This study examined the existence of genetically diverse population of Cucumber mosaic virus (CMV), known as quasispecies, from lily, Nicotiana benthamiana and from purified virions. Based on the conserved sequences of CMV lily isolates in intergenic region (IR) on RNA3, the genetic variation of IR from three different sources was investigated by a specific restriction endonuclease hydrolysis of amplified reverse transcription-polymerase chain reaction (RT-PCR) products using virusspecific primers, and was compared with IR sequences. The IR nucleotide sequences of CMV lily isolates were highly conserved, however, quasispecies was detected from all three sources in low level, containing subpopulations of RNA3. These subpopulations of RNA3 were inoculated onto zucchini squash by in vitro transcripts from corresponding full-length cDNA clones together with Fny RNA1 and 2 transcripts. The systemic symptom of zucchini plants infected by these quasispecies was chlorotic spotting, which was milder than severe mosaic and stunt symptom caused by Fny-CMV. The severity of symptom was correlated with RNA accumulation of viruses. These results suggest that the genome of CMV lily isolates consists of quasispecies populations.