Avirulent Race Research Articles

Mechanism of hypersensitive cell collapse: correlation of increased lipoxygenase activity with membrane damage in leaves of Phaseolus vulgaris (L) inoculated with an avirulent race of Pseudomonas syringae pv. phaseolicola

Increases in lipolytic acyl hydrolase and lipoxygenase isoenzyme activities in leaves of Phaseolus vulgaris (L.) cv. Red Mexican during the incompatible reaction to Pseudomonas syringae pv. phaseolicola race 1 cells coincided with electrolyte leakage characteristic of the early stages of the hypersensitive response. Increase in lipoxygenase activity was inhibited by cycloheximide. Evolution of ethane indicated that membrane-lipid peroxidation had occurred in vivo. Superoxide dismutase (SOD) and peroxidase (POX) activities also increased in the incompatible combination, but at 12 h after inoculation, somewhat later than lipoxygenase. It is possible that SOD and POX were induced as a protective response of the cells against active oxygen species produced as a consequence of lipoxygenase action. The role of lipoxygenase in initiating processes that lead to hypersensitive cell death is discussed.

Chitinase mRNA and enzyme activity in Phaseolus vulgaris (L.) increase more rapidly in response to avirulent than to virulent cells of Pseudomonas syringae pv. phaseolicola

Changes in chitinase mRNA concentration in leaves of French bean cultivar Red Mexican, after inoculation with virulent or avirulent races of Pseudomonas syringae pv. phaseolicola, were estimated by probing Northern and dot blots with radiolabelled cDNA for bean chitinase. In leaves inoculated with the avirulent race 1 isolate of the pathogen, chitinase mRNA levels had begun to increase by 6 h after inoculation. In contrast, chitinase mRNA levels first began to increase between 20 to 24 h after inoculation of leaves with a virulent race 3 isolate of the pathogen. Chitinase mRNA activity was estimated by immunoprecipitation of in vitro translation products using antiserum raised against bean chitinase. Changes in chitinase mRNA activity reflected closely, changes in mRNA concentration. Increases in extractable chitinase enzyme activity occurred 3–6 h after increases in mRNA concentration and activity. As far as we are aware this is the first report of the differential induction of chitinase mRNA and enzyme activity in a host plant by virulent and avirulent races of a homologous bacterial plant pathogen. Heat-killed and UV-killed cells of both avirulent and virulent races of P.s. pv. phaseolicola induced chitinase enzyme activity. This shows that chitinase is induced separately from the hypersensitive reaction and phytoalexin synthesis; both of which are induced only by metabolically active, living bacteria.

Increase of Chalcone Synthase mRNA in Pathogen-Inoculated Soybeans with Race-Specific Resistance Is Different in Leaves and Roots

Soybeans (Glycine max [L.] Merr.) respond to pathogens by producing isoflavonoid-derived phytoalexins. Chalcone synthase (CHS) is the first enzyme of the flavonoid/isoflavonoid biosynthetic pathway. We investigated changes in the steady state levels of CHS mRNA and other specific mRNAs at increasing times after inoculation in two different race-specific interactions, one between leaves and the bacterium Pseudomonas syringae pv glycinea (Psg), and one between roots and the fungus, Phytophthora megasperma f. sp. glycinea (Pmg). The amount of CHS mRNA increases significantly in soybean leaves inoculated with an avirulent race of Psg but not with a virulent race or water. In contrast, the increase in CHS mRNA is similar in roots inoculated with zoospores of either an avirulent or virulent race of Pmg. CHS mRNA increases significantly in pathogen inoculated roots but not in uninoculated controls. Hydroxyproline-rich glycoprotein (HRGP) has been observed by others to increase in wounded or pathogen-inoculated plants. We report here that HRGP mRNA levels are greater in roots inoculated with an avirulent Pmg race than with a virulent race, but inoculation with either race causes a significant increase in HRGP mRNA with respect to controls. Calmodulin or ubiquitin mRNA do not increase in either uninoculated or inoculated roots and leaves. The possibility that race-specific resistance in soybeans is expressed differently in different organs of the plant is discussed.

Effects of heat treatment or mixed inoculation on the development of compatible and incompatible bean rust infections

Dipping leaves bearing 3-day-old infections into water at 50 °C for 25 sec caused the haustoria to shrink, the intercellular hyphae to stop growing, and some host cells to respond, but only at or near infection sites. Responses at all infection sites were synthesis of encasing material around many haustoria and death of a few of the cells which had been penetrated by haustoria or contacted by intercellular hyphae. Another response, seen around potentially incompatible infections only, was death of many cells surrounding fungal colonies; this occurred in a pattern characteristic of the necrosis that would have occurred naturally at a later stage of infection. Mixed inoculations with virulent and avirulent races affected the ways in which different types of infection sites developed, but only where these sites were close together. Necrosis was prevented on borders of avirulent colonies facing nearby virulent colonies, and the avirulent colonies grew more in these positions. The growth of virulent intercellular hyphae appeared to be less near the necrosis of incompatible infections. The results of these experiments are discussed in relation to hypotheses about underlying mechanisms regulating the development of incompatible and compatible interactions.

Local and Systemic Resistance Induced in Watermelons by Formae Speciales ofFusarium oxysporum

Avirulent races of F. o. niveum induced a higher level of resistance than did F. o. cucumerinum. An interval of 24 hr between induction and challenge provided significant protection; a 72-hr interval further enhanced resistance. When roots of the wilt-susceptible watermelon cultivar Black Diamond were induced with F. o. cucumerinum and the leaves inoculated with Colletotrichum lagenarium 24 or 72 hr later, 50% fewer lesions developed on leaves of induced plants than on noninduced inoculated controls. This suggests that induced resistance to F. o. niveum is both local and systemic, as well as nonspecific

Abstracts of papers presented at the annual meeting of the Canadian Phytopathological Society University of British Columbia, Vancouver 14-17 August 1988

Abstracts of papers presented at the annual meeting of the Canadian Phytopathological Society University of British Columbia, Vancouver 14-17 August 1988

Pyramiding Major Genes for Resistance to Maintain Residual Effects

In 1978, R. R. Nelson (26) concluded his chapter in a previous volume of the Annual Review of Phytopathology entitled Genetics of Horizontal Resis­ tance with the advice, Go back young man and gather up your weary and defeated genes of the past, take your currently successful genes, find some new ones if you can, and build yourself a pyramid. Nearly ten years later, it is our privilege to take another look at that counsel. A number of papers have been published that deal with the effects resistance genes confer when in­ oculated with compatible isolates of the pathogen (2, 6, 8, 20-25, 28, 29, 31, 32). Reviews of some of this literature have already appeared (9, 16, 26, 27). Despite this important progress, plant pathology is still not at a point where the notion of pyramiding resistance genes to provide durable resistance to both virulent and avirulent races of a pathogen is widely accepted. We feel that the research that has been accomplished can be divided into three categories. The initial research, much of which has been completed, focused on determining if host genes do confer resistance to isolates of a pathogen that

Comparison of resistance‐inducing abilities of virulent and avirulent races of Erysiphe graminis f.sp. hordei and a race of Erysiphe graminis f.sp. tritici in barley

Resistance to powdery mildew was induced in barley by preinoculation with virulent and avirulent races of barley powdery mildew (Erysiphe graminis f.sp. hordei), and with a race of wheat powdery mildew (E. graminis f.sp. tritici). Four inducer densities were tested in 13 different induction periods between 1 and 24 h. Generally, the resistance induced by barley powdery mildew increased up to 10‐12 h of induction and was maintained in longer induction periods. The inducing abilities of virulent and avirulent races could not be distinguished up to 10‐12 h of induction, after which the inducing ability of avirulent races increased significantly in relation to virulent races. Wheat powdery mildew was able to induce more resistance than barley powdery mildew in induction periods up to 8 h. In a single inoculation procedure the number of haustoria developing from virulent barley powdery mildew decreased as inoculum density increased. The effect was ascribed to induction of resistance. This reduction of infection efficiency in the compatible interaction was compared to induced resistance. However, the inoculum density needed for 50% resistance induction in the double inoculation procedure was approximately 40 times higher than the inoculum density needed for 50% reduction in infection efficiency in the single inoculation procedure.

Host responses to fungal penetration in Evysiphe graminis f.sp. hordei infections in barley

Resistance mechanisms restricting penetration and establishment were investigated in an incompatible interaction using an avirulent race CC/3 (BMV 1 + 4) of Erysiphe graminis f.sp hordei on barley cv. Athos (BMR 2 + 5) and a compatible interaction using the universal susceptible cv. Golden Promise (BMR 0). In both interactions: (i) auto‐fluorescence of the host cell wall occurred adjacent to the primary germ tube tip after 4 h, and by 10 h near to the appressorial germ tube; (ii) strongly fluorescing papillae developed after 12 h, being frequently associated with failed penetrations; and (iii) only 30% of the attempted penetrations from appressorial lobes resulted in an incipient haustorium at 16 h. Hypersensitive reactions occurred in cells with an incipient haustorium in 30% of appressorial penetrations on Athos, but only in 4% of those on Golden Promise. Twenty per cent of hypersensitively reacting cells in Athos appeared dead by 14 h using trypan blue (membrane exclusion test), compared with 5% for neutral red, suggesting that plasma membrane damage is an early event in the hypersensitive response. Haustorial death was associated with host cell death, but did not precede it; appressorial death occurred 2‐4 h after that of the host cell.

Arachidonic and Linoleic Acids Elicit Isoflavonoid Phytoalexin Accumulation in Phaseolus vulgaris (French bean)

AbstractArachidonic and linoleic, but not α‐linolenic or γ‐linolenic acids, induced necrosis and the accumulation of isoflavonoid phytoalexins in leaves of French bean (Phaseolus vulgaris L.) cv. ‘Red Mexican'. The levels of phytoalexins which accumulated were similar to those found in an incompatible interaction between cv. ‘Red Mexican’and an avirulent race 1 isolate of Pseudomonas syringae pv. phaseolicola. Maximum amounts of phytoalexins accumulated in response to 1.6 mM fattyacids, as opposed to 3.3 or 0.6 mM. The most severe tissue necrosis was not accompanied by the highest levels of phytoalexin accumulation. In contrast to leaves, only trace amounts of phytoalexins were elicited in hypocotyls. The possible significance of, these results is discussed in terms of mechanisms leading to plant cell death and phytoalexin accumulation.

Transient Induction of Phenylalanine Ammonia-Lyase and 4-Coumarate: CoA Ligase mRNAs in Potato Leaves Infected with Virulent or Avirulent Races of Phytophthora infestans

Infection of potato leaves with the fungal pathogen Phytophthora infestans (Pi) resulted in the rapid stimulation of phenylpropanoid metabolism. Increases in the activities of several mRNAs, including those encoding phenylalanine ammonia-lyase (PAL) and 4-coumarate:CoA ligase (4CL), were detectable within a few hours postinoculation, as demonstrated by two-dimensional gel electrophoresis of proteins synthesized in vitro. This effect was closely mimicked by application of Pi culture filtrate through cut leaf stems. PAL and 4CL mRNA activities were also rapidly and transiently induced in potato cell suspension cultures by treatments with Pi culture filtrate or arachidonic acid. This induction was exploited to generate cDNA probes complementary to PAL and 4CL mRNAs. Blot hybridizations using these probes revealed almost immediate, transient and coordinate increases in the transcription rates and subsequent changes in the amounts of PAL and 4CL mRNAs in leaves treated with Pi culture filtrate. Similar changes in the mRNA amounts were found in infected leaves of potato cultivars carrying resistance genes R1 (cv Datura) or R4 (cv Isola), independent of whether a virulent or an avirulent Pi pathotype was used for inoculation. These results are discussed in relation to recent cytological observations with the same potato cultivars and Pi pathotypes.

Induction of defence reactions in plants

Induced local resistance presumably involves the same mechanisms in the plants as resistance elicited during normal plant-pathogen interactions. In many cases resistance elicitors from pathogens have been found to be non-specific, i.e. unrelated to race-cultivar specificity. Thus, existence of specific resistance suppressors has been suggested to make the virulent races able to infect. In other cases specific resistance elicitors have been indicated to exist in a virulent races, by which the race specific resistance may be accomplished. At our Department resistance has been induced in the barley—powdery mildew interaction by use of double inoculation procedures. Both virulent and avirulent races of barley powdery mildew can induce resistance, but avirulent races show an increased resistance induction ability in relation to virulent races from 12 hours after inoculation. In barley plants wheat powdery mildew induced more resistance than barley powdery mildew 1 to 8 hours after inoculation. Induced resistance was mainly localized to the epidermal cells attacked by the inducer, but an effect was also present in the surrounding epidermal cells. The energetic consequences of resistance in barley to barley powdery mildew have been found to be reflected in an increased respiratory rate at the time of infection attempt. Further, these energy costs appeared to reduce grain yield by 7 %. The expression of resistance in barley is thought to involve de novo synthesis mRNAs and proteins, which makes it possible to apply gene technological methods to study induced resistance. Research of this kind is in progress at our Department, which hopefully will give information on the mechanisms of resistance triggering and resistance expression.

Zur Auswirkung von Resistenzreaktionen auf Pflanzenwachstum und Ertrag in inkompatiblen Wirt‐Parasit‐Systemen

Abstract Effects of defense reactions on plant growth and yield in incompatible host‐parasite‐systems Investigations in incompatible host‐parasite‐systems (barley –Erysiphe graminis and lettuce –Bremia lactucae) were carried out to prove, whether the induction of defense reactions leads to measurable effects on the yield performance of plants. Permanent or frequent inoculations with avirulent races decreased yield of barley and lettuce remarkably, expressed in reduced fresh weight and drym, atter. Less grains were produced and their contents of carbohydrates and amino acids were diminished significantly. In young plants a reduction in sugar content was evident already after a single inoculation. In barley as in lettuce this effect was shown, not only for inoculated, but also for plant parts not present at the time of inoculation. There, in addition especially aromatic amino acids increased, while glutamate, aspartate and their amides diminished. The consequences of a permanent maintenance of postinfectional resistance mechanisms either of a hypersensitive or non‐hypersensitive reaction type for growth and yield performance are discussed with regard to an increased consumption of energy and carbohydrates.

Changes in gene activity during expression of the hypersensitive response in Phaseolus vulgaris cv. Red Mexican to an avirulent race 1 isolate of Pseudomonas syringae pv. phaseolicola

Total cellular and polysomal RNAs were isolated from leaves of the French bean cultivar Red Mexican which had been inoculated with virulent or avirulent races of Pseudomonas syringae pv. phaseolicola . The mRNAs were translated in vitro and the polypeptide fingerprints obtained from the various treatments indicated specific changes in host mRNA activities in leaf tissue expressing a hypersensitive reaction (HR).The mRNAs coded for several high M r polypeptides, and coordinated decreases and transient increases in activity were observed, defining a cascade of changes in gene expression in tissues undergoing the HR, Some changes were apparent as early as 2 h after inoculation. Some mRNA activities which decreased rapidly during the HR also decreased in the susceptible reaction but at a slower rate. The possible function of some of the polypeptides is discussed.

Protein synthesis in flax following inoculation with flax rust

Resistance to flax rust Melampsora lini (Ehrenb.) Lév. in flax carrying the N resistance gene is determined by 24 h postinoculation, at which time hypersensitivity is observed. We have examined protein synthesis in cotyledons inoculated with both virulent and avirulent races of rust by in vivo labelling with [35S]methionine. The pattern of protein synthesis was assessed by one- and two-dimensional electrophoresis 8, 13, and 18 h after inoculation. No changes in protein synthesis were observed in the first 14 h following inoculation; however, by 18 h after inoculation the susceptible combination showed a marked decrease in protein synthesis (22%; P = 0.01). This could be largely accounted for by the reduced synthesis of the ribulose 1,5-bisphosphate carboxylase large subunit, which was readily quantified on electrophoresis gels. In addition, a 30-kDa polypeptide also declined in the susceptible combination. Two-dimensional electrophoresis enabled changes to be detected in the synthesis of other minor polypeptides. None of these changes were observed in the resistant combination in which a small increase in the synthesis of the ribulose 1,5-bisphosphate carboxylase large subunit and the 30-kDa polypeptide was found. These results indicate that the outcome of the host–pathogen interaction has already been determined by 18 h after inoculation.

Response of tomato to leaf injection with conidia of virulent and avirulent races of Cladosporium fulvum

Injection of tomato leaflets of various cultivars with conidia of various races of Cladosporium fulvum produced the expected disease reactions. In all interactions, histological study of leaflets sampled up to 6 or 8 days post-inoculation revealed localized development of wal appositions (composed of aniline blue fluorescent material, most likely callose) in those host cells in direct contact with a conidium or a young germ tube. In compatible interactions (Potentate, no known Cf gene, with race 1, 7 or 12; Vinequeen, Cf genes 2 and 4, with race 12) and a delayed incompatible interaction (Stirling Castle, gene Cf 1, with race 7), germ tubes eliciting wall apposition formation often continued to develop, usually by producing a new branch which failed to elicit a visible host response. In incompatible interactions (Vinequeen with race 1 or 7), germ tubes either did not develop further on encountering the wall apposition response or they evoked a response in the next cells contacted and ceased growth. In vitro, the outer surface of germ tubes was readily stained with fluorescein labelled concanavalin A, supporting the possibility that previously described non-specific glycopeptide elicitor molecules are localized on the surface. This leaf-injection method may be useful in studies concerning the molecular interactions of C. fulvum and tomato.

Origin and Phylogenetical Differentiation of Wild Tetraploid Wheats Inferred from Their Susceptibility to Leaf Rust (Preliminary Note) : Susceptibility of the Genus Triticum to Two Races of Puccinia recondita f.sp. tritici

To study the relationship between the origin and phylogenetic differentiation of the tetraploid wheats and their susceptibility to leaf rust, a model experiment has been carried out. Namely, using 8 species which had played an important role on the origin and evolution of the genus Triticum, the seedling reactions of them to 2 races of Puccinia recondita Rob. ex Desm. f.sp, tritici were examined. In the consequence of the test, Tritcium monococcum belonging to the Einkorn. group, l strain of T. dicoccoides belonging to the Emmer group, T. araraticum and T. tinoheevi belonging to the Timopheevi group and 2 strains of T. spelta belonging to the Dinkel group were resistant, T. aegilopoides belonging to the Einkorn group, 1 strain of T. durum belonging to the Emmer group, 1 strain of T. spelta and T. araraticum were susceptible, and 2 strains of T. dicoccoides and 2 strains of T. durumshowed various types of responses to avilulent race 1 B. On the other hand, T. monococcum and T. timopheevi were resistant, 2 strains of T. durum and 1 strain of T. spelta showed several types of reactions, respectively, but other wheat strains were sesceptible to virulent race 21 B .Three wild species except T. aegilopoides showed different reactions to 2 races. Namely they were susceptible to race 21 B in all cases, change of the reactions was observed. As a whole, the tendency to show unstable responses to avirulent race 1 B, but stable ones to virulent race 21 B was recognized.

Differential agglutination of races 1 and 2 of Pseudomonas phaseolicola by a fraction from cotyledons of Phaseolus vulgaris cv. Red Mexican

A fraction obtained from cotyledons of Phaseolus vulgaris cv. Red Mexican agglutinated cells of avirulent Race 1 isolates of Pseudomonas phaseolicola more actively than it agglutinated cells of virulent Race 2 isolates; it also agglutinated human erythrocytes. Carbohydrate in the fraction agglutinated the bacteria whereas haemagglutination was caused by protein. A fraction with similar properties could not be obtained from cotyledons of cv. Canadian Wonder, susceptible to Races 1 and 2. Agglutination of bacteria was assayed by a modification of a standard microhaemagglutination assay.

イネおよびシコクビエ葉身のいもち病り病性病はん部における非病原性イネいもち病菌レースの増殖

Leaves of rice variety Aichiasahi were previously inoculated with virulent race 003 or 303 of rice blast fungus and those of Kanto 51 were inoculated with virulent race 031. Leaves of Ragi (finger millet) were previously inoculated with virulent ragi blast fungus. After development of lesions, the infected leaves of Aichiasahi, Kanto 51 and Ragi were reinoculated with avirulent race 031, 003 and 001 of rice blast fungus respectively. A few conidia of avirulent challenge races 031, 003 or 001 were produced on lesions of Aichiasahi, Kanto 51 and Ragi leaves 7 or 10 days after reinoculation.

Responses of wheat cells around heat-inhibited rust mycelia and associated with the expression of the Lr20, Sr6 and Sr15 alleles for resistance

Mycelia of avirulent and virulent races of Puccinia recondita and P. graminis were established in leaves of different wheat cultivars at 30 °C where the Lr20, Sr6 and Sr15 alleles are not expressed. Leaves were then dipped in water at 50 °C for 15 s, which prevented further mycelial growth. Plants were then held at 20 °C for several days. Two different types of symptom appeared at infection sites, distinguishing the expression of the Lr20 and Sr15 alleles from that of the Sr6 allele. White sunken lesions, containing masses of mesophyll cells with collapsed protoplasts, developed around mycelia avirulent with respect to the Lr20 and Sr15 alleles. Yellow-brown lesions, containing some necrotic mesophyll cells in contact with hyphae, developed at the sites of mycelia avirulent with respect to the Sr6 allele. Similar yellow-brown lesions developed at the sites of mycelia virulent with respect to each allele. The cause of the white sunken lesions is discussed in relation to the action of hypothetical, diffusible and gene-specific fungal products. A different cause is suggested for the yellow-brown lesions.