Despite decreasing prevalence, rotavirus infections still rank among the most important viral infections in colonies of laboratory mice. Although the disease is characterized by low mortality and a relatively short and mild clinical period, the infection has the potential to alter the outcome of experiments substantially. For animal facilities, it is therefore essential to eradicate the virus. Here we report a successful sanitation of a rotavirus-infected mouse colony in an animal facility. Despite a high ratio of transgenic and partially immunodeficient strains, a permanent eradication of the virus was achieved by euthanasia of highly susceptible mice, a prolonged breeding cessation in areas containing immunocompromised mice and a strict hygienic management. The management of a rotavirus infection reported here is a feasible and inexpensive opportunity for sanitation that benefits from maintaining most of the animal population, even in today's mouse colonies comprising mainly transgenic mice with unknown or compromised immune status.