Cord blood (CB) progenitor cells are increasingly used for transplantation in children because of the lower risk of graft-versus-host disease compared with unrelated bone marrow and comparable rates of disease-free survival. There is concern that CB might carry a higher risk of opportunistic infections. Human herpesviruses (HHV) are common pathogens in transplant recipients. CB donors are routinely tested for the presence of anti-cytomegalovirus (CMV) immunoglobulin M to reduce the risk of collecting CMV-infected CB. To assess the incidence of beta and gamma HHV infection of CB collected under standard procedures, we tested 362 CB samples for the presence of CMV; HHV-6, -7, and -8; and Epstein-Barr virus DNA by polymerase chain reaction. HHV-6 DNA was found in 2 samples, yielding an incidence of 0.55% (95% confidence interval, 0.1%-2%). None of the other viral DNAs was found, resulting in a 95% confidence interval of 0% to 1% for the incidence of CMV, Epstein-Barr virus, HHV-7, and HHV-8. Because the seroprevalence of HHV-8 among the CB donors in this study was only 4%, these findings cannot be extended to HHV-8-endemic areas. Our data show that screening prospective CB donors with anti-CMV immunoglobulin M practically eliminates the risk of CB CMV transmission, but HHV-6 warrants CB testing by polymerase chain reaction.