The common pathogenic bacteria in the eye include Staphylococcus epidermidis, Staphylococcus aureus, Staphylococcus pneumoniae and Pseudomonas spp. These pathogenic bacteria cause keratitis and endophthalmitis that can often lead to blindness, so finding broad-spectrum, highly effective, low-toxicity antibiotics is particularly important for effective control of ocular infections. Fourth-generation fluoroquinolones have a bactericidal effect by inhibiting the DNA gyrase of Gram-negative bacteria and topoisomerase of Gram-positive bacteria to block the replication of bacterial DNA. The fourth-generation fluoroquinolones have overcome the inability of the first three generations to combat Gram-positive bacteria and have become broad-spectrum antibiotics that can effectively treat drug-resistant bacterial infections and slow the emergence of new drug-resistant strains. In this study, the antimicrobial activity of gatifloxacin and moxifloxacin against 14 pathogenic bacteria in the eye was evaluated by measuring their minimum inhibitory concentration (MIC) in vitro. METHODS: Fourteen pathogenic bacteria, including seven Gram-positive, four Gram-negative and three uncommon strains, were obtained from patients with suspected bacterial keratitis and endophthalmitis. The minimum inhibitory concentrations of gatifloxacin and moxifloxacin were observed by microassay for each strain according to NCCLS (National Committee for Clinical Laboratory Standard) standards. Results: 1. Gatifloxacin and moxifloxacin showed similar antibacterial activity (MIC values of 0.08 mg/ml to 0.57 mg/ml) against six Gram-positive bacteria (Staphylococcus epidermidis, Staphylococcus aureus, Staphylococcus pneumoniae, Streptococcus pyogenes, Bacillus cereus and Enterococcus); while for Streptococcus griseus, the MIC values were 0.22 mg/ml for gatifloxacin and 0.73 mg/ml for moxifloxacin. Among the Gram-negative bacteria, the MIC value of gatifloxacin against Pseudomonas aeruginosa was 1.28 mg/ml, which was significantly lower than that of moxifloxacin, which was 2.60 mg/ml; while the MIC values of gatifloxacin against Pneumocystis pneumoniae and Aeromonas aeruginosa were 1/4 to 1/5 of those of moxifloxacin. 3. Conclusion: The fourth-generation fluoroquinolones, especially gatifloxacin, have broad-spectrum anti-microbial activity, and it has efficient killing effect on common pathogenic bacteria in ophthalmology, which is stronger than moxifloxacin; for opportunistic pathogenic infections with strong resistance, such as post-LASIK mycobacterium keratitis, gatifloxacin has stronger bactericidal effect.