SOME EFFECTS OF CHANGING TEMPERATURE AND OF VIRUS INHIBITORS ON INFECTION BY CUCUMBER MOSAIC VIRUS

Whereas the spinach strain of cucumber mosaic virus fails to multiply and cause symptoms in tobacco plants kept above 30° C., the yellow strain infects at 36° C. and causes more severe symptoms than at 20° C. Increasing the temperature up to 28° C. increases the initial rate at which the spinach strain multiplies, but the virus later reaches much higher concentrations in leaves at lower temperatures, presumably because it is rapidly inactivated at 28° C. Exposing inoculated plants to 36° C. for 6 hr. decreases the number of infections by the spinach strain when the exposure starts within 6 hr. of inoculation, but not afterwards.
Pancreatic ribonuclease inhibits infections by strains of cucumber mosaic virus; inhibition is greatest when the enzyme is present in the inoculum, and when applied to inoculated leaves its effect decreases rapidly with increasing time after inoculation.
Infection by and the multiplication of strains of cucumber mosaic virus in tobacco are only slightly affected by thiouracil and greatly by azaguanine, whereas strains of tobacco mosaic virus are inhibited much more by thiouracil than by azaguanine. Like tobacco mosaic virus, cucumber mosaic virus multiplies more when inoculated leaves are floated in nutrient solutions than in water, but unlike tobacco mosaic virus, its multiplication is not inhibited by thiouracil more in nutrient solutions than in water.     

THE EFFECT OF TEMPERATURE ON INFECTION WITH STRAINS OF CUCUMBER MOSAIC VIRUS

Cucumber mosaic virus strains differed in their ability to multiply in plants at 37° C. Some strains multiplied in inoculated leaves and produced systemic symptoms in plants at this temperature; plants systemically infected with one such strain remained infected after prolonged treatment at 37° C. Other strains did not appear to multiply in inoculated leaves at 37° C. and heat treatment was successful in freeing plants from infection with these. Tests with one strain of each type showed both to be rapidly inactivated in expressed sap at 37° C.
Strains of cucumber mosaic virus forming small necrotic local lesions in leaves of french bean var. Canadian Wonder, produced many fewer lesions in plants kept after inoculation at 25° C. for 24 hr. and then at 15° C. than in plants kept continuously at the lower temperature.     

SOME EFFECTS OF TEMPERATURE AND OF VIRUS INHIBITORS ON INFECTION OF FRENCH-BEAN LEAVES BY RED CLOVER MOTTLE VIRUS

Keeping French-bean plants before inoculation at 36, 32 or 28°C. for 1–2 days increased their susceptibility to infection with red clover mottle virus, but longer exposures to 36 and 32°C. decreased susceptibility. Susceptibility increased most rapidly at 36°C. The number of infections was unaffected by changes in post-inoculation temperatures between 12 and 24°C., but decreased above 24°C. The rate virus multiplied increased with increase of temperature up to 28°C., but the maximum virus concentrations reached at 18, 24 and 28°C. were very similar and above the maximum reached at 30°C.
Thiouracil inhibited infection slightly but neither it nor azaguanine affected the multiplication of red clover mottle virus in French bean. Trichothecin inhibited infection and interfered with virus accumulation. Inhibition of infection was associated with macroscopic injury to the leaves, and washing leaves up to 1 hr. after inoculation prevented both inhibition and leaf damage. Virus multiplication was not resumed when leaves were transferred from trichothecin solutions to water.     

SOME EFFECTS OF HIGH TEMPERATURE ON THE SUSCEPTIBILITY OF PLANTS TO INFECTION WITH VIRUSES

When plants were kept at 36°C. for some time before inoculation, their susceptibility to infection by five mechanically transmissible viruses was greatly increased. When kept at 36° after inoculation, fewer local lesions were produced than at lower temperatures, but the effects of the post-inoculation treatment differed with different viruses. Tomato spotted wilt and tobacco mosaic viruses multiply in plants at 36°, and the post-inoculation treatment reduced the local lesions they caused to numbers that varied between 10 and 90% of the control; these two viruses also have large thermal coefficients of heat inactivation. By contrast, tobacco necrosis, tomato bushy stunt and cucumber mosaic viruses, were much affected by post-inoculation treatment, lesion formation being completely prevented by exposure to 36° for a day or more. These three viruses appear not to multiply in plants at 36°, and although they have high thermal inactivation points, they have small temperature coefficients of thermal inactivation.
The extent to which lesion formation was affected by pre- or post-inoculation exposure of plants to 36° depended not only on the length of the treatment, but also on the physiological condition of the plants.
The symptoms of infected plants changed considerably if kept at 36°. At 36° Nicotiana glutinosa , inoculated with tobacco mosaic virus, gave chlorotic local lesions instead of necrotic ones, and became systemically infected. When systemically infected plants were brought to ordinary glasshouse temperature, the infected tissues all collapsed and died in a day.     

THE EFFECTS OF INFECTION WITH TOBACCO MOSAIC VIRUS ON THE PHOTOSYNTHESIS OF TOBACCO LEAVES

The rate of photosynthesis of tobacco leaves infected with the Rothamsted type culture of tobacco mosaic virus was lower than that of comparable healthy tobacco leaves. The lower rate was inferred from Net Assimilation Rates of whole plants and confirmed by direct comparisons of photosynthetic rates of inoculated and healthy leaves. The effect began within 1 hr. of inoculation. It was not caused by an effect of the virus on the stomata, and inactivated virus inoculum did not change the rates. The results indicate either a more rapid movement of virus from the epidermis into the chlorenchyma than has been previously recorded or an effect of virus infection at a site distant from the cells containing virus.     

FACTORS AFFECTING THE PRODUCTION OF LOCAL LESIONS BY PLANT VIRUSES

Beans inoculated with tobacco necrosis virus were kept in the dark at different temperatures for 1 hr. before and 1 hr. after inoculation; in this experiment the number of lesions increased with temperature over the range 55–82° F.
The effect of 30 min. periods of darkness before or after inoculation depended on the time of day, the number of local lesions usually being decreased. Prolonging the night period before inoculation sometimes increased the number of lesions.
Light appeared to be more important than temperature in controlling the daily variation in susceptibility. However, in a test over a 30 hr. period this variation continued even when plants were placed under constant conditions before and after inoculation.
When plants that had been kept in the dark were exposed to light of about 800 f.c. intensity for 1 min. immediately before inoculation the number of local lesions was doubled.     

ANEMONE MOSAIC–A VIRUS DISEASE

The name anemone mosaic is proposed for a previously unrecorded virus disease of Anemone coronaria L.; infected plants have mottled leaves, and broken and distorted flowers. This virus can cause winter browning, and can contribute to crinkle in anemones.
The virus infected forty-seven out of ninety plant species tested; it was transmitted by mechanical inoculation, and by four of the six aphid species tested. Most aphids ceased to be infective within 30 min. when continuing to feed after leaving an infected plant.
Properties in vitro varied according to conditions of the tests; the thermal inactivation point was always below 62°C., the dilution end-point did not exceed 1/2500, and the virus inactivated at 18°C., the fewer than 72 hr.
Intracellular inclusion bodies were produced in all hosts examined.
Anemone mosaic virus is very similar to viruses placed in the turnip virus 1 group of Hoggan & Johnson, and is serologically related to cabbage black ringspot virus, although AMV infection did not protect plants against infection with cabbage black ring-spot virus.
Weeds naturally infected with AMV were found in anemone plantations, and this virus was detected, together with cucumber mosaic and tobacco necrosis viruses, in corms imported into this country.     

THE TRANSMISSION BY MITES, HOST-RANGE AND PROPERTIES OF RYEGRASS MOSAIC VIRUS

A virus that causes chlorotic streaks on ryegrass leaves was transmitted by the eriophyid mite Abacarus hystrix (Nalepa). Virus-free mites acquired the virus in 2 hr. feeding on infected ryegrass and the proportion that became infective increased with increased feeding time up to 12 hr.; vectors lost infectivity within 24 hr. of leaving the infected leaves. All instars of A. hystrix transmitted the virus.
The virus was transmitted by manual inoculation of sap to other species of Gramineae, including oats, rice, cocksfoot and meadow fescue, but none of these hosts seemed to contain as much virus as ryegrass; their saps did not precipitate specifically with antiserum prepared against the virus in ryegrass, whereas sap from infected ryegrass precipitated up to a dilution of 1/32. Infective sap of S22 Italian ryegrass contained flexuous rod-shaped particles; the dilution end-point of the virus was about 1 in 1000; the virus was inactivated when held for 10 min. at 60°C. and most of its infectivity was lost after 24 hr. at room temperature.     

Effect of Temperature on Susceptibility of the Primary Leaves ofPhaseolus vulgaris L. to Red Clover Mottle Virus

Red clover mottle virus isolated in Czechoslovakia was studied in relation to its reaction to varying temperature on primary French bean leaves (Phaseolus vulgaris L.) on which it forms local necrotic lesions. The plants were kept 24 or 48 h before, or 24 or 48 h after inoculation at the temperatures 23, 25, 27, 30, 33 and 36°C. After such exposures the French beans were kept at a constant temperature of 25°C. The lesions were counted at various intervals. In the experiment the optimal temperature for the maximum number of lesions seems to be 36°C 48 h before inoculation. The temperature above 25°C applied 24 h after inoculation seems to have a decreasing effect upon the number of lesions formed by RCMV on primary leaves of French beans and the lesions appeared several hours later, especially at 30, 33 and 36°C. The temperatures 27, 30 and 33°C applied 48 h after inoculation have a further decreasing effect on the number of lesions. The temperature of 36°C applied 48 h after inoculation has an inactivating effect upon RCMV inoculated on French bean leaves and no lesions appeared 5 days after inoculation.     

Effects of heat shock treatment on symptom development in tobacco leaves inoculated with potato mop-top virus

PMTV normally causes necrotic ringspot local lesions in Xanthi-nc tobacco leaves at 15 °C but not at 22°. Dipping the leaves once in hot water at different intervals after inoculation induced necrotic reactions even at 22°. Successive concentric rings, one for each day of the interval between inoculation and treatment, were sometimes induced at 22°; these even formed in continuous lighting, suggesting an intrinsic diurnal rhythm. No lesions were induced at 22° by dipping inoculated leaves in ice-cold water. Samsun tobacco inoculated with PMTV developed very few lesions at 15°. Many more were induced however, by dipping the leaves in hot water after inoculation. The results suggest some similarities between PMTV and TMV, and the two viruses have been reported as serologically related; but PMTV did not protect plants against TMV, for TMV challenge-inoculations induced lesions within existing PMTV lesions.     

THE EFFECT OF INFECTION WITH TOBACCO MOSAIC VIRUS ON THE RESPIRATION OF TOBACCO LEAVES OF VARYING AGES IN THE PERIOD BETWEEN INOCULATION AND SYSTEMIC INFECTION

Leaves of tobacco plants inoculated with tobacco mosaic virus were divided into three groups: ( a ) inoculated leaves; ( b ) younger non-inoculated leaves present at the time of inoculation; ( c ) leaves formed since inoculation. The respiration rate of each group was compared with that of similar leaves from healthy plants. The respiration rate of inoculated leaves was increased by a constant amount for 3 weeks after inoculation, when it decreased. The respiration rate of group ( b ) leaves was not affected at any time, and that of group ( c ) leaves was decreased by 10% when they showed symptoms. The increased respiration in the inoculated leaves occurred too soon to reflect virus formation, and it is suggested that it reflects an initial change in infected cells preparatory to virus synthesis. The subsequent decrease in respiration may be due to the accumulation of virus which does not contribute to the total leaf respiration.     

PHOTOSYNTHESIS AND RESPIRATION RATES OF LEAVES OF NICOTIANA GLUTINOSA INFECTED WITH TOBACCO MOSAIC VIRUS AND OF N. TABACUM INFECTED WITH POTATO VIRUS X

The rates of respiration and of photosynthesis of tobacco leaves infected with potato virus X were not affected until the leaves showed symptoms; the respiration rate was then increased by more than 30% and the photosynthesis rate decreased by 20%. When local lesions appeared on the leaves of Nicotiana glutinosa infected with tobacco mosaic virus, but not before, the respiration rate was increased by an amount, up to 30%, that varied with the number of lesions. The photosynthesis rate was decreased by 20%, but there was no effect on photosynthesis or respiration until symptoms appeared. These results differ from those previously reported for tobacco leaves infected with tobacco mosaic virus, in which both respiration and photosynthesis were affected within 1 hr. of inoculation. The validity of extrapolating arguments based on the results obtained with other combinations to this commonly used combination and vice-versa is questioned.     

Effect of thermal shock treatments on symptom expression in test plants inoculated with potato aucuba mosaic virus

Potato aucuba mosaic virus (PAMV) has few reliable local lesion assay hosts. However, lesions formed when PAMV-inoculated leaves were exposed to thermal shock (dipping for 40 s in water at 50 or 2 °C). Leaves of Nicotiana tabacum (cv. Xanthi-nc, Samsun or Samsun NN), Hyoscyamus niger and Datura metel consistently developed necrotic lesions, leaves of Chenopodium amaranticolor developed whitish rings, and leaves of N. glutinosa developed diffuse cream-coloured rings and spots. In PAMV-inoculated leaves of Xanthi-nc tobacco, C. amaranticolor and D. metel, lesions formed only in areas exposed to light. Thermal shocks applied to systemically infected leaves of Xanthi-nc tobacco induced necrotic vein banding patterns. In inoculated Xanthi-nc tobacco leaves, PAMV seemed confined to local lesions. The rate of lesion enlargement was therefore a measure of rate of virus spread. Lesion size increased as the interval between inoculation and shock treatment increased. The mean rates of increase in lesion radius were 17 and 27 Cμm/h at 15 and 25°C respectively. ‘Target’ lesions, composed of concentric necrotic rings, formed when inoculated Xanthi-nc tobacco leaves were given two or more 50°C shocks. The first of two 50°C treatments decreased subsequent rates of lesion enlargement.     

Can Salicylic Acid Affect the Intercellular Transport of the Tobacco Mosaic Virus by Changing Plasmodesmal Permeability?

We studied the effects of salicylic acid (SA) on the plasmodesmal permeability as evaluated by the tobacco mosaic virus (TMV) spreading in tobacco Nicotiana glutinosaleaves, where TMV induces necrotic lesions. When leaves were treated with SA simultaneously with their viral inoculation, SA retarded the development of necrotic lesions and reduced their number. When inoculated leaves were kept on the SA solution at an elevated temperature (31°C) for a short period of time, the size of the necrotic lesions, which developed after leaf transfer to room temperature, was decreased. SA stimulated the formation of rapid callose involved in the control of the plasmodesmal permeability, which was assessed from fluorescence after tissue staining with Aniline Blue. On the basis of these data, we suggest that SA suppressed TMV spreading in the inoculated tobacco leaves by reducing the plasmodesmal permeability.     

HEAT-THERAPY OF VIRUS-INFECTED PLANTS

Virus-free plants were produced from parents systemically infected with the following five viruses: tomato bushy stunt, carnation ring spot, cucumber mosaic, tomato aspermy and Abutilon variegation. The leaves formed while the infected plants were kept at 36°C. were free from symptoms, and test plants inoculated from these remained uninfected. When cuttings were taken from the infected plants at the end of the treatment most grew into healthy plants. The treated plants themselves usually developed symptoms after varying lengths of time at 20°C, but some that before treatment were infected with tomato aspermy, cucumber mosaic or Abutilon variegation viruses, remained permanently healthy.
The same method failed to cure plants infected with tomato spotted wilt, potato virus X and tobacco mosaic virus, although it decreased their virus content. Heat-therapy seems not to be correlated with the thermal inactivation end point of the virus in vitro.     

THE EFFECT OF INFECTION WITH TOBACCO ETCH VIRUS ON THE RATES OF RESPIRATION AND PHOTOSYNTHESIS OF TOBACCO LEAVES

Unlike tobacco mosaic virus, which increases the respiration of tobacco leaves within an hour of their being inoculated, a virulent strain of tobacco etch virus did not change respiration rates until leaves showed external symptoms. The respiration rates of inoculated or systemically infected leaves with symptoms rose to 40% above that of healthy leaves, three times the increase produced by tobacco mosaic virus. The increased respiration rate occurred at all times of the year and was maintained through the life of the leaves.
Leaves infected with tobacco etch virus and showing symptoms had a photo-synthetic rate 20% lower than that of healthy leaves.     

SOME EFFECTS OF VIRUS INFECTION ON LEAF WATER CONTENTS OF NICOTIANA SPECIES

Infection with tobacco mosaic virus decreases the water content which detached tobacco leaves attain when kept for 20 hr. in conditions of minimum water stress, and does so more when the plants are kept in light before inoculation than when they are kept in darkness. No such effects of infection during the first day after inoculation were obtained with tobacco leaves infected with either tobacco etch virus or potato virus X , or with Nicotiana glutinosa leaves infected with tobacco mosaic virus. These results, like those showing early effects of TMV on respiration and photosynthesis of tobacco leaves, suggest that inoculation with TMV affects deeper leaf tissues than the epidermis earlier in tobacco leaves than in other leaves, and earlier than other viruses in tobacco leaves.     

THE INFLUENCE OF TEMPERATURE AND DAYLENGTH ON SPOTTED WILT VIRUS DISEASE OF TOMATO

The responses of young tomato plants to infection with tomato spotted wilt virus were studied at constant temperatures. The plants were grown in nutrient solutions and given three daylength treatments.
The length of the incubation period was positively correlated with temperature and a regression equation for this relationship is given. The calculated temperature at which this period becomes infinite is 6.7°C. The temperature coefficient ( Q ₁₀) varied from 1.7 to 2.9.
Dry-weight determinations were made of stems, leaves and roots. In healthy plants the leaf/stem ratio generally fell with rise of temperature and the shoot/root ratio rose. Infection changed these ratios, which is interpreted as indicating that it interfered with supplies of carbohydrate to the growing regions.
Systemic symptoms are described for plants raised at 11.4°, 18.3°, 23.9°, and 29.4°C.
Changes in daylength did not affect the systemic symptoms or incubation period. With a 9 hr. day at 29.4°C., the virus content of sap was significantly lower than with 12 or 15 hr., and with a 12 hr. day was significantly higher than with a 15 hr. day.     

VIRUSES OF COWPEA, VIGNA UNGUICULATA L. (WALP.), IN NIGERIA

The cowpea strain of tobacco mosaic virus was isolated from a range of leguminous hosts at Ibadan, but was rare in cultivated crops. Systemic symptoms in species infected experimentally are described.
A new virus of cowpea was also found in Nigeria. The physical properties (thermal inactivation point 56° C., dilution end-point 1/50,000 and longevity in vitro 4 days at 25° C.) differ from those for cowpea viruses reported elsewhere and the name cowpea yellow mosaic virus is proposed. This virus produces local lesions in French bean ( Phaseolus vulgaris L.) and local and systemic lesions in Bengal bean ( Mucuna aterrima Holland), but does not infect other leguminous hosts. The virus was purified and an antiserum prepared against it.
Both viruses are transmitted by a beetle ( Ootheca mutabilis Sahlb.) which loses infectivity within 48 hr. of leaving plants infected with either or both viruses.     

Effects of Salicylate on Virus-Infected Tobacco Plants

Salicylate watered onto the soil of tobacco plants in pots reduced the antigen accumulation and local lesion growth of tobacco necrosis virus mechanically inoculated on the leaves. It also retarded the growth of the necrotic centres of lesions and, in parallel, inhibited ethylene production from infected leaves. However, the therapeutic index of salicylate was very small and the chemical had to be applied in advance of, or at the same time as virus inoculation to give good levels of resistance. The number of lesions and their rate of appearance were not affected by salicylate. In addition, it did not induce resistance against multiplication, systemic spread or symptom severity in tobacco plants inoculated with a necrotic strain of potato virus Y. These findings suggest that salicylate is not likely to prove useful as polyvalent chemotherapeutic agent.