Severe adenovirus infections are an increasing threat for immunosuppressed individuals, in particular those receiving stem cell transplants. It has been previously hypothesized that severe infections might be due to reactivation of a persistent infection, but this hypothesis has been difficult to test due to the lack of a permissive in vivo model of human adenovirus (HAdV) infection. Here we established a humanized mouse model that reproduces both features of acute and persistent HAdV infection. In this model, acute infection correlated with high mortality, weight loss, liver pathology and expression of viral proteins in several organs. In contrast, persistent infection was asymptomatic and led to establishment of HAdV-specific adaptive immunity and expression of early viral genes exclusively in the bone marrow. These findings validate the use of humanized mice to study HAdV acute and persistent infection, and strongly suggest the presence of cellular reservoirs in the bone marrow.