C.difficile is a gram positive, toxigenic and anaerobic bacterium that cause diarrhea and psudomembrane colitis. The am of this study was rapid detection of toxigenic C.difficile in children with cancer that received chemotherapy.Fecal specimens obtained from children with cancer that undergoing antineoplastic chemotherapy. After collection of specimens by swab, primary isolation and extracted DNA was performed and genes of tpi , toxins A and B were detected by Multiplex PCR.In this study, incidence of C.difficille in children with cancer that undergoing chemotherapy, were 18 out of 105 patients. 12 out of 18 cases were toxigenic and 6 out of 18 were non- toxigenic . 3 out of 12 cases had both toxin A ,B and 4 out of 12 cases had only toxin B and 2 out of 12 had only toxin A.