Mice infected with the Tucson strain of coxsackievirus B1 (CVB1T) develop chronic T cell-mediated polymyositis that is manifest as the acute infection resolves and is characterized by hindquarter weakness and muscle inflammation. This model system was used to study persistence of CVB1T RNA by using reverse transcriptase-polymerase chain reaction (RT-PCR). For the most part, RNA persistence reflected the myotropic and neurotropic nature of the virus. At 1 month after infection, infectious virus was not detected in muscle, but persistent viral RNA was found in both skeletal and cardiac muscle, brain, and spinal cord. The kidney was weakly positive for viral RNA, whereas the liver and spleen were negative. Hindquarter muscle was assayed for persistent viral RNA at 1, 3, 6, 9, and 12 months after infection. In a few cases, persistent viral RNA was detected as late as 12 months after infection. The incidence of persistent viral RNA was high at 1 month after infection and gradually declined until, at 6 months and beyond, it was maintained in 3% to 12% of the muscles tested. Long-term viral RNA persistence was not more common in severely weak animals. However, the degree of hindquarter weakness that developed by 1 month was static thereafter and did not change over the 12-month study period. In contrast, separate experiments revealed that typical mononuclear cell (MNC) infiltration of muscle followed a time course similar to that of viral RNA persistence, peaking at 1 month and gradually resolving by 6 months. Infiltrating polymorphonuclear leukocytes (PMNs) and mast cells were present at 3 to 12 months after infection, signifying that some inflammatory activity remained. Other signs of myopathy that persisted for 12 months included a lack of muscle regeneration, variations in fiber size, and myofiber atrophy with increased perimysial and endomysial connective tissue. These results demonstrate that coxsackievirus RNA can persist in muscle for extended periods of time and are compatible with the idea that persistent virus is involved in maintaining the chronic MNC inflammation observed in murine polymyositis.