Swine testicle (ST) cells and Madin-Darby canine kidney (MDCK) cells differ in their ability to support replication of porcine parvovirus (PPV). Viral replication events in ST cells, a permissive cell type, and MDCK cells, a nonpermissive cell type, were compared in an attempt to elucidate putative mechanisms of restrictive virus replication. Radiolabeled PPV bound to the cell surface of both cell types equally well and the binding was shown to be PPV specific, indicating that the restriction was not at the cell surface level. In contrast, profound differences in intracellular events in PPV replication were observed between these two cell types. Synthesis of viral DNA was limited in MDCK cells in that the percentage of cells with replicative-form DNA as determined by strand-specific probe in situ hybridization was approximately 100-fold lower in MDCK cells than in ST cells at the same multiplicity of infection. Northern (RNA) blot analysis, using oligonucleotide probes derived from both structural and nonstructural protein-coding regions of the PPV genome, revealed four PPV mRNA transcripts from infected ST cells. Comparatively, RNA species from the structural protein coding region were actively transcribed in MDCK cells, but synthesis of RNA species from the nonstructural protein coding region was negligible. Immunoprecipitation of viral polypeptides revealed the three characteristic structural polypeptides, VP1, VP2, and VP3, along with the nonstructural polypeptide, NS-1, from ST cells. In contrast, neither viral structural or nonstructural polypeptides nor progeny virions were produced from MDCK cells. The data suggest that mechanisms controlling permissiveness of cells to PPV infection are associated with the level of viral DNA replication, RNA transcription, and viral antigen expression but not absorption to the cell surface.