We determined MICs of antibacterial agents against 1145 clinical strains of aerobic Gram-negative bacteria (22 species) isolated at 16 Japanese facilities in 2008. MICs were determined using mostly broth microdilution method and antibacterial activity was assessed. Strains producing extended-spectrum beta-lactamases (ESBL) accounted for 3.8% of Escherichia coli, 2.6% of Klebsiella pneumoniae, 6.8% of Klebsiella oxytoca, 5.5% of Proteus mirabilis and 1.8% of Proteus vulgaris. ESBL produced strains were 6.8% at K. oxytoca that increased compared with 3.2% and 5.5% at P. mirabilis that decreased compared with 18.8% in 2006. Among Haemophilus influenzae, 61.7% that decreased compared with 67.7% in 2006, equaled 58.7% in 2004, were strains when classified by penicillin-binding protein 3 mutation. Against Pseudomonas aeruginosa, the activity of most antibacterial agents was similar to that in 2006. Although two antibacterial agents that tobramycin showed an MIC90 of 1 microg/mL and doripenem showed an MIC90 of 4 microg/mL against P. aeruginosa have potent activity. Of all P. aeruginosa strains, 4.3% were resistant to six agents of nine antipseudomonal agents, that decreased compared to 12.2% in 2004 and 5.7% in 2006. Against other glucose-non-fermentative Gram-negative rods, the activity of most antibacterial agents was similar to that in 2006.