Effects of phenylpropanoid and energetic metabolism inhibition on faba bean resistance mechanisms to rust

ABSTRACT Effects on penetration and hypersensitive resistance of the cinnamyl acid dehydrogenase (CAD) suicide inhibitor ([(2-hydroxyphenyl) amino] sulphinyl) acetic acid, 1.1 dimethyl ester, which suppresses phenylpro-panoid biosynthesis, and of D-mannose, which sequesters phosphate and reduces energy available in host cells, were studied in faba bean (Vicia faba) genotypes with differing resistance mechanisms to faba bean rust (Uromyces viciae-fabae). Inhibition of CAD reduced penetration resistance in lines 2N-34, 2N-52, V-1271, and V-1272, revealing an important role for phenylpropanoid biosynthesis in the resistance of these lines. Inhibition of CAD also inhibited hypersensitive cell death in these lines. D-mannose had little or no effect on resistance. By contrast, CAD inhibition did not affect penetration resistance of line BPL-261, which has a high degree of penetration resistance not associated with hypersensitive cell death. In BPL-261, D-mannose inhibited penetration resistance. The parallelism between the faba bean genotype responses to rust observed here and the response of barley genotypes with differing resistance mechanisms to powdery mildew after similar inhibitor treatments is analyzed and discussed.     

Cytological and pharmacological evidence that biotrophic fungi trigger different cell death execution processes in host and nonhost cells during the hypersensitive response

Living epidermal cells undergoing a hypersensitive response to penetration by a rust or a powdery mildew fungus exhibited two distinct patterns of cellular changes that occurred from the time of the cessation of cytoplasmic streaming to protoplast collapse. In nonhost cells, this death process was completed in less than 1 h, trans-vacuolar strands initially remained visible after streaming stopped, Brownian-like motion was seen in particulate cytoplasmic components, and the plant nucleus was usually associated with the intracellular fungus. In resistant host cells, trans-vacuolar strands disappeared as cytoplasmic streaming stopped, no motion was seen in the cytoplasm, there was no consistent association of the plant nucleus with the fungus, and the death process took 2 h or more. Neither type of cell death was mimicked by the application of CuCl₂, salicylic acid, or KCN, nor did the individually distinctive morphological features of the cell deaths that they triggered change with chemical concentration. Pharmacological studies suggested that host and nonhost cell death required an influx of extracellular calcium, protein kinase activity and protein synthesis, but not the generation of extracellular reactive oxygen species. The application of peptides with an RGD motif delayed cell death in only one resistant host, and inhibitors of caspase activity delayed cell death only in the nonhost combinations. These and other results suggest that some rust- and powdery mildew-specific resistance genes in host cultivars may control a different cell dismantling process during the hypersensitive response than that triggered in nonhost plants.     

Compatible Puccinia hordei infection in barley induces basal defense to subsequent infection by Blumeria graminis

Rusts and powdery mildews employ different strategies to suppress defense during penetration. We observed that a compatible interaction of barley-Puccinia hordei induced increased penetration resistance to a challenge infection by powdery mildew. This induced resistance is local and its level is not determined by the virulence spectrum of the challenger isolate. Our data suggest that the inducer effect is due to rust-stoma communication during penetration, to the presence of the rust hyphae in the apoplast, or to penetration resistance mounted by the rust attacked mesophyll cells. We hypothesized that the rust “primes” the basal defense prior to the mildew infection.     

Nonhost versus host resistance to the grapevine downy mildew, Plasmopara viticola, studied at the tissue level

Following inoculation of host and nonhost plants with Plasmopara viticola, the grapevine downy mildew, a histological survey was undertaken to identify the stage where its development is contained in nonhosts and in resistant host plants. Three herbaceous nonhost species, Beta vulgaris, Lactuca sativa, and Capsicum annuum, and three grapevine species displaying different level of resistance (Vitis vinifera [susceptible], Vitis riparia [partially resistant] and Muscadinia rotundifolia [totally resistant]) where inoculated by P. viticola using a controlled leaf disk inoculation bioassay. During the early steps of infection, defined as encystment of zoospores on stomata, penetration of the germ tube, and production of the vesicle with the primary hypha, there was no evidence of a clear-cut preference to grapevine tissues that could attest to host specificity. The main difference between host grapevine species and nonhosts was observed during the haustorium formation stage. In nonhost tissues, the infection was stopped by cell wall-associated defense responses before any mature haustorium could appear. Defense responses in resistant grapevines were triggered when haustoria were fully visible and corresponded to hypersensitive responses. These observations illustrate that, for P. viticola, haustorium formation is not only a key stage for the establishment of biotrophy but also for the host specificity and the recognition by grapevine resistance factors.     

Primary Haustorial Development of Striga asiatica on Host and Nonhost Species

ABSTRACT Initial interactions of Striga asiatica with a susceptible host and non-host plants were examined by histological methods. Haustorial development was initiated when radicles of S. asiatica were placed in contact with host or nonhost roots. Reorganization of the S. asiatica root apical meristem was rapid and involved the formation of a distal group of cells that penetrated the host or nonhost root. Penetration of the epidermis of the host (sorghum) roots and advance into the cortex occurred within 24 to 48 h of inoculation. Penetration of the endodermis by the developing endophyte was delayed for 72 to 96 h after initial contact. However, upon penetration vascular continuity was established between parasite and host. In contrast, interactions with nonhosts provided evidence of active resistance mechanisms. Penetration of lettuce, marigold, and cowpea roots by S. asiatica was most frequently arrested in the cortex, and endophytic cells were necrotic 72 h after inoculation. Some species-specific differences were observed in the reactions of nonhosts to penetration, although in their general nature the interactions with S. asiatica were similar.     

Local and systemic effects of oxylipins on powdery mildew infection in barley

Treatment of the first leaves of barley seedlings with the oxylipin colneleic acid, or the two trihydroxy oxylipins 9,12,13-trihydroxy-11(E)-octadecenoic acid and 9,12,13-trihydroxy-10(E)-octadecenoic acid, reduced infection of that leaf by the powdery mildew fungus Blumeria graminis Speer f sp hordei Marchal. When applied to first leaves, etheroleic acid and colneleic acid, as well as the trihydroxy oxylipin 9,12,13-trihydroxy-10(E),15(Z)-octadecadienoic acid, also reduced mildew infection in second leaves. In all cases where local and systemic effects against mildew were observed, activity of the defence-related enzyme phenylalanine ammonia lyase (PAL) was increased, but only following challenge inoculation with powdery mildew. Peroxidase activity was not affected by oxylipin treatment or mildew inoculation. Whether the effects observed were due to the oxylipins or to breakdown products is not known, since no information is available on the stability of these particular oxylipins on leaf surfaces. Nevertheless, these data represent the first report of systemic effects against pathogen infection following pre-treatment with oxylipins.     

Genes Governing Resistance to Puccinia hordei in Thirteen Spring Barley Accessions

ABSTRACT Leaf rust, caused by Puccinia hordei, is an important disease of barley in many parts of the world. In the eastern United States, this disease was effectively controlled for over 20 years through the deployment of cultivars carrying the resistance gene Rph7. Isolates of P. hordei with virulence for Rph7 appeared in this region in the early 1990s rendering barley cultivars with this gene vulnerable to leaf rust infection. From a preliminary evaluation test, 13 accessions from diverse geographic locations possessed resistance to P. hordei isolate VA90-34, which has virulence for genes Rph1, 2, 4, 6, 7, 8, and 11. Each of these 13 accessions was crossed with susceptible cvs. Moore or Larker to characterize gene number and gene action for resistance to P. hordei. Additionally, the 13 accessions were intercrossed and crossed to host differential lines possessing genes Rph3, Rph5, and Rph9 to determine allelic relationships of resistance genes. Seedlings of F(1), F(2), and BC(1)F(1) populations were evaluated in the greenhouse for their reaction to P. hordei isolate VA90-34. Leaf rust resistance in six of the accessions including Collo sib, CR270.3.2, Deir Alla 105, Giza 119, Gloria, and Lenka is governed by a single dominant gene located at or near the Rph3 locus. All accessions for which the gene Rph3 was postulated to govern leaf rust resistance, except for Deir Alla 105, likely possess an allele different than Rph3.c found in Estate based on the differential reaction to isolates of P. hordei. The resistance gene in Grit and Donan is located at or near the Rph9 locus. Alleles at both the Rph3 and Rph9 loci confer resistance in Femina and Dorina. In addition to Rph3, Caroline and CR366.13.2 likely possess a second unknown recessive gene for leaf rust resistance. Resistance in Carre 180 is governed by a recessive gene that is different from all other genes considered in this study. Identification of both known and unique genes conferring leaf rust resistance in the barley germplasm included in this study provides breeding programs with the knowledge and opportunity to assess currently used sources of leaf rust resistance and to incorporate new sources of resistance into their programs.     

Specificity of Prehaustorial Resistance to Puccinia hordei and to Two Inappropriate Rust Fungi in Barley

ABSTRACT To elucidate the specificity of prehaustorial resistance to inappropriate rust fungi, we studied two populations of recombinant inbred lines of barley that segregated for partial resistance (PR) to Puccinia hordei and for the resistance to the inappropriate rust species P. recondita f. sp. tritici and P. hordei-murini. PR to P. hordei is prehaustorial and nonhypersensitive, and its level can be assessed accurately by measuring the latent period of the fungus. The resistance to the inappropriate rust species is a combination of prehaustorial (nonhypersensitive) and posthaustorial (hypersensitive) mechanisms. The amount of nonhypersensitive, early abortion of P. recondita f. sp. tritici and P. hordei-murini sporelings reflects the degree of prehaustorial defense to the two inappropriate rust species. All lines showing a long latent period of P. hordei also had a relatively high level of early abortion of the growth of P. recondita f. sp. tritici and P. hordei-murini. This indicates that genes for PR to P. hordei are also effective against these two inappropriate rust species. The reverse was not necessarily true; some lines showing a high level of early abortion of P. recondita f. sp. tritici and P. hordei-murini had a low level of PR to P. hordei. Moreover, lines with a similar level of prehaustorial resistance to P. recondita f. sp. tritici could differ considerably in their prehaustorial resistance to P. hordei-murini. This indicates that genes for prehaustorial resistance may exhibit rust species specificity.     

Genetic analysis and molecular mapping of wheat genes conferring resistance to the wheat stripe rust and barley stripe rust pathogens

ABSTRACT Stripe rust is one of the most important diseases of wheat and barley worldwide. On wheat it is caused by Puccinia striiformis f. sp. tritici and on barley by P. striiformis f. sp. hordei Most wheat genotypes are resistant to P. striiformis f. sp. hordei and most barley genotypes are resistant to P. striiformis f. sp. tritici. To determine the genetics of resistance in wheat to P. striiformis f. sp. hordei, crosses were made between wheat genotypes Lemhi (resistant to P. striiformis f. sp. hordei) and PI 478214 (susceptible to P. striiformis f. sp. hordei). The greenhouse seedling test of 150 F(2) progeny from the Lemhi x PI 478214 cross, inoculated with race PSH-14 of P. striiformis f. sp. hordei, indicated that Lemhi has a dominant resistance gene. The single dominant gene was confirmed by testing seedlings of the F(1), BC(1) to the two parents, and 150 F(3) lines from the F(2) plants with the same race. The tests of the F(1), BC(1), and F(3) progeny with race PSH-48 of P. striiformis f. sp. hordei and PST-21 of P. striiformis f. sp. tritici also showed a dominant gene for resistance to these races. Cosegregation analyses of the F(3) data from the tests with the two races of P. striiformis f. sp. hordei and one race of P. striiformis f. sp. tritici suggested that the same gene conferred the resistance to both races of P. striiformis f. sp. hordei, and this gene was different but closely linked to Yr21, a previously reported gene in Lemhi conferring resistance to race PST-21 of P. striiformis f. sp. tritici. A linkage group consisting of 11 resistance gene analog polymorphism (RGAP) markers was established for the genes. The gene was confirmed to be on chromosome 1B by amplification of a set of nullitetrasomic Chinese Spring lines with an RGAP marker linked in repulsion with the resistance allele. The genetic information obtained from this study is useful in understanding interactions between inappropriate hosts and pathogens. The gene identified in Lemhi for resistance to P. striiformis f. sp. hordei should provide resistance to barley stripe rust when introgressed into barley cultivars.     

The importance of non‐penetrated papillae formation in the resistance response of triticale to powdery mildew (Blumeria graminis)

Triticale is the intergeneric hybrid between wheat and rye. With the expansion of the triticale growing area, powdery mildew has emerged and become a significant disease on this new host. Recent research demonstrated that this ‘new’ powdery mildew on triticale has emerged through a host range expansion of powdery mildew of wheat. Moreover, isolates sampled from triticale still infect their previous host, wheat, but isolates sampled from wheat hardly infect triticale. Race‐specific and adult‐plant resistance have been identified in triticale cultivars. The main objective of this study was to characterize the cellular basis of powdery mildew resistance in triticale. Commonalities with resistance responses in other cereals such as wheat, barley and oat are discussed. A detailed comparative histological study of various resistance responses during cross‐inoculation of either virulent or avirulent wheat and triticale isolates on both hosts was carried out. The present data provide evidence that for incompatible interactions, the formation of non‐penetrated papillae is the predominant resistance response, while the hypersensitive response (HR) acts as a second line of defence, to cut the fungus off from nutrients, if penetration resistance fails. It is not clear yet what causes the slower growth and reduced colony size of triticale isolates when inoculated on wheat. Possibly, post‐penetration resistance mechanisms, other than HR, are switched on during these (semi‐) compatible interactions. Molecular studies on gene expression and gene function of defence‐related genes might reveal further insights into the genetic basis of these resistance responses.     

Genetics of resistance to wheat leaf rust, stem rust, and powdery mildew in Aegilops sharonensis

Aegilops sharonensis (Sharon goatgrass) is a wild relative of wheat and a rich source of genetic diversity for disease resistance. The objectives of this study were to determine the genetic basis of leaf rust, stem rust, and powdery mildew resistance in A. sharonensis and also the allelic relationships between genes controlling resistance to each disease. Progeny from crosses between resistant and susceptible accessions were evaluated for their disease reaction at the seedling and/or adult plant stage to determine the number and action of genes conferring resistance. Two different genes conferring resistance to leaf rust races THBJ and BBBB were identified in accessions 1644 and 603. For stem rust, the same single gene was found to confer resistance to race TTTT in accessions 1644 and 2229. Resistance to stem rust race TPMK was conferred by two genes in accessions 1644 and 603. A contingency test revealed no association between genes conferring resistance to leaf rust race THBJ and stem rust race TTTT or between genes conferring resistance to stem rust race TTTT and powdery mildew isolate UM06-01, indicating that the respective resistance genes are not linked. Three accessions (1644, 2229, and 1193) were found to carry a single gene for resistance to powdery mildew. Allelism tests revealed that the resistance gene in accession 1644 is different from the respective single genes present in either 2229 or 1193. The simple inheritance of leaf rust, stem rust, and powdery mildew resistance in A. sharonensis should simplify the transfer of resistance to wheat in wide crosses.     

Adult plant resistance to powdery mildew (Erysiphe graminis)in three barley cultivars

The mechanism(s) of adult plant resistance (a.p.r.) to Erysiphe graminis f. sp. hordei were examined in two spring barley cultivars, Athos and Porthos, which possess similar identified mildew resistance genes and seedling resistance, but differential field resistance. These cultivars were compared with Golden Promise, a universally susceptible cultivar. Differential a.p.r. to a compatible race was detected as a reduction in the number and sporulation of colonies on later-formed Athos leaves. Golden Promise exhibited some a.p.r., but much less than either Athos or Porthos. Differential a.p.r. was expressed at several infection stages, viz. appressorial formation, penetration and sporulation. Epidermal hypersensitivity occurred infrequently in all leaf position/cultivar interactions. Papillae were associated with >90% of all penetration failures. The incidence of papillae in successful penetrations at 96 h was 20% on later-formed leaves of Athos compared to c. 60% on all other leaves.     

Saccharin-induced protection against powdery mildew in barley: effects on growth and phenylpropanoid metabolism

Treatment of barley ( Hordeum vulgare ) with 3 m m saccharin, applied as a foliar treatment to the first leaf or as a soil drench, provided significant control of powdery mildew ( Blumeria graminis f.sp. hordei ) on first and second leaves. This was unlikely to be the result of a direct effect of saccharin on the fungus, as application of the chemical to first leaves 2 h before inoculation did not affect conidial germination or formation of appressoria. Saccharin treatment had no significant effect on plant growth, except for a reduction in total leaf area in plants treated with a saccharin drench 14 days before inoculation with mildew. Phenylalanine ammonia-lyase activity was reduced significantly in second leaves 18 and 48 h after inoculation in plants treated with saccharin 14 days earlier. Peroxidase activity increased significantly in plants challenged with mildew within 6 days of saccharin application, although changes were not apparent until 48 h after pathogen challenge. On these occasions, treatment with saccharin resulted in a 33% increase in peroxidase activity compared with controls. In plants inoculated 10 or 14 days after saccharin application, cinnamyl alcohol dehydrogenase (CAD) activity increased prior to, and 18, 24 and 48 h after, inoculation of the barley plants with mildew. CAD activity increased approximately twofold compared with controls. However, in contrast to peroxidase, CAD activity was significantly higher in saccharin-treated plants prior to and after inoculation with powdery mildew, suggesting that saccharin primes CAD activity prior to pathogen challenge.     

Potassium phosphate induces systemic protection in barley to powdery mildew infection

In laboratory tests, treatment of the first leaves of barley (Hordeum vulgare L cv Golden Promise) with potassium phosphate led to significant reduction in infection of the second leaves with the powdery mildew fungus Blumeria graminis f sp hordei Marchal, with a 25 mM treatment giving 89% reduction in infection. Although the optimal interval between phosphate treatment of the first leaves and mildew inoculation of the second leaves was 2 days, significant protection was still obtained if the interval was increased to 12 days. Protection against powdery mildew infection was not as effective when the potassium phosphate was applied as a seed treatment or root drench. Phosphate treatment of the first leaves led to significant increases in activities of phenylalanine ammonia lyase (PAL), peroxidase and lipoxygenase in second leaves. Enzyme activities, especially PAL and peroxidase, were increased further when second leaves of phosphate-treated plants were inoculated with powdery mildew. Phosphate treatment of the first leaves did not adversely affect plant growth and, in a field trial, 25 mM potassium phosphate provided 70% control of mildew and gave a small increase in grain yield.     

微管骨架在辣椒-黄瓜炭疽病菌非寄主互作中的作用

 以辣椒“A11”和黄瓜炭疽病菌（Colletotrichum orbiculare）组成的非寄主互作体系为研究对象,利用组织化学方法研究了辣椒叶片微管骨架解聚后接种黄瓜炭疽病菌,对抗病反应相关的乳突、H₂O₂和过敏性坏死反应的影响以及黄瓜炭疽病菌在非寄主辣椒上的侵入状况,揭示微管骨架在辣椒-黄瓜炭疽病菌非寄主互作中的作用。结果表明,微管骨架解聚显著抑制了乳突的形成,H₂O₂的积累也明显减弱,使黄瓜炭疽病菌能成功侵入辣椒细胞,表明微管骨架参与了辣椒的非寄主抗病性反应,微管的聚合在辣椒非寄主抗病性中具有重要的作用。     

A Compromised Mlo Pathway Affects the Response of Barley to the Necrotrophic Fungus Bipolaris sorokiniana (Teleomorph: Cochliobolus sativus) and Its Toxins

ABSTRACT In search of new durable disease resistance traits in barley to control leaf spot blotch disease caused by the necrotrophic fungus Bipolaris sorokiniana (teleomorph: Cochliobolus sativus), we developed macroscopic and microscopic scales to judge spot blotch disease development on barley. Infection of barley was associated with cell wall penetration and accumulation of hydrogen peroxide. The latter appeared to take place in cell wall swellings under fungal penetration attempts as well as during cell death provoked by the necrotrophic pathogen. Additionally, we tested the influence of a compromised Mlo pathway that confers broad resistance against powdery mildew fungus (Blumeria graminis f. sp. hordei). Powdery mildew-resistant genotypes with mutations at the Mlo locus (mlo genotypes) showed a higher sensitivity to infiltration of toxic culture filtrate of Bipolaris sorokiniana as compared with wild-type barley. Mutants defective in Ror, a gene required for mlo-specified powdery mildew resistance, were also more sensitive to Bipolaris sorokiniana toxins than wild-type barley but showed less symptoms than mlo5 parents. Fungal culture filtrates induced an H2O2 burst in all mutants, whereas wild-type (Mlo) barley was less sensitive. The results support the hypothesis that the barley Mlo gene product functions as a suppresser of cell death. Therefore, a compromised Mlo pathway is effective for control of biotrophic powdery mildew fungus but not for necrotrophic Bipolaris sorokiniana. We discuss the problem of finding resistance traits that are effective against both biotrophic and necrotrophic pathogens with emphasis on the role of the anti-oxidative system of plant cells.     

Temporary partial breakdown of Mlo-resistance in spring barley by the sudden relief of soil water stress

Seedlings of spring barley cultivars expressing mlo genes for resistance to mildew (Erysiphe graminis f.sp. hordei) in different genetic backgrounds were subjected to degrees of water stress under different nutrient and soil conditions. The temporary breakdown in the host of the expression of Mlo-resistance, reported occasionally under field conditions, was successfully simulated when stress treatments were relieved. Differences in the degree of temporary breakdown were attributable to nutrient status, soil compaction and genetic background of the mlo gene.