) were extensively drug-resistant (XDR) and 5 (8.3%) were multi-drug resistant (MDR). 93.3% of Acinetobacter isolates were resistant to imipenem and meropenem, and 90% were resistant to doripenem. The prevalence of bla genes in isolates were bla OXA-51 (93.3%), bla , bla , bla OXA-58 (10%), bla SIM-1 (46.7%), and bla VIM (8.3%). bla IMP-1 was not detected in any isolate. Conclusion: CRA infections were considerable in our ICUs. The high prevalence of XDR Acinetobacter spp. observed in our study was very alarming and requires critical intervention. METHODOLOGY Specimen collection:This study was conducted on 1204 clinical samples obtained from 928 patients with HAI who were hospitalized in different ICUs of Assiut University Hospitals. Samples included endotracheal aspirates (n= 546), sputum (n= 218), blood (n= 217), urine (n= 161), throat swabs (n= 35), wound swabs (n= 24), ear swab