The 3'-untranslated region of RNA1 as a primary determinant of temperature sensitivity of Red clover necrotic mosaic virus Canadian strain.

Red clover necrotic mosaic virus Canadian strain (RCNMV-Can) induces symptoms on host plants at 17 degrees C, but not at 25 degrees C. We investigated the temperature sensitivity of RCNMV-Can in Nicotiana benthamiana plants and protoplasts using infectious transcripts of genomic RNAs 1 and 2. Viral RNAs accumulated in both inoculated and noninoculated leaves at 17 degrees C, whereas no viral RNAs were detected at 25 degrees C in either inoculated or noninoculated leaves. Similar temperature sensitivity in RNA accumulation was observed in protoplasts, and no viral RNAs were detected at temperatures above 22 degrees C. These results indicate that the temperature sensitivity of RCNMV-Can occurs at an early stage of infection, including during RNA replication. Using reassortant viruses and chimeric RNAs 1 between RCNMV-Can and the RCNMV Australian strain, which accumulates viral RNAs at nonpermissive temperatures for RCNMV-Can, we demonstrated that a viral determinant for the temperature sensitivity resides in the 3'-untranslated region of RNA1.