Abstract: PnV (Perina nuda iflavirus) was found to infect persistently its heterologous cell line, NTU-LY-1. The cytopathic effect of PnV to NTU-LY-1 cells was different from that of PnV to its homologous NTU-PN-HF cells. The membrane-bound inclusion bodies (IBs) were found in newly seeded NTU-LY-1 cells. The lysed NTU-LY-1 cells formed many microsomes 15 days post passage. Three different inclusion bodies (IBs) could be distinguished among the microsomes: membrane-bound IBs, fibril-bound IBs, and sheath-bound IBs. At 15 days post-passage, the cytoplasm of un-lysed NTU-LY-1 cells contained zipper-like array of unmembrane-bound virogenic stroma, the parallel rows of viral particles were restricted by two membranes. These viral particles were purified and showed a typical PnV morphology by negative stain. They were further examined by RT-PCR using a primer set designed from the sequence of a putative PnV helicase gene and by Western blot. Indirect immunofluorescent assay with a specific anti-P. nuda picorna-like virus (PnV) antiserum confirmed that PnV extends its in vitro host range to NTU-LY-1 cell line. These results suggest that PnV could extend its in vitro host range to NTU-LY-1 cell line and this persistent infection of PnV is a model for the membrane-associated replication and translation for positive sense ssRNA viruses.