Intracellular Inclusions in Potato and other Solanaceous Plants affected by the X-and Y-Viruses of Potatoes

Abstract

The Lugol-Giemsa method described by Bald (1949), trypan-blue method by McWhorter (1941), and-pyronin-methyl green stain were tested for selective staining of the intracellula inclusions in potato and other Solanaceous plants produced by the X-and Y-virues of the potato. The tissue observed was integumentary, consisting of epidermal peelings from disaesed leaves and petioles. The Giemsa method brought about the best results, staining the viroplasm and the nucleolus violet (not purple) and the nucleus light blue; while the pyronin-methyl green stain failed to provide any differentiation, staining both the healthy and the diseased plasm red. The trypan-blue staining was found to be unsatisfactory.
Potato virus X (PXV)-Deeply staining material, presumably virus, was found to extend around the cells in potato, tobacco and tomato plants infected with the mottle strain of virus X (Fig. 3, C, D, G; and Fig. 1 in Plate). A more disperse material, also staining violet with Giemsa, was detected in the cytoplasm or near the nuclei (Fig. 3, A, B, E, H, I), which, in some cases, entirely embedded the whole cell content (Fig. 3, F). The apparently healthy potato carieties, carrying virus X, contained the same inclusions in the later stage of growth. The needle-like crystalline inclusions, stained with Giemsa and dissolved in acid and alkali, were also observed in the same varieties (Fig. 1).
Potato virus Y (PYV)-A small granular inclusions were present in potato, tabacco and tomato plants infected with the ordinary strain of virus Y (Fig. 3, J, K, O; and Fig. 2-4 in Plate). The deeply staining and the disperse materials which somewhat resemble those caused by virus X, were also detected in the same Plants (Fig. 3, L-N). Potato variety, Iwate No. 4, which carries virus Y, has contained the same inclusions. The intranuclear crystalline inclusions showing the proteinaceous reactions were found in fall in tobacco plants grown in the dry soils (Fig. 2).
Potato virus X plus virus Y (PXV+PYV)-In this case, the PXV-type inclusios alone or associated with the PYV-type inclusions were observed in one cell in potato and tobacco plants (Fig. 3, P, Q; and Fig. 5-7 in Plate). On the other hand, a large vacuolated inclusions (X+Y type) were also prevalent, especially in potato plants (Fig. 3, R-U; and Fig. 8 in Plate). The chemical nature of the inclusions-Results obtained with the materials infected with virus Y are now presented below. The Millon reaction and the xanthoprotei test provided evidence of protein in the inclusions. They were also positive to Nadi and benzidin reactions (but not inhibited by KCN). They were not stained in the Feulgen reaction. When the materials were fixed with Regaud's solution or treated with hot (60°C) hydrochloric acid (both the treament to remove ribonucleic acid), the inclusions were not stained with thionin, toluidin blue, methylene blue, and oresyl blue. The materials pretreated with tri chloracetic acid (90°C, 15min.), did not absorb any dyes tested except the trypan-blue. These results seem to suggest the presence of ribonucleic, acid in the inclusions. the reason why the Nadi and benzidi reactions were positive in the inclusions remains unknown, but the writers should like to consider that the non-crystalline inclusions are composed of both viruses and some of the host materials. The hest materials, like mitochondria and so on, have been demonstrated to react to Nadi solution when the host cells resist to the infection by viruses or fungi. It seems reasonable, therefore, to conclude that the inclusions, especially non-crystalline, may be the product of the interaction of viruses and host materials, though the crystalline inclusions within the nuclei have proved to be a pure nucleoprotein.