Clostridium difficile is a gram-positive, spore-forming anaerobic bacterium. C.difficile is the leading cause of antibiotic associated diarrhea and colitis. The clinical spectrum of C.difficile infection (CDI) is highly variable, ranging from mild diarrhea to severe forms of intestinal illness including toxic megacolon, ileus, bowel perforation, and pseudomembranous colitis. Advanced age, long duration of hospitalization, and exposure to certain antimicrobial agents are the most common risk factors for CDI. The main virulence determinants of C.difficile are toxin A (enterotoxin) and toxin B (cytotoxin). Diagnosis of CDI is based on the identification of C.difficile toxin A or B in diarrheal stool. Various laboratory tests have been currently developed for the detection of C.difficile or its toxins in stool samples. The cell culture cytotoxicity assay and toxigenic culture have been regarded as the reference standard methods for the detection of C.difficile toxins. However, both of the reference methods are laborious, time consuming, and need expert personnel. Therefore, many microbiology laboratories use enzyme immunoassays, glutamate dehydrogenase antigen tests and real-time polymerase chain reaction (PCR) which are more rapid and practical. First-line antibiotics for CDI treatment are metronidazole and vancomycin. In this review, epidemiology, clinical spectrum, risk factors, pathogenesis, diagnostic methods and treatment of CDI have been summarized.