The prevalence of Clostridium difficile and its toxins (A and B) in HIV-positive children in Poland was investigated in a group of 18 children, aged 6 months to 8 1/2 years. Stool samples were tested using an antigen detection method for toxin A/B, cytotoxicity-neutralization and culture. In 3 cases (17%) C. difficile toxins were detected in both stool samples and strains recovered from culture. The three strains isolated were shown by PCR methods to contain toxins A and B genes. All children had been treated previously with antimicrobial and antiviral agents. All three C. difficile-positive children had mild diarrhea that resolved without specific therapy. Further studies involving a large number of children and molecular analyses of isolated C. difficile strains are necessary to determine the frequency and rate of carriage of C. difficile strains among HIV-positive children in Poland.