Acquired Resistance in Hypersensitive Tobacco against Tobacco Mosaic Virus, Induced by Plant Cell Wall Components

Isolated cell walls from Nicotiana tabacum L. cv. Xanthi-nc were partially digested with enzymes present in the fungal preparation cellulase Onozuka R-10. Released carbohydrate material was separated from the enzymes by gel filtration and fractions were injected into the intercellular spaces of one half of each of several Xanthi-nc leaves. Two to three days after injection the whole leaves were inoculated with TMV. A large reduction in mean lesion diameter was observed in the injected areas compared with those in untreated tissue.     

The Effect of Light on Tobacco Mosaic Virus (TMV) Formation

Discs were punched from TMV-inoculated tobacco leaves (Nicotiana tabacum L.) and illuminated while floating on half strength Vickery's solution maintained at 24°C. After 48 hours some discs were placed in the dark for 24 hours and the amount of TMV formed in the light and dark compared. Discs from young leaves formed more virus in the light than in the dark. Discs from older leaves produced less virus, but as much in the dark as in the light.     

The Effect of Post Inoculation Treatment with Mineral Oil on TMV Multiplication in Tobacco Leaves and Protoplasts

Half leaves of N. tabacum dipped into a 0.2 % emulsion of meneral oil 15min after inoculation with tabacco masaic virus (TMV) developed significantly fewer lesion than when not dipped Dipping reduced lesion numbers when applied up to 2½ h after inoculation and multiple dippings were more effective than one. The effect of oil was the same whether the inoculum was whole virus or RNA. Tobacco protoplasts treated with mineral oil contained less virus than untreated protoplasts. The oil probably acted by killing the protoplasts and was effective only when protoplasts were centrifuged through the oil emulsion, When water-treated leaves were dipped into a TMV solution there was an effect on TMV infection similar to that caused by dipping in oil after TMV inoculation.     

Retarded Stomatal Closure by Phenylmercuric Acetate

Excised leaves of Nerium oleander, which were treated with phenylmercuric acetate (PMA) 1_1/2 h before excising, transpired faster than untreated excised leaves. Similarly, PMA-treated oleander plants transpired more than untreated plants in the dark. These effects were due to retarded stomatal closure caused by PMA. Measurements of stomatal apertures on disks of Vicia faba leaves kept in the dark, and of diffusive resistance to water vapor from Phaseolus vulgaris leaves, confirmed that PMA retards stomatal closing as well as stomatal opening. However, day-time reductions in transpiration by PMA greatly exceed night-time increases in water loss. The mechanisms of stomatal movement, as affected by PMA, are discussed. PMA may conceivably decrease the permeability of guard cell membranes to solutes, thereby retarding all stomatal movements that are osmotically induced.     

Influence of foliar and systemically applied azadirachtin on host‐plant evaluation behaviour of the sweetpotato whitefly,Bemisia tabaci

Abstract The behaviour of female adult Bemisia tabaci is observed for a period of 20 min after initial contact with untreated cucumber leaves, or leaves either foliar or systemically treated with azadirachtin, to determine whether application of azadirachtin affects the host‐evaluation behaviour and whether the behaviour on treated leaves differs between application methods. Application of azadirachtin deters settling of the whiteflies on host plants. The whiteflies probe for shorter duration and less frequently but spend longer and engage more frequently in labial grooming on treated leaves than on untreated leaves. Behavioural transition between probing and other behavioural elements is less common, and that between labial grooming and other behavioural elements more common, on both treated leaves than on untreated leaves. No difference is detected in the host‐evaluation behaviour of B. tabaci between leaves foliar treated and systemically treated with azadirachtin.     

Phytochemical Studies on Tobacco Alkaloids

From the findings of the feeding experiments of d- and l-nicotine-¹⁴CH₃ and d-nicotine to the excised tobacco leaves, it was demonstrated that demethylation of nicotine was stereospecific for the d-form in tobacco leaves. Such preferential demethylation to the enantiomer was also observed with N-methylanabasine and 1-(3′-pyridyl)-1-methylaminoethane which are analogous compounds to nicotine.     

Lipid Compositions of Photomixotrophic Green Calluses and Chlorophyll Deficient Leaves of Tobacco

Among photomixotrophic green calluses tested (N. rustica. N. tobacum L. cv. BY-4 and Samsun), the callus of Samsun had the highest contents of chlorophyll and chloroplast lipids, such as monogalactosyldiglyceride (MGDG), digalactosyldiglyceride (DGDG), sulfoquinovosyldigly-ceride (SQDG) and phosphatidylglycerol (PG). However, the chlorophyll and chloroplast lipids in the green callus of Samsun were still 1/6 and 1/3 of that in the parent leaves, respectively. The relative content of a-linolenate in MGDG, DGDG and SQDG of the green calluses were higher than that of the white calluses. The ratios of hexadecatrienoate in MGDG and hexadeceno-ate (Δ³-trans) in PG in the green calluses were trace or less compared with that of the parent leaves. The crude lipids and total fatty acid contents of the chlorophyll deficient leaves (N. taba-cum L. cv. Consolation 402 and Dominant Aurea Su/su) were almost the same as those of the normal leaves (cv. BY-4 and Samsun), although the chlorophyll contents of the chlorophyll deficient leaves were 1/3 ~ 1/4 of that of the normal leaves. The ratios of chloroplast lipids in the total polar lipids in the chlorophyll deficient leaves were a little lower than that in the normal green leaves, but the former had a slightly higher ratio of phospholipids such as phosphatidylcholine and phosphatidylethanolamine than the latter. There were few differences in the fatty acid compositions of each individual lipid betweeen both types of leaves.     

Species‐specific elicitors induce tea leaf to arrest the endoparasitoid Ascogaster reticulata (Hymenoptera: Braconidae)

Ascogaster reticulata Watanabe (Hymenoptera: Braconidae) is an egg‐larval endoparasitoid of the smaller tea tortrix, Adoxophyes honmai Yasuda. Recent studies have examined tritrophic interactions among Camellia sinensis, A. honmai and A. reticulata, but the effect of non‐host insects on the induction of tea plant that may affect foraging behaviour of A. reticulata remains unclear. In this study, we selected two non‐host insects, Homona magnanima Diakonoff and Ostrinia furnacalis (Guenée), as representative species in our bioassays. Tea leaves were treated with homogenized female reproductive tissues of a non‐host insect in comparison with untreated leaves in a choice test. Residence times of parasitoids on both leaves were recorded. The parasitoids seemed to prefer walking on leaves treated with homogenates of H. magnanima over untreated leaves, but the difference in residence times was not significant. In contrast, its residence time on leaves treated with homogenates of O. furnacalis was significantly shorter than that on untreated leaves. Thus, the induction of tea leaf surface chemicals may differ among moth species, which may produce different types of elicitors. This difference may, in turn, affect the host‐searching behaviour of A. reticulata.     

Effect of Tobacco Mosaic Virus on Phospholipid Content and Phospholipase D Activity in Tobacco Leaves

The phospholipid content and phospholipase D activity in the leaves of two tobacco (Nicotiana tabacum L.) cultivars were investigated. These cultivars are characterized by different response to the infection with tobacco mosaic virus (TMV). In the infected leaves of a susceptible cv. Samsun, phospholipid content and phospholipase D activity did not change within seven days after TMV infection. The development of a hypersensitive response in the leaves of a resistant cv. Xanthy necrotic was not accompanied by a change in the total phospholipid content as compared to the noninfected leaves. However, the appearance of necrotic lesions and their subsequent expansion resulted in a steady decrease in the level of phosphatidylglycerol in infected leaves. At the same time, phosphatidic acid and diphosphatidylglycerol contents increased. Leaf zones remote from the regions of necrosis development were also characterized by an increased level of phosphatidic acid. There was a tendency for an increase in phospholipase D activity in both the sites of necrosis development and in the leaf regions remote from these sites. The changes in phosphatidic acid content were of similar nature, and therefore a relative increase in phosphatidic acid could result from the phospholipase D activity. This fact suggests a possible involvement of phospholipase D in the development of the hypersensitive response, and this suggestion is supported by a higher enzyme activity in the leaves of healthy plants of the resistant cultivar as compared to the susceptible one. Causes for the changes in the content of some phospholipids, as well as the physiological role of phospholipase D in the hypersensitive response are discussed.     

Short- and Long-term Effects of High Temperatures (47–49 °C) on Tobacco Leaves. I. Photosynthesis

Detached Nicotiana rustica leaves were exposed for 2 minutes to temperatures in the range 36° to 51°C. Above 45°C, ¹⁴CO₂ fixation was reduced by half as compared with controls. The fall in ¹⁴CO₂ fixation continued for 2 1/2 h. Recovery was completed 45 h after the treatment. Above 45°C there was an increase in the labeled cationic and anionic fractions and a decrease in the neutral fractions, both increase and decrease being associated with the impaired CO₂ fixation. However, these changes in products were also demonstrated when leaves were exposed to the high temperature after labeling.     

Hormonal Aspects of Senescence in Detached Tobacco Leaves

The objective of the present work was to describe the simultaneous changes in endogenous levels of cytokinins, abscisic acid, indoleacetic acid and ethylene in detached, senescing tobacco (Nicotiana rustica L.) leaves. These measurements were related to changes in chlorophyll contents, ¹⁴CO₂ fixation and proline contents — three parameters which have been considered to reflect senescence. Effects of exogenous hormonal treatments on these parameters, as well as on endogenous hormonal levels, provided further evidence for the interrelationships between hormones and for their roles in senescence. Starting with actively growing attached leaves and ending with well-advanced senescence in detached leaves, our data indicate a chronological sequence of three hormonal states: (a) cytokinins — high activity, abscisic acid, auxin and ethylene — low contents (actively growing, attached leaves); (b) cytokinins — low activity, abscisic acid — high, auxin and ethylene — low contents (apparent induction of senescence in detached leaves); and (c) cytokinins and abscisic acid — low, auxin and ethylene — high contents (senescence proper in detached leaves).     

Preconditioning of Tobacco and Bean Leaves to Heat Shock by High Temperature or NaCl

The present paper examines whether leaves subjected to heat or salt pretreatment are better adapted to heat shock than untreated leaves. Such adaptation could result from either acquired thermo-stability and/or better recovery. The rates of metabolic processes such as ¹⁴CO₂ fixation, ¹⁴C-leucine incorporation into the TCA insoluble fraction, ³²P incorporation into phospholipids and ³⁶Cl efflux were used as indicators of adaptability. It was found that pretreatments (heat or salt) protect leaves only slightly against the short-term effects of a heat treatment, but have a pronounced effect on recovery. Cytokinin level was reduced by heat shock, and as in the case of metabolic response, pretreatment had no effect on this reduction.     

Studies on Induced Resistance to Tobacco Mosaic Virus, in Samsun NN Tobacco and Changes in Ribosomal Fractions

Resistance to tobacco mosaic virus (TMV) was activated by various forms of induction in Samsun NN tobacco leaves, and the intensity of the different forms was compared. Induced resistance was highest in leaf tissue between TMV inoculated stripes parallel to the mid-vein and after injection of ethylene maleic anhydride copolymer (EMA), followed by that induced in distal half leaves after inoculating the basal halves with TMV. Resistance in upper leaves following inoculation of the lower leaves with TMV was relatively low, while induction due to lesions caused by ethrel gave an intermediate degree of resistance. Estimation of resistance by size and number of local lesions was correlated with the amount of extractable virus as measured by enzyme-linked immunosorbent assay (ELISA), thus indicating that in the resistant tissue virus replication, and not only the development of necrotic local lesions, is suppressed. An increase in a specific ribosomal fraction (R₂), recovered by a two-step procedure, was observed in tissues where resistance was most intense, i.e., between TMV stripes and after EMA injection. It may be that this specific ribosomal fraction participates in maintaining the resistant state.     

Carbohydrate Accumulation Affects the Redox State of Ascorbate in Detached Tobacco Leaves

A decreased utilization of NADPH for CO₂ fixation as a result of carbohydrate accumulation in chlorotic leaves is generally believed to be associated with an increase in oxidative stress. Molecular oxygen may serve as an alternative electron acceptor of photosynthesis under these conditions. In order to test this hypothesis mature leaves of tobacco plants (Nicotiana tabacum) were detached, fed with glucose (50 mM) via the petiole, and used to study the effect of carbohydrate accumulation on the pigment content and the components of the superoxide dismutase-ascorbate-glutathione cycle. During a period of five days the concentration of total soluble carbohydrates increased substantially in leaves supplied with glucose in comparison with control leaves supplied only with water. This increase was accompanied by a twofold decrease in the chlorophyll content. In detached water-fed leaves the levels of most of the antioxidative components increased, whereas glucose feeding had no or only little additional effect on the activities of the protective enzymes, but caused a 1.6- and 4-fold increase in the contents of glutathione and ascorbate, respectively. In relation to the total foliar ascorbate pool, the amount of reduced ascorbate increased from about 30 % to 60 % upon feeding with glucose. These results do not support the idea that the accumulation carbohydrates per se cause an increased rate of superoxide production which necessitates increased activities of antioxidative enzymes. It rather appears that glucose-fed leaves have an increased reductive capacity that can be released via glutathione into the ascorbate pool, thereby, regulating the redox state of ascorbate.     

Effect of Tobacco Mosaic Virus Infection on Amino Acid Incorporation into Isolated Mitochondria from Tobacco Leaves

Mitochondria isolated from tobacco leaves incorporated ¹⁴C-leucine into the protein and the rate was enhanced by tobacco mosaic virus (TMV) infection as compared with noninfected level. In vitro amino acid incorporation by mitochondria required adenosine triphosphate (ATP), Mg²⁺, and KC1 and the energy sources from oxidative phosphorylation as well as from ATP-generating system. This incorporation was inhibited by ribonuclease (RNase), deoxyribonuclease (DNase), actinomycin D, mitomycin C, puromycin, and chloramphenicol added in the reaction medium. The pretreatment of the mitochondria with DNase and actinomycin D reduced the rate of incorporation. The mitochondria incorporated ³H-guanosine triphosphate (GTP) and this activity was blocked by actinomycin D. The presence in this system of 15,000 g supernatant cell sap fraction or bacterial contamination was carefully checked obtaining a negative result. The reaction product into which ^l4C-amino acids incorporated was solubilized by trypsin. The nature of the amino acid incorporating activity of isolated mitochondria obtained from TMV-infected tobacco leaves is discussed.     

Jasmonic acid treatment to part of the root system is consistent with simulated leaf herbivory, diverting recently assimilated carbon towards untreated roots within an hour

It is known that shoot application of jasmonic acid (JA) leads to an increased carbon export from leaves to stem and roots, and that root treatment with JA inhibits root growth. Using the radioisotope ¹¹C, we measured JA effects on carbon partitioning in sterile, split-root, barley plants. JA applied to one root half reduced carbon partitioning to the JA-treated tissue within minutes, whereas the untreated side showed a corresponding – but slower – increase. This response was not observed when instead of applying JA, the sink strength of one root half was reduced by cooling it: there was no enhanced partitioning to the untreated roots. The slower response in the JA-untreated roots, and the difference between the effect of JA and temperature, suggest that root JA treatment caused transduction of a signal from the treated roots to the shoot, leading to an increase in carbon allocation from the leaves to the untreated root tissue, as was indeed observed 10 min after the shoot application of JA. This supports the hypothesis that the response of some plant species to both leaf and root herbivores may be the diversion of resources to safer locations.     

Induced Resistance in Hypersensitive Tobacco Against Tobacco Mosaic Virus by Injection of Intercellular Fluid from Tobacco Plants with Systemic Acquired Resistance

Acquired resistance in hypersensitive tobacco plants against tobacco mosaic virus (TMV) was induced by components occurring in the intercellular fluid (IV) obtained from virus-infected plants or by plant cell wall components. Induced resistance could be transmitted through seed to the progeny. Lesion size and number were reduced significantly when the progeny was tested by TMV-inoculation. IV was extracted from the upper uninoculated leaves of four times TMV-inoculated Nicotiana tabacum cv. ‘Xanthi’ nc plants. Injection of IV from induction-inoculated plants (SAR-IV) into leaves of healthy plants followed by TMV-infection reduced lesion size significantly. A concentration of 5 × 10^?7 g SAR-IV/ml was still active. IV from healthy plants was inactive. The IV's were partly purified by gel filtration on a Sephadex G-50 column. Some fractions were active in inducing resistance as expressed in reduction of lesion size. Fractions of control-IV were inactive. It is still unknown whether the active substances in SAR-IV are in fact cell wall fragments acting as regulatory molecules in disease resistance.     

Studies on Nitrogen Metabolism in Tobacco Plants A

The effect of age on non-protein constituents of tobacco leaves (N. tabacum L., var. Bright Yellow) has been studied. For this purpose leaves in three different stalk positions, upper, middle and lower, which represent young, mature and over-mature leaves, respectively, were harvested in the day-time and at night.

The total amino nitrogen content both in the day-time and at night decreases from the upper leaf position downwards. As for the individual groups, the content of both upper and middle leaves increases in the day-time and that of the lower ones increases at night.

In general, the content of the individual amino acids is high in the upper leaves and low in the lower ones. Proline and γ-aminobutyric acid, as a ratio of the total amino acid content, show a marked difference with position, in other words with age of the leaves.

The levels of proline decrease very sharply from the upper leaf position downwards and that of γ-aminobutyric acid exhibits an opposite trend in both samples at night and in the day-time. These trends are very prominent in the case of the midribs.

The contents of other amino acids, regardless of position, show similar trends with time to those reported in the previous paper₁₎, and the aspartic acid content increases at night.     

Studies on Nitrogen Metabolism in Tobacco Plants

Δ¹-Pyrroline-5-carboxylate reductase has been considerably purified from tobacco leaves. This enzyme uses NADPH or NADH for the formation of proline, although the former is better used. This enzyme was found in washed chloroplast extract as well as in cytoplasmic fluid and utilized NADPH, formed by the photosynthetic NADP reduction, for the sythesis of proline in the light.