Abstract Introduction Our objectives were to describe the incidence, clinical characteristics, and risk factors for Clostridium difficile infection (CDI) in critically ill patients and to determine C. difficile PCR-ribotypes. Methods Prospective, observational study in 26 Spanish ICUs. Patients with diarrhea meeting ESCMID criteria for CDI were included. Molecular characterization of isolates was performed using PCR ribotyping. Results Of 4258 patients admitted to the ICUs, 190 (4.5%) developed diarrhea. Only 16 patients (8.4%) were diagnosed with CDI. Ribotype 078/126 (25.0%) was the most frequently identified. The mortality rate was similar in patients with ICD compared to patients with diarrhea not caused by C. difficile ( p = 0.115). Chronic renal insufficiency was identified as the only factor independently associated with the development of CDI (OR 5.87, 95% CI 1.24–27.83; p = 0.026). Conclusions The incidence of CDI in Spanish ICUs is low. Only chronic renal insufficiency was observed to be a risk factor for CDI development.