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This study evaluated the effect of the antibiotics oxytetracycline (OTC) and florfenicol (FLO) on the operation of two EGSB (expanded granular sludge bed) reactors. The experiment was conducted for 210 d in reactor R1 and 245 d in reactor R2. The reactors were inoculated with granular sludge from a upflow anaerobic sludge blanket (UASB) reactor on a local dairy farm. The sludge had an average pellet size of 2.35 mm, good sedimentability and a high percentage of organic material. The antibiotic tolerance and the inhibitory action on the bacterial population were different for each antibiotic studied. The results showed a more severe inhibitory effect on microorganisms that were in contact with increases in loads of FLO than those that were in contact with increasing loads of OTC, a condition reflected in the chemical oxygen demand (COD) removal efficiency.