This study analysed 330 Clostridium difficile strains isolated from patients with C. difficile infection who were hospitalized in two university hospitals (H1 and H2) in Warsaw, Poland, over the period 2004-2006. Strains were investigated for the presence of tcdA (A), tcdB (B) and binary toxin (CDT) genes, and antimicrobial susceptibility was determined against nine agents. Among the 330 C. difficile isolates, 150 (45.4 %) were classified as A(+)B(+)CDT(-), 18 (5.5 %) as A(+)B(+)CDT(+), 144 (43.6 %) as A(-)B(+)CDT(-) and 18 (5.5 %) as A(-)B(-)CDT(-). The predominant PCR ribotype in hospitals H1 and H2 was type 017 and accounted for 48.3 and 40.0 %, respectively. Only one PCR ribotype 027 strain was found. The rates of resistance to erythromycin and clindamycin in hospitals H1 and H2 were 53.6 and 53.6 %, and 48.6 and 47.5 %, respectively, whereas resistance rates to the newer fluoroquinolones gatifloxacin and moxifloxacin were 38.5 and 38.5 % (H1) and 38.4 and 40.1 % (H2). Erythromycin resistance was frequently associated with resistance to clindamycin and newer fluoroquinolones in strains belonging to type 017. No metronidazole- and vancomycin-resistant isolates were found, although two C. difficile isolates had elevated MIC values of metronidazole (MIC range 1.0-1.5 mg l(-1)) and 15 strains revealed elevated MIC values for vancomycin (MIC range 1.5-2.0 mg l(-1)). In conclusion, an increase in non-027 CDT-producing C. difficile strains was observed in Poland, but C. difficile PCR ribotype 017 remains a major circulating type.