During a natural outbreak of mouse hepatitis virus (MHV) infection in an animal facility in Rome, 6-week-old, outbred, immunocompetent Hsd:ICR (CD-1) and immunodeficient Hsd:athymic nude-nu sentinel mice (barrier maintained) were exposed to MHV in order to study tissue distribution and duration of the virus in naturally infected mice. Infection was diagnosed by serology and by reverse transcriptase-polymerase chain reaction (RT-PCR) using primers directed to two separate but highly conserved regions of the MHV genome. Faeces, colons, spleens, lungs, brains, livers, epididymides, testes, uteri and ovaries from sentinels were tested by RT-PCR after 1, 2, 4, 6, 8 and 12 weeks. A second round of amplification with nested primers was performed to increase the sensitivity of detection. The results indicated that all the organs tested became infected with the virus at various times. Furthermore, male and female reproductive organs were infected within 6 weeks of the beginning of exposure. Investigation of MHV transmission by ovarian transplantation and by in vitro fertilization (IVF) revealed that MHV was transmitted by infected ovaries transplanted into both immunocompetent and immunodeficient mouse strains but transmission was not observed when sperm from infected testes were used for IVF. These results suggest that sperm do not transmit infection from actively infected animals and that IVF could be considered a cleansing procedure.