Cefquinome is the only fourth-generation cephalosporin used solely for veterinary applications.  In this study, we established the wild-type cut-off (CO_WT) and pharmacokinetic/pharmacodynamic cut-off (CO_PD) of cefquinome against Escherichia coli and Staphylococcus aureus.  A total of 210 E. coli and 160 S. aureus isolates were collected from pigs in Guangdong Province between 2014 and 2018.  The minimum inhibitory concentrations (MICs) were determined using a microdilution broth method.  MIC₅₀ and MIC₉₀ were 0.06 and 0.25 μg mL^–1 for E. coli and 0.5 and 1 μg mL^–1 for S. aureus, respectively.  Statistical analysis and the ECOFFinder Program showed that the COWT for cefquinome against E. coli and S. aureus were 0.125 and 2 µg mL^–1, respectively.  The resistance rates were 11.9% for E. coli and 6.25% for S. aureus.  Based on a 5 000-subject Monte Carlo simulation, the CO_PD value for cefquinome against E. coil and S. aureus was 0.25 µg mL–¹ under the recommended dose (2 mg kg^–1, twice a day for 3 days), confirming that infections caused by strains with MIC≤0.25 μg mL^–1 could be effectively treated.  Following adjustment of the dosing regimen to 4.5 mg kg^–1, effective treatment (>90) was achieved for S. aureus infections with MIC₉₀ 1 μg mL^–1.  This susceptibility breakpoint determination is significant for resistant surveillance and cefquinome dosage guidance against E. coli and S. aureus in pigs.