MICs were determined for 15 antimicrobial agents against 37 Mycoplasma putrefaciens isolates. The most effective antimicrobial drug classes were the fluoroquinolones, the tetracyclines, the lincosamide lincomycin, and the macrolides. The susceptibility profile of the isolates correlated with the geographic origin. This is the first report of decreased susceptibility to the macrolides, lincomycin, and the tetracyclines in M. putrefaciens strains.