A mutagenesis-derived broad-spectrum disease resistance locus in wheat

Wheat leaf rust, stem rust, stripe rust, and powdery mildew caused by the fungal pathogens Puccinia triticina, P. graminis f. sp. tritici, P. striiformis f. sp. tritici, and Blumeria graminis f. sp. tritici, respectively, are destructive diseases of wheat worldwide. Breeding durable disease resistance cultivars rely largely on continually introgressing new resistance genes, especially the genes with different defense mechanisms, into adapted varieties. Here, we describe a new resistance gene obtained by mutagenesis. The mutant, MNR220 (mutagenesis-derived new resistance), enhances resistance to three rusts and powdery mildew, with the characteristics of delayed disease development at the seedling stage and completed resistance at the adult plant stage. Genetic analysis demonstrated that the resistance in MNR220 is conferred by a single semidominant gene mapped on the short arm of chromosome 2B. Gene expression profiling of several pathogenesis-related genes indicated that MNR220 has an elevated and rapid pathogen-induced response. In addition to its potential use in breeding for resistance to multiple diseases, high-resolution mapping and cloning of the disease resistance locus in MNR220 may lead to a better understanding of the regulation of defense responses in wheat.     

A certain but non-exclusive association between Polymyxa graminis special forms and cereals

Polymyxo graminis, a ubiquitous plasmodiophorid obligate root endoparasite, is recognized as the vector of about 15 viruses on cereals and groundnut in temperate and tropical areas. Within the species, five special forms have been distinguished on the basis of specific ribotypes. Three of them occur in tropical areas: P. graminis f.sp. colombiana on rice, P. graminis f.sp. subtropicalis on cereals cropped in the tropics such as maize, pearl millet and sorghum but also on barley and/or wheat, and P. graminis f.sp. tropicalis mainly on maize, pearl millet and sorghum. Their particular host ranges distinguish them significantly from P. graminis f.sp. temperata and P. graminis f.sp. tepida found in temperate areas on barley and wheat. In order to assess whether these special forms commonly infect these cereals, barley and wheat plants were grown under controlled conditions on two soils from Belgium and France and both infested by P. graminis f.sp. temperata and P. graminis f.sp. tepida. The infection of each cereal species by each form was quantified by real-time quantitative PCR with specific primers and Taqman probes. The infection of P. graminis f.sp. temperata was significantly higher on barley than on wheat, whereas the quantities of P. graminis f.sp. tepida on wheat were higher than on barley. These results show that the distinction between these special forms, based on the ribotype, reflects differences in ecological features.     

Effect of temperature, light and host on prepenetration development of Puccinia graminis avenae and Puccinia coronata avenae

The influence of temperature and light on prepenetration development of single and mixed isolates of Puccinia graminis avenae and Puccinia coronata avenae was studied on 0–2% water agar and on leaves of three oat cultivars and on three non-cultivated species of Avena. Germination of uredospores of P. graminis avenae and P. coronata avenae occurred best at 10–30^oC and at 20^oC respectively. The optimum temperature for germ-tube growth and for appressorial formation was 20^oC for both rusts. An inverse relationship was observed between light intensity and prepenetration development with maximal germination of uredospores, germ-tube growth and appressorial formation occurring in darkness. Under optimum conditions maximum percentage germination and appressorium formation of both rusts was attained within 4 and 12 h after inoculation respectively. The proportion of germinated uredospores of crown rust which gave rise to appressoria was about twice that observed for stem rust. No significant differences were observed in prepenetration development between the single and mixed race inocula of the two rusts. Although germination of uredospores was significantly greater on water agar than on oat leaves, there were no significant differences in prepenetration development of the rusts on the various oat cultivars and species examined. Consequently, the data failed to indicate the presence of resistance mechanisms operating during the prepenetration phase of the infection process on the cultivars and species examined.     

Host and environmental effects on the penetration of oats by Puccinia graminis avenae and Puccinia coronata avenae

The frequency of penetration from appressoria of Puccinia graminis avenae and P. coronata avenae varied among Avena species and between oat cultivars, although both rusts produced susceptible infection type pustules on the cultivars tested. Penetration on cv. Garry was significantly less than that on the Avena species (A. barbata, A.fatua and A. sterilis) studied and penetration of these Avena species was significantly less than on the cvs Algerian and Fulmark. When the rusts were allowed to develop into pustules on seedlings which had been inoculated with fixed amounts of inoculum, there was a direct relationship between number of pustules produced and penetration frequency. The effects of temperature, light and dew period on penetration from appressoria of ‘single race’ and ‘mixed race’ inocula was also studied on these cultivars and species. Penetration by P. graminis avenae was greatest at 30–35 °C and at light intensities of 5625 lux and above, whereas that by P. coronata avenae was greatest at 20 °C and was unaffected by artificial light intensities up to n 250 lux. Maximal penetration by P. graminis avenae and P. coronata avenae was observed after inoculated plants had been exposed to dew periods of 16 and 12 h respectively. Some penetration was observed after a dew period of 8 h. The time taken for each rust to attain maximum penetration varied from 36 to 52 h after inoculation, depending on the environment, and was usually less for P. coronata avenae than for P. graminis avenae.     

Nonhost resistance of rice to rust pathogens

Rice is atypical in that it is an agricultural cereal that is immune to fungal rust diseases. This report demonstrates that several cereal rust species (Puccinia graminis f. sp tritici, P. triticina, P. striiformis, and P. hordei) can infect rice and produce all the infection structures necessary for plant colonization, including specialized feeding cells (haustoria). Some rust infection sites are remarkably large and many plant cells are colonized, suggesting that nutrient uptake occurs to support this growth. Rice responds with an active, nonhost resistance (NHR) response that prevents fungal sporulation and that involves callose deposition, production of reactive oxygen species, and, occasionally, cell death. Genetic variation for the efficacy of NHR to wheat stem rust and wheat leaf rust was observed. Unlike cereal rusts, the rust pathogen (Melampsora lini) of the dicotyledenous plant flax (Linum usitatissimum) rarely successfully infects rice due to an apparent inability to recognize host-derived signals. Morphologically abnormal infection structures are produced and appressorial-like structures often don't coincide with stomata. These data suggest that basic compatibility is an important determinate of nonhost infection outcomes of rust diseases on cereals, with cereal rusts being more capable of infecting a cereal nonhost species compared with rust species that are adapted for dicot hosts.     

Host and environmental effects on post-penetration development of Puccinia graminis avenae and P. coronata avenae

After inoculation of Avena sterilis and the oat cultivars Algerian and Garry with Puccinia graminis avenae the time required for the eruption of pustules of the rust was markedly less at 30–35 than at 20–25 ^oC. In addition, the area of pustules and number of uredospores produced were significantly greater at 30–35 than at 20 ^oC. Peaks of uredospore production occurred between 12 and 22 days after inoculation. In comparable experiments, the time required for pustules of P. coronata avenae to erupt, and the size of pustules, were relatively insensitive to change of temperature, although weight of uredospores produced was greater at 20 than at 30 ^oC. Peaks of uredospore production occurred between 14 and 18 days after inoculation. Both rusts showed straight-line relationships between pustule area and number of uredospores produced. The percentage of infection foci that developed into pustules was similar with both rusts and on all the oat cultivars examined. Both rusts produced susceptible reaction types on all the hosts tested. Pustules of P. graminis avenae were smaller and fewer and generation time longer on cv. Garry than on cv. Algerian or Avena sterilis and the numbers of pustules per unit of inoculum of both rusts were greatest on Algerian, least on Garry. It is suggested that these quantitative differences in phases of the infection process contribute towards the ‘slow-rusting’ reaction of cv. Garry.     

Production of teliospores by some races of Puccinia graminis f. sp. tritici in detached wheat and barley leaves

Five races of Puccinia graminis f. sp. tritici cultured on wheat and barley leaves in two nutrient solutions were studied for teliospore formation by subjecting them to varying treatments of temperature and light. The early appearance as well as a high percentage of teliospore formation occurred in 100 ppm benzimidazole solution on wheat or barley leaves kept at 30°C and 500 footcandles of light. The feasibility of maintaining and multiplying races of Puccinia graminis f. sp. tritici in continuous cultures on detached leaves depends on various factors, the most important being the onset of the teliostage of the fungus. The appearance of the teliospore denotes the culmination of the sporophytic or repeating stage of the wheat rusts. In this paper, some factors that influence the production of teliospores by certain races of Puccinia graminis tritici in detached leaves are discussed.     

Gene-for-gene interactions between the rye mildew fungus and wheat cultivars.

Genetic mechanisms of the incompatibility between Erysiphe graminis f.sp. secalis and wheat cultivars were analyzed using F1 hybrids between E. graminis f.sp. secalis, Sk-1, and f.sp. tritici, Tk-1. The avirulence of Sk-1 on Triticum aestivum 'Norin 4', 'Chinese Spring', and 'Kokeshi-komugi' was controlled by a single gene. The resistance of the three cultivars to Sk-1 was also controlled by a single gene, Pm15, a gene for resistance to E. graminis f.sp. agropyri. Implications of these results were discussed in terms of host-parasite coevolution.     

The barley apoptosis suppressor homologue BAX inhibitor-1 compromises nonhost penetration resistance of barley to the inappropriate pathogen Blumeria graminis f. sp. tritici

BAX inhibitor-1 (BI-1) proteins have been characterized as suppressors of programmed cell death in mammals and plants. The barley BI-1 is a suppressor of nonspecific background resistance and mlo-mediated penetration resistance to the biotrophic fungal pathogen Blumeria graminis f. sp. hordei when overexpressed in epidermal cells of barley. We report here that BI-1 expression is also slightly up-regulated during interaction with the inappropriate wheat pathogen Blumeria graminis f. sp. tritici. Significantly, overexpression of BI-1 in single epidermal cells of barley by microprojectile-mediated transformation rendered cells susceptible to penetration by inappropriate B. graminis f. sp. tritici. The degree of transgene-induced accessibility to B. graminis f. sp. tritici was thereby similar to the effect achieved by overexpression of the defense suppressor gene Mlo and could not be further enhanced by double expression of both BI-1 and Mlo. Confocal laser scanning microscopy was used to locate a functional green fluorescing GFP:BI-1 fusion protein in endomembranes and the nuclear envelope of barley epidermal cells. Together, enhanced expression of barley BI-1 suppresses penetration resistance to B. graminis f. sp. tritici, linking barley nonhost resistance with cell death regulation.     

Preliminary Investigations on the Rusts of Perennial Ryegrass in Central Italy

The results of a four-year investigation carried out in Central Italy on the occurrence of perennial ryegrass rusts from 1989—92 are reported. The crown rust (Puccinia coronata) was the most frequently recorded in the surveyed are while P. graminis was only occasionally detected. The causal agents were identified by the morphological criteria proposed by CUMMINS (1971).
A single-pustule isolate of P. coronata was used to test the resistance of seven ryegrass cultivars under greenhouse conditions. Two cultivars, 'Artal' and 'Score', proved resistant to the fungus isolate while the remaining ('Elka', 'Patora', 'Pleno', 'Sisu' and 'Vejo') were high susceptible.     

Protein polyubiquitination plays a role in basal host resistance of barley

To study protein ubiquitination pathways in the interaction of barley (Hordeum vulgare) with the powdery mildew fungus (Blumeria graminis), we measured protein turnover and performed transient-induced gene silencing (TIGS) of ubiquitin and 26S proteasome subunit encoding genes in epidermal cells. Attack by B. graminis hyperdestabilized a novel unstable green fluorescent protein fusion that contains a destabilization domain of a putative barley 1-aminocyclopropane-1-carboxylate synthase, suggesting enhanced protein turnover. Partial depletion of cellular ubiquitin levels by TIGS induced extreme susceptibility of transformed cells toward the appropriate host pathogen B. graminis f. sp hordei, whereas papilla-based resistance to the nonhost pathogen B. graminis f. sp tritici and host resistance mediated by the mlo gene (for mildew resistance locus O) remained unaffected. Cells were rescued from TIGS-induced ubiquitin depletion by synthetic genes encoding wild-type or mutant barley monoubiquitin proteins. The strongest rescue was from a gene encoding a K63R mutant form of ubiquitin blocked in several ubiquitination pathways while still allowing Lys-48-dependent polyubiquitination required for proteasomal protein degradation. Systematic RNA interference of 40 genes encoding all 17 subunits of the proteasome 19S regulatory particle failed to induce hypersusceptibility against B. graminis f. sp hordei. This suggests a role for Lys-48-linked protein polyubiquitination, which is independent from the proteasome pathway, in basal host defense of barley.     

Isogamous,hermaphroditic inheritance of mitochondrion-encoded resistance to Qo inhibitor fungicides in Blumeria graminis f. sp. tritici

A mutation of glycine to alanine at position 143 in the mitochondrial cytochrome b amino acid sequence of Blumeria graminis f. sp. tritici cosegregated with the QoI-resistant phenotype in a ratio of 1:1 in a cross between a sensitive and a resistant isolate. This mutation was used as a mitochondrial marker to determine whether mitochondrial inheritance in B. graminis was anisogamous, as in heterothallic Neurospora sp., or isogamous and hermaphroditic, as in Aspergillus nidulans. Segregation of mitochondrial genotypes in B. graminis f. sp. tritici was consistent with inheritance of mitochondria being hermaphroditic and isogamous, in that all ascospores from an individual cleistothecium had the same mitochondrial genotype and that either parent could act as the maternal parent of a cleistothecium. Within each cleistothecium, nuclear segregation occurred independently of mitochondrial inheritance, as shown by segregation of resistance to the fungicide triadimenol and by segregation of avirulences to the wheat cultivars Galahad (Pm2), Armada (Pm4b), and Holger (Pm6).     

Role of topography sensing for infection-structure differentiation in cereal rust fungi

Over 90% of the germ tubes of Puccinia graminis tritici (wheat stem rust) and Puccinia hordei (barley brown rust) differentiate appressoria on encountering stomata.There has been controversy as to the role of host topographical signals in the highly precise and efficient induction of these infection structures over stomata by cereal rusts. In the present study, polystyrene replicas of microfabricated silicon wafers, bearing precise microtopographies of defined dimensions, were used to investigate the influence of ridge spacing and height on infection-structure induction by P. graminis tritici and P. hordei. It was found that artificial topographical signals alone can induce a reproducibly high percentage (83–86%) of germ tubes to differentiate infection structures. Multiple, closely spaced (1.5 μm) ridges which were 2.0 μm high provided the most inductive topography. Differentiation on flat surfaces and over single ridges was < 4%. Appressorium induction commonly initiated a cascade of differentiation events involving the formation of infection pegs, vesicles, infection hyphae, and occasionally haustorial mother cells. It is suggested that the close spacing of cell junctions associated with the dumbbell-shaped guard cells of cereal stomatal complexes provide inductive signals for infection-structure formation by cereal rusts in vivo. Received: 17 October 1996 / Accepted: 5 December 1996     

Control of Gaeumannomyces graminis Var. tritici in Wheat by Isolates of the G. graminis var. graminis/Phialophora sp. (Lobed Hyphopodia) Complex under Field Conditions

In an experiment carried out under field conditions, wheat inoculated with Gaeumanmyces graminis var. tritics had less take-all and yielded significantly more grain when simultaneously inoculated with G. graminis var. graminis and a lobed hyphopodiate phialophora sp. then when unprotected with these fungi.     

Colonization of barley roots by endophytic fungi and their reduction of take-all caused by Gaeumannomyces graminis var. tritici

Fungal root endophytes obtained from natural vegetation were tested for antifungal activity in dual culture tests against the root pathogen Gaeumannomyces graminis var. tritici. Fifteen isolates, including Acremonium blochii, Acremonium furcatum, Aspergillus fumigatus, Cylindrocarpon sp., Cylindrocarpon destructans, Dactylaria sp., Fusarium equiseti, Phoma herbarum, Phoma leveillei, and a sterile mycelium, selected based on the dual culture test, were inoculated on barley roots in growth tubes under axenic conditions, both in the absence and presence of G. graminis var. tritici. All isolates colonized the rhizosphere and very often the root cortex without causing disease symptoms and without affecting plant growth. Eight isolates significantly reduced the symptoms caused by G. graminis var. tritici, and 6 of them reduced its presence in the roots.     

普通小麦99-2439 中的白粉病抗性遗传

普通冬小麦品系99-2439在郑州连续4年对田间白粉菌(Blumeria graminis sp. tritici)表现高抗,但其抗性基因来源不清。通过染色体C-分带和1RS染色体特异性SCAR标记鉴定, 表明它是一个小麦-黑麦(Triticum aestivum - Secale cereale)1BL/1RS异易位系。通过对中国春×99-2439杂交F2代分离群 体抗性鉴定和1RS染色体臂检测结果分析, 证明该抗病基因不在1RS染色体臂上。用单孢小麦白粉菌分离株对其抗性遗传进行研究, 结果表明, 99-2439的白粉病抗性由一对小种专化、隐性抗病基因控制。由于携带Pm5a的Hope/8Cc对中国的21个小麦白粉菌分离菌株均高度感病, 而99-2439高抗混和白粉菌和5个单孢分离菌株, 所以, 99-2439所携带的抗白粉病基因不同于Pm5a。     

普通小麦99-2439中的白粉病抗性遗传

普通冬小麦品系99-2439在郑州连续4年对田间白粉菌(Blumeria graminis sp.tritici)表现高抗,但其抗性基因来源不清.通过染色体C-分带和IRS染色体特异性SCAR标记鉴定,表明它是一个小麦-黑麦(Triticum aestivum-Secale cereale)lBL/1RS异易位系.通过对中国春×99-2439杂交F2代分离群体抗性鉴定和1RS染色体臂检测结果分析,证明该抗病基因不在1RS染色体臂上.用单孢小麦白粉菌分离株对其抗性遗传进行研究,结果表明,99-2439的白粉病抗性由一对小种专化、隐性抗病基因控制.由于携带Pm5a的Hope/8Cc对中国的21个小麦白粉菌分离菌株均高度感病,而99-2439高抗混和白粉菌和5个单孢分离菌株,所以,99-2439所携带的抗白粉病基因不同于Pm5a.     

小麦硫代硫酸硫转移酶类似基因的克隆与定位

小麦-簇毛麦6VS/6AL易位系92R137含有抗白粉病基因Pm21。为了研究该易位系的抗病机理,应用mRNA差异显示和快速扩增cDNA未端（Rapid Amplification of cDNAEnd,RACE)技术对在白粉菌诱导后表达增强的基因进行了克隆,分离到1个命名为TaTST的全长cDNA序列。Northern杂交分析表明,TaTST基因在白粉菌诱导后表达明显增强,24h达到峰值,氨基酸序列同源性分析表明,TaTST与Datisca glomerata的硫代硫酸硫转移酶基因（rho-danese,EC,2.8.1.1)序列有64%相同,80%相似,用中国春缺体/四体系和端体系Southern杂交和基因特异性引物扩增（gene specific primer-PCR)将TaTST基因定位在小麦6B染色体短臂上,Southern杂交表明,该基因为单拷贝基因,由于在杨麦5号和6VS/6AL易位系间存在明显多态,可以推测在6VS上有TaTST的同源基因,TaTST是从小麦中分离的新基因。白粉菌诱导后的表达变化提示;TaTST与小麦抗白粉病反应有关。