Studies of the prevalence of antibody to murine cytomegalovirus (MCMV) in free-living wild mice (Mus domesticus) trapped in diverse regions of Australia and on a sub-Antarctic island indicated that 90% of 468 mice had serum antibody to MCMV. Twenty-six field isolates of MCMV were plaque-purified from salivary gland extracts of representative seropositive mice. These isolates varied considerably in their ability to replicate in the salivary glands of weanling BALB/c mice with 9 of 15 failing to reach significant titres in this organ and the titres of the remaining 6 strains varying by at least 100-fold. The high frequency of restriction fragment length polymorphisms observed suggests widespread genetic heterogeneity exists among the strains. This observation was mirrored at the polypeptide level by Western blot analyses with polyclonal antisera to MCMV. The isolation in this study of four genetically distinct strains of MCMV from a single wild mouse and several strains from other individual mice demonstrates that multiple infections with MCMV may be commonplace in wild mice.