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SUMMARY: By treating leaves of French bean (Phaseolus vulgaris L.) in various ways at intervals after they were inoculated with the Rothamsted tobacco necrosis virus, a series of events in infected cells was detected and approximately timed. Treating leaves with ribonuclease, or floating them in water, in the first hour after inoculation decreases the number of infections ; these treatments fail to do so later, perhaps because the virus has become firmly attached to some host component, or because cells injured at inoculation have healed and no longer allow virus particles to diffuse out or ribonuclease in. After 2 hr. at 25" the virus is less readily inactivated by ultraviolet radiation than previously, suggesting that infected cells now contain more of substances that absorb radiation of 2537A. After 4 hr. the inactivation curve with ultraviolet radiation deviates from the course of a first-order reaction, probably because some infected cells now contain more than one potentially infective particle. After 6 hr. the forma- tion of some lesions cannot be prevented by irradiation, suggesting that newly formed virus has spread from initially infected cells to deeper tissues, where it is protected from inactivation. At 16" the mean time for virus to spread from the initially infected cells is 12.5 hr. and it probably spreads from some cells by 8 hr. and from most by 15 hr. Newly formed virus is not detectable in extracts of inoculated leaves until after it has spread from the epidermis to deeper tissues. Up to 20 hr. after inoculation most of the extractable virus seems to come from the epidermis, but later it comes predominantly from other tissues. When leaves of susceptible plants are rubbed with solutions of plant viruses, washed to remove surplus inoculum, and then macerated at intervals, there follows a ' latent ' period during which the infectivity of successive samples of leaf sap not only fails to increase but usually decreases. With any given inoculum and host plant, provided the conditions are kept constant, the interval between inoculation and the time when new infective particles become detectable in the sap is reasonably constant, but the latent period differs with different viruses and hosts, and it also varies with changes in either the tem- perature or the virus content of the inoculum. With the Rothamsted tobacco necrosis virus in French bean, the virus-host combination with which we shall be mainly concerned, the minimum latent period at 22" is about 10 hr. (Harrison, 1955). Little or nothing is known about what goes on during this period, and the experiments we describe below, in which leaves were sub- jected to a variety of treatments at different times after they were inoculated, were made to gain some indirect evidence about the sequence of events between the application of the inoculum and the occurrence of new infective virus.

Studies on the Multiplication of a Tobacco Necrosis Virus in Inoculated Leaves of French-bean Plants