Laboratory of Plant Pathology, Faculty of Agriculture, Tohoku University, Tsutsumidori-amamiyamachi 1-1, Aoba-ku, Sendai 981-8555, Japan

Background and objectives
Cucumber mosaic virus strain Y (CMV-Y) induces severe yellow/white systemic mosaic symptoms in tobacco, the degree of which is different in the upper leaves. We report a comparative study of leaf number, timing and spatial pattern of the symptomatic expression as well as the virus accumulation in tobacco plants infected with CMV-Y.

Materials and methods
Nicotiana tabacum cv. Ky57 was grown in a growth cabinet at 27C for a 14-h light period (light intensity 100-130 mol/m2/s) and at 22C for a 10 h dark period. CMV-Y [1] without satellite was used at a concentration of 100 g/ml in 0.01 M potassium phosphate buffer (pH 8.0).

Results and conclusions
We inoculated CMV-Y on the 6th leaf when the plants had developed to the stage of 15 cm in length in the 6th leaf and 1 cm in length in the 9th leaf. Systemic mosaic symptoms were observed and divided into three types.

The first mosaic symptom appeared on the 8th to 9th leaf, 4 days post-inoculation. This symptom was in the form of a veinal chlorotic yellowing area. Depending on the development of these leaves, they became completely white within 10-20 days. The amount of virus coat protein and activity in the 9th leaf increased dramatically at the early stage of leaf development, and decreased quickly.

The second mosaic symptom at the 10th to 14th leaf was in the form of interveinal yellow/white chlorotic areas. The chlorotic areas were observed from the early stage of each leaf development. The unaffected green area remained so until the later stage. Some infected plants showed the form of an almost green area at the 14th leaf. The amount of virus in the chlorotic area was highest at the early stage, followed by a sharp decrease; in the green area, virus was at a very low level from the early stage, but increased a little at the later stage. We suggest that the virus was synthesized in the green areas or transported from the other areas. CMV RNA segments in the green area were analysed by Northern blot hybridization. The positive and negative strands of RNA1 or RNA2 could be detected at very low levels compared with those of RNA3 or RNA4. Some defective negative-strand RNAs were also detected. We suggest that this was one of the characteristics of the green area.

The third mosaic symptom appeared on the uppermost part of the 15th leaf. Different types of chlorotic areas were observed randomly on each leaf. One of the types was a scattering of very small, fibrous spots, one type made up of small, square, chlorotic spots, and the other type a chlorotic section. Some of the leaves developed completely green. The virus accumulation of the many green areas was very low, but in some of the green areas it was very high. These areas were observed as unrelated either to the leaf number or to the stage of leaf development, even if these were cultured under the same conditions.

We confirm that the three types of mosaic symptoms (1-3) appeared in this order when the plants were inoculated in the 6th leaf with CMV-Y at the different stages of development (8-21 cm in the 6th leaf and 0-7.5 cm in the 9th leaf). We suggest that each symptom is unique.

1. Takahashi H, Ehara Y, 1992. Molecular Plant-Microbe Interactions 5, 269-272.