The antimicrobial susceptibilities of 49 Clostridium difficile isolates obtained from patients with C. difficile-associated diarrhea to metronidazole, vancomycin, rifampicin, fusidic acid, doxycycline, and linezolid were determined by the disc diffusion and Etest (Biodisk, Solna, Sweden). Random amplification of polymorphic DNA-PCR amplification assay was performed for studying clonality of isolates. Resistance to metronidazole was found in 2% (1/49 isolates; MIC > or = 256 microg/mL) of isolates and resistance to linezolid in 2% (1/49 isolates; MIC = 24 microg/mL). One isolate showed combined resistance to fusidic acid (by disc diffusion test) and rifampicin (MIC > or = 32 microg/mL). All isolates were sensitive to doxycycline and vancomycin. Molecular typing revealed an absence of clonality among the resistant isolates, whereas the sensitive isolates were monoclonal. Resistance of C. difficile to metronidazole and other antimicrobials including linezolid exists in our institution. This finding should promote exploration of this problem in Israel and clarify the impact of resistance on outcome.