Three seropositive pediatric recipients of allogeneic stem cell transplantation out of a group of 42 patients receiving T-cell-depleted, unrelated transplants and 37 patients receiving T-cell-depleted, haploidentical transplants were monitored longitudinally for human cytomegalovirus (HCMV) infection and the emergence of antiviral drug resistance. Early in the posttransplant course, all 3 patients developed HCMV mutations conferring drug resistance to ganciclovir. One child additionally developed multidrug resistance to foscarnet and cidofovir, with mutations in the viral phosphotransferase gene (UL97) and the DNA-polymerase gene (UL54) being found. These data show that resistant HCMV infection does not necessarily correlate with a severe clinical outcome. The early detection of genotypic resistance up to 129 days before the emergence of phenotypic resistance and the dissociation of resistance patterns among different body sites emphasize the importance of genotypic analyses of different DNA specimens for an efficient antiviral therapy. T-cell-depleted children having transplantation might be at an increased risk for the development of drug resistance.