Background and Objectives: Clostridium difficile infection (CDI) has become a significant healthcare-associated infection throughout the world and is particularly important in developing countries. This study aimed to investigate clinical characterization and risk factors related to toxigenic C. difficile infection in adult and pediatric patients, antimicrobial susceptibility pattern. Also, to evaluate different diagnostic methods for rapid detection of C. difficile associated diarrhea (CDAD) in Egypt. Materials and Methods: Stool samples were collected from 95 pediatric patients and 37 adult patients suffering from antibiotic associated diarrhea and were subjected to direct toxin immunoassay and culture on cycloserine/cefoxitin/fructose agar. The presence of tcdA and tcdB genes was tested by PCR. Results: Toxigenic C. difficile was isolated from pediatric and adult patients at a rate of 17.89% (17/95) and 27% (10/37) respectively. The sensitivity and specificity of direct PCR from stool are (100%, 100% and 82.4%, 100%) in adult and pediatric samples respectively. The susceptibility of C. difficile to vancomycin and metronidazole were found to be 66.7% and 48.2% respectively. Conclusion: Diabetes mellitus, prior antibiotic treatment, hematological malignancy on chemotherapy, malnutrition, neutropenia and Ryle feeding are risk factors for development of CDAD. Tight restriction of unnecessary antibiotic uses is necessary in our locality. Direct detection of toxin genes in stool by PCR is sensitive and specific method for early detection of C. difficile.