This study was to investigate antimicrobial resistance and bacterial distribution of gram negative bacilli isolated from intensive care unit (ICU) in the First Bethune Hospital in 2009. Disk diffusion test was used to study the antimicrobial resistance. The data were analyzed by WHONET 5 software according to Clinical and Laboratory Standards Institute (CLSI). Most of 593 gram negative bacilli isolated were isolated from sputum (39.3%), urine (23.1%) and blood (9.8%). Escherichia coli, Klebsiella.spp, pseudomonas aeruginosa, Acinetobacter baumannii were the most common strains among the isolates. The rates of extended-spectrum β-lactamases (ESBLs) in Escherichia coli and Klebsiella.spp were 53.9% and 47.9%, respectively. Carbapenems, cefepime, ceftazidime piperacillin/tazobactam, cefoperazone/sulbactam and amikacin were showed excellent activity. Bacterial distribution and the change of drug resistance should be investigated in order to guide rational drug usage in the clinic and prevent bacterial strains from being transmitted.