To determine the distribution and resistance profiles of Gram-negative bacteria (GNB) in intensive care units (ICUs) at King Abdullah Hospital in Bisha, Saudi Arabia. Methods: A record based retrospective study was conducted from December 2016 to January 2018. In total, 3736 non-duplicate clinical specimens from the general intensive care unit (ICU), neonatal ICU (NICU), and coronary CU (CCU) were analyzed for pathogens. Results: Of 3736 specimens, 9.6% (358) were positive for pathogens, and GNB constituted the majority (290/358; 81%). Acinetobacter is predominant in the general ICU, whereas Klebsiella pneumoniae is common in the NICU and CCU. Overall, GNB revealed a high resistance rate for cefuroxime (75.8%) trimethoprim/sulfamethoxazole (73.4%), cefotaxime (72.9%), aztreonam (64.6%), piperacillin (62.1%), and ciprofloxacin (61.5%). Acinetobacter revealed a high resistance (93.4% to 97.5%) to all antimicrobials except colistin (4%). Klebsiella pneumoniae showed a high resistance to trimethoprim/sulfamethoxazole (71.8%), cefotaxime (71.4%) and aztreonam (65.2%). Pseudomonas aeruginosa showed good activity for aminoglycosides but increasing resistance for cephalosporins and meropenem. GNB exhibited a high rate of multi-drug-resistant (MDR) phenotypes (67.9%) with a higher level among Acinetobacter spp. (97.5%). There were no significant differences in the resistance rates of GNB from different ICUs except for imipenem (p=0.002) and ciprofloxacin (p=0.003).  Conclusions: Increased antimicrobial resistance with high proportions of MDR patterns were found among GNB from ICUs. Comprehensive surveillance programs are needed to track the origins and emergence pathways of resistant pathogens.