To investigate the genetic determinants of severe chlorotic spot symptoms in cucumber mosaic virus (CMV[Y])-inoculated tobacco (Nicotiana tabacum 'Ky57') leaves, symptom expression was compared among tobaccos (N. tabacum 'Ky57', N. tabacum 'Xanthi nc', and their hybrids) inoculated with CMV(Y), CMV(O), or pseudorecombinants of both strains of CMV. Xanthi nc showed very mild chlorotic spot symptoms in CMV(Y)-inoculated leaves. Both CMV(O)-inoculated Ky57 and Xanthi nc leaves showed no symptoms. In pseudorecombinant-inoculated Ky57 leaves, severe symptoms were determined by chimeric CMV(O) and CMV(Y) RNA3 transcripts, which contained most of the coat protein gene of CMV(Y). Genetic analysis of severe symptom formation in CMV(Y) or pseudorecombinant-inoculated tobaccos indicated that two recessive nuclear-coded host genes determined severe symptom formation in CMV(Y)-inoculated Ky57. The two recessive host genetic loci completely cosegregated with the two recessive burley loci (yb1 and yb2 genes) of Ky57, whose character is an apparent reduction in chlorophyll of the stems. When several cultivars of N. tabacum were inoculated with CMV(Y), all burley tobaccos clearly showed severe symptoms, but most other cultivars showed very mild symptoms. These results suggest that severe chlorotic spot symptom formation in CMV(Y)-inoculated Ky57 is induced by a combination of the virus coat protein gene and two recessive host genes that are closely linked to or identical to yb genes.