The Arabidopsis genes RPW8.1 and RPW8.2 confer induced resistance to powdery mildew diseases in tobacco

Plant disease resistance (R) gene products recognize pathogen avirulence (Avr) gene products and induce defense responses. It is not known if an R gene can function in different plant families, however. The Arabidopsis thaliana R genes RPW8.1 and RPW8.2 confer resistance to the powdery mildew pathogens Erysiphe orontii, E. cichoracearum, and Oidium lycopersici, which also infect plants from other families. We produced transgenic Nicotiana tabacum, N. benthamiana, and Lycopersicon esculentum plants containing RPW8.1 and RPW8.2. Transgenic N. tabacum plants had increased resistance to E. orontii and O. lycopersici, transgenic N. benthamiana plants had increased resistance to E. cichoracearum, but transgenic L. esculentum plants remained susceptible to these pathogens. The defense responses induced in transgenic N. tabacum and N. benthamiana were similar to those mediated by RPW8.1 and RPW8.2 in Arabidopsis. Apparently, RPW8.1 and RPW8.2 could be used to control powdery mildew diseases of plants from other families.     

Natural genetic resources of Arabidopsis thaliana reveal a high prevalence and unexpected phenotypic plasticity of RPW8-mediated powdery mildew resistance

Here, an approach based on natural genetic variation was adopted to analyse powdery mildew resistance in Arabidopsis thaliana. Accessions resistant to multiple powdery mildew species were crossed with the susceptible Col-0 ecotype and inheritance of resistance was analysed. Histochemical staining was used to visualize archetypal plant defence responses such as callose deposition, hydrogen peroxide accumulation and host cell death in a subset of these ecotypes. In six accessions, resistance was likely of polygenic origin while 10 accessions exhibited evidence for a single recessively or semi-dominantly inherited resistance locus. Resistance in the latter accessions was mainly manifested at the terminal stage of the fungal life cycle by a failure of abundant conidiophore production. The resistance locus of several of these ecotypes was mapped to a genomic region containing the previously analysed atypical RPW8 powdery mildew resistance genes. Gene silencing revealed that members of the RPW8 locus were responsible for resistance to Golovinomyces orontii in seven accessions. These results suggest that broad-spectrum powdery mildew resistance in A. thaliana is predominantly of polygenic origin or based on RPW8 function. The findings shed new light on the natural variation of inheritance, phenotypic expression and pathogen range of RPW8-conditioned powdery mildew resistance.     

亚精胺诱导黄瓜幼苗对白粉病抗性的研究

以黄瓜品种‘长春密刺’幼苗为材料,研究了亚精氨(Spd)诱导黄瓜幼苗对白粉病的抗性,并测定Spd处理和白粉菌接种对黄瓜叶片4种防御酶活性及3种防卫基因表达的影响。结果显示:(1)0.2～1.0mmol.L-1 Spd对黄瓜幼苗白粉病抗性均有不同程度的诱抗效果,并以0.8mmol.L-1 Spd处理效果最明显,诱导效率可达55.3%。(2)喷施Spd或接种白粉菌均可提高黄瓜叶片过氧化物酶(POD)、苯丙氨酸解氨酶(PAL)、几丁质酶和β-1,3-葡聚糖酶的活性,且诱导并接种处理的植株叶片上述酶活性均比只诱导不接种处理的上升速度更快;同时,Spd处理和接种白粉菌可以提高植株叶片中POX、PAL、PR-1a基因的表达量。研究表明,Spd处理可以诱导防卫基因表达的增强,提高防御酶活性,显著降低病情指数,增强黄瓜幼苗对白粉病的抗性。     

Powdery mildew (Ascomycotina: Erysiphales) — an alternative food for the predatory mite Typhlodromus pyri Scheuten (Acari: Phytoseiidae)

The ability of Typhlodromus pyri Scheuten to feed on powdery mildew was investigated. Adult females of T. pyri which had previously been fed on the pollen of Pinus sylvestris L. were offered three powdery mildew species and their feeding behaviour was observed. The females fed readily on the conidia of Erysiphe orontii Cast. from tobacco and Oidium fragariae Harz. from strawberry while no feeding was observed on Erysiphe polygoni DC. from red clover, even after the mites were starved for 2 days. Erysiphe orontii was tested for its effect on the survival, oviposition and development of T. pyri. The results showed that the conidia of E. orontii can supply mites with the water and nutrients necessary for their normal development. Mature females successfully mated and reproduced, although their oviposition rate was low. This revised version was published online in November 2006 with corrections to the Cover Date.     

Detection and identification of bacteria intimately associated with fungi of the order Sebacinales

Because of their beneficial impact on plants, the highly diverse mycorrhizal fungi grouped in the order Sebacinales lay claim to high ecological and agricultural significance. Here, we describe for the first time associations of Sebacinoid members with bacteria. Using quantitative PCR, denaturating gradient gel electrophoresis and fluorescence in situ hybridization, we detected an intimate association between Piriformospora indica and Rhizobium radiobacter, an alpha-Proteobacterium. The stability of the association, vertical transmission of the bacteria during asexual fungal reproduction and fungal plant colonization was monitored using R. radiobacter-specific primers. Treatment of mycelium or fungal protoplasts with antibiotics highly efficient against the free bacteria failed to cure the fungus. Barley seedlings dip-inoculated with R. radiobacter showed growth promotion and systemic resistance to the powdery mildew fungus Blumeria graminis comparable to P. indica inoculation. By screening additional isolates of the Sebacina vermifera complex, three species-specific associations with bacteria from the genera Paenibacillus, Acinetobacter and Rhodococcus were found. These findings suggest that Sebacinales species regularly undergo complex interactions involving host plants and bacteria reminiscent of other ectomycorrhizal and endomycorrhizal associations.     

A SNARE-protein has opposing functions in penetration resistance and defence signalling pathways

Penetration resistance is often the first line of defence against fungal pathogens. Subsequently induced defences are mediated by the programmed cell death (PCD) reaction pathway and the salicylic acid (SA), jasmonic acid (JA) and ethylene (ET) signalling pathways. We previously demonstrated that full penetration resistance in Arabidopsis against the non-host barley powdery mildew fungus (Blumeria graminis f.sp. hordei) requires the syntaxin SYP121 (PEN1). Here we report that SYP121, together with SYP122, functions as a negative regulator of subsequently induced defence pathways. The SA level in the syntaxin double mutant syp121-1 syp122-1 is dramatically elevated, resulting in necrosis and dwarfism. This phenotype is partially rescued by introducing the SA-signalling mutations eds1-2, eds5-3, sid2-1 and npr1-1 as well as the NahG transgene. These partially rescued triple mutants have an unknown defence to Pseudomonas syringae pv. tomato, and have increased HR-like responses to non-host and host powdery mildew fungi. The HR-like responses cause efficient resistance to the latter. These defence pathways are SA-independent. Furthermore, the JA/ET signalling marker, PDF1.2, is highly upregulated in the triple mutants. Thus SYP121 and SYP122 are negative regulators of PCD, SA, JA and ET pathways through a molecular function distinct from that of SYP121 in penetration resistance. Our data suggest that individual cells preferentially express either penetration resistance or the subsequently induced defences.     

Correlation of defense gene induction defects with powdery mildew susceptibility in Arabidopsis enhanced disease susceptibility mutants

We investigated the relative importance of specific Arabidopsis thaliana genes in conferring resistance to bacterial versus fungal pathogens. We first developed a pathosystem involving the infection of Arabidopsis accession Columbia with a virulent isolate of the obligate biotrophic fungal pathogen Erysiphe orontii. E. orontii elicited the accumulation of mRNAs corresponding to the defense-related genes PR1, BGL2 (PR2), PR5 and GST1 , but did not elicit production of the phytoalexin camalexin or the accumulation of defensin ( PDF1.2 ) or thionin ( THI2.1 ) mRNAs. We tested a set of 15 previously isolated Arabidopsis phytoalexin deficient (pad), non-expresser of PR (npr) and enhanced disease susceptibility (eds) mutants that are more susceptible to Pseudomonas syringae for their susceptibility to E. orontii. Four of these mutants ( pad4–1, npr1–1, eds5–1 and a double npr1–1 eds5–1 mutant) as well as Arabidopsis lines carrying a nahG transgene exhibited enhanced susceptibility to E. orontii and reduced levels of PR gene expression . Comparison of the PR gene induction patterns in response to E. orontii in the various mutants and in the nahG transgenics suggests the existence of NPR1 -independent salicylate-dependent and NPR1 -independent salicylate-independent defense gene activation pathways. Eleven other eds and pad mutants did not show measurable enhanced susceptibility to E. orontii , suggesting that these mutants are defective in factors that are not important for the limitation of E. orontii growth.     

Cell-autonomous expression of barley Mla1 confers race-specific resistance to the powdery mildew fungus via a Rar1-independent signaling pathway

The barley Mla locus encodes 28 characterized resistance specificities to the biotrophic fungal pathogen barley powdery mildew. We describe a single-cell transient expression assay using entire cosmid DNAs to pinpoint Mla1 within the complex 240-kb Mla locus. The MLA1 cDNA encodes a 108-kD protein containing an N-terminal coiled-coil structure, a central nucleotide binding domain, and a C-terminal leucine-rich repeat region; it also contains a second short open reading frame at the 5' end that has a possible regulatory function. Although most Mla-encoded resistance specificities require Rar1 for their function, we used the single-cell expression system to demonstrate that Mla1 triggers full resistance in the presence of the severely defective rar1-2 mutant allele. Wheat contains an ortholog of barley Mla, designated TaMla, that is tightly linked to (0.7 centimorgan) but distinct from a tested resistance specificity at the complex Pm3 locus to wheat powdery mildew. Thus, the most polymorphic powdery mildew resistance loci in barley and wheat may have evolved in parallel at two closely linked homeoloci. Barley Mla1 expressed in wheat using the single-cell transformation system failed to trigger a response to any of the wheat powdery mildew Avr genes tested, indicating that AvrMla1 is not genetically fixed in wheat mildew strains.     

Powdery Mildew Resistance in Barley Landraces from Morocco

Nineteen barley landraces collected from Morocco were screened for resistance to powdery mildew. The landraces originated from the collection at the Polish Gene Bank, IHAR Radzików, Poland. The fifteen landraces tested showed powdery mildew resistance reactions and 35 single plant lines were selected. Twenty-one of these lines were tested in the seedling stage with 30, four lines with 17 and another 10 lines with 23 differential isolates of powdery mildew, respectively. The isolates were chosen according to their virulence spectra observed on the Pallas isolines differential set. Nine lines (E 1029-1-1, E 1042-2-2, E 1050-1-1, E 1054-5-1, E 1056-2-5, E 1056-3-1, E 1061-1-1, E 1061-1-3 and E 1067-1-2) which originated from seven landraces showed resistance to all prevalent European powdery mildew virulence genes. The most frequent score was 2 and 16 lines showed this reaction for inoculation with most isolates used. The distribution of reaction type indicated that about 77% of all reaction types observed were classified as powdery mildew resistance (scores 0, 1 and 2). In all lines the presence of unknown genes alone or in combinations with specific ones was postulated. Four different resistance alleles ( Mlat , Mla6 , Mla14 and Mla12 ) were postulated to be present in 10 tested lines alone or in combination. Alleles Mlat , Mla6 and Mla14 were postulated to be present in four and Mla12 in two tested lines, respectively. The value of barley landraces for diversification of resistance genes for powdery mildew is discussed.     

QTLs for powdery mildew resistance in peach ×Prunus davidiana crosses: consistency across generations and environments

Powdery mildew, caused by Sphaerotheca pannosa var. persicae is one of the most important diseases in European peach orchards. Quantitative trait loci controlling powdery mildew resistance were detected using three related F1, F2 and BC2 populations derived from the cross between the resistant parent P. davidiana clone P1908 and the susceptible peach cultivar Summergrand. Powdery mildew resistance of each population was evaluated under natural exposure, in several locations and over several years. Thirteen QTLs were detected. For nine of them, the favourable allele came from the resistant parent. Five QTLs were consistently detected across the three populations. The F1 hybrid used to produce F2 and BC2 populations had not inherited the favourable allele from P1908 for QTL detected on LG3 and LG8 in F1 population. QTLs were not detected in the corresponding regions in F2 and BC2 populations. In two other genomic areas, significant substitution effects between P1908 alleles were evidenced in the F1 population, but the favourable allele came from Summergrand in the F2 and BC2 populations. Analysis of phenotypic data suggested an important qualitative change in the distribution of powdery mildew resistance after 1996, confirmed by QTL analysis. Indeed, a dramatic decrease of the effect of the major QTL previously detected on LG6 was observed after 1996, while the QTL on LG8 was increasingly involved in the control of powdery mildew resistance. Consequences for peach breeding strategies to improve powdery mildew resistance are discussed.     

TaRPP13-3, a CC-NBS-LRR-like gene located on chr 7D,promotes disease resistance to wheat powdery mildew in Brock

Disease resistance (R) gene, RPP13, plays an important role in the resistance of plants to pathogen infections; its function in resistance of wheat to powdery mildew remains unknown. In this study, a RNA-Seq technique was used to monitor expression of genes in susceptible wheat ‘Jing411’ and resistant near-isogenic line ‘BJ-1’ in response to powdery mildew infection. Overall, 413 differential expression genes were observed and identified as involved in disease resistance. RPP13 homologous gene on wheat chromosome 7D was preliminarily identified using the wheat 660K SNP chip. RPP13 was highly expressed in ‘BJ-1’ and encodes 1,027 amino acids, including CC, NB and LRR domain, termed TaRPP13-3. After inoculation with powdery mildew, expression of TaRPP13-3 in resistant wheat changed with time, but average expression was higher when compared to susceptible variety, thus indicating that TaRPP13-3 is involved in resistance to powdery mildew. Virus-induced gene silencing (VIGS) was used to inhibit expression of TaRPP13-3 in resistant parent ‘Brock’. Results indicated that silencing of TaRPP13-3 led to decreased disease resistance in ‘Brock’. Overall results of this study indicate that TaRPP13-3 gene is involved in the defence response of wheat to powdery mildew and plays a positive role in wheat powdery mildew interactions.     

Linkage of a RAPD marker with powdery mildew resistance

The aim of this study was to investigate the inheritance of powdery mildew disease and to tag it with a DNA marker to utilize for the marker-assisted selection (MAS) breeding program. The powdery mildew resistant genotype Fallon(er) and susceptible genotype 11760-3ER were selected from 177 genotypes by heavy infestation of germplasm with Erysiphe pisi through artificial inoculation. The F1 plants of the cross Fallon/11760-3 indicated the dominance of the susceptible allele, while F2 plants segregated in 3 : 1 ratio (susceptible : resistant) that fit for goodness of fitness by chi2 (P > 0.07), indicating monogenic recessive inheritance for powdery mildew resistance in Pisum sativum. A novel RAPD marker OPB18 (5'-CCACAGCAGT-3') was linked to the er-1 gene with 83% probability with a LOD score of 4.13, and was located at a distance of 11.2 cM from the er-1 gene.     

Simultaneous modification of three homoeologs of TaEDR1 by genome editing enhances powdery mildew resistance in wheat

Wheat (Triticum aestivum L.) incurs significant yield losses from powdery mildew, a major fungal disease caused by Blumeria graminis f. sp. tritici (Bgt). enhanced disease resistance1 (EDR1) plays a negative role in the defense response against powdery mildew in Arabidopsis thaliana; however, the edr1 mutant does not show constitutively activated defense responses. This makes EDR1 an ideal target for approaches using new genome‐editing tools to improve resistance to powdery mildew. We cloned TaEDR1 from hexaploid wheat and found high similarity among the three homoeologs of EDR1. Knock‐down of TaEDR1 by virus‐induced gene silencing or RNA interference enhanced resistance to powdery mildew, indicating that TaEDR1 negatively regulates powdery mildew resistance in wheat. We used CRISPR/Cas9 technology to generate Taedr1 wheat plants by simultaneous modification of the three homoeologs of wheat EDR1. No off‐target mutations were detected in the Taedr1 mutant plants. The Taedr1 plants were resistant to powdery mildew and did not show mildew‐induced cell death. Our study represents the successful generation of a potentially valuable trait using genome‐editing technology in wheat and provides germplasm for disease resistance breeding.     

Effects of fungal infection and wounding on the expression of chitinases and β-1,3 glucanases in near-isogenic lines of barley

Chitinases (EC 3.2.1.14) and β -1.3 glucanases (EC 3.2.1.39) have been known to play a vital role in the defense of plants against fungal pathogens. The pattern of induction of these two enzymes subsequent to infection by powdery mildew was studied in 10 pairs of near-isogenic lines of barley ( Hordeum vulgare L.) which possess powdery mildew resistance genes. These isogenic lines have been grotiped according to their reaction to the fungus. The induction patterns varied between the resistant and the susceptible cultivars within each group and between different groups. More tsozymcs were induced in susceptible varieties of highly resistant groups and the overall levels and the number of isozymes of chitinases and β -1.3 glucanases were lower in groups with low resistance. The effect of powdery mildew infection and mechanical wounding on the cellular localization of chitinases and β -1.3 glucanases in barley leaves has also been studied. The 31 kDa leaf chitinase, L-CH2, and trace amounts of a 25 kDa chitinase. L-CH3. were present in healthy leaves. Wounding increased the levels of L-CH3 within I ft h. Powdery mildew infection increased the levels of L-CH3 both in intercellular fluid and in intracellular extract of leaves. A /3-I.3 glucanase. GH, also increased after infection and wounding. In infected barley leaves, GL-1 was present both in intercellular space and intracellular extract. It is concluded that powdery mildew resistance genes exhibit qualitative and quantitative differences in the expression of chitinases and β -1.3 glucanases. Further, chitinases and β -1.3 glucanases appear to be a response to active infection rather than the factors responsible for disease resistance.     

Mapping of novel powdery mildew resistance gene(s) from <Emphasis Type="Italic">Agropyron cristatum</Emphasis> chromosome 2P

Key message

A physical map of Agropyron cristatum 2P chromosome was constructed for the first time and the novel powdery mildew resistance gene(s) from chromosome 2P was(were) also mapped.

Abstract

Agropyron cristatum (L.) Gaertn. (2n = 28, PPPP), a wild relative of common wheat, is highly resistant to powdery mildew. Previous studies showed that wheat-A. cristatum 2P disomic addition line II-9-3 displayed high resistance to powdery mildew, and the resistance was attributable to A. cristatum chromosome 2P. To utilize and physically map the powdery mildew resistance gene(s), 15 wheat-A. cristatum 2P translocation lines and three A. cristatum 2P deletion lines with different chromosomal segment sizes, obtained from II-9-3 using ⁶⁰Co-γ ray irradiation, were characterized using cytogenetic and molecular marker analysis. A. cristatum 2P chromosomal segments in the translocations were translocated to different wheat chromosomes, including 1A, 4A, 5A, 6A, 7A, 1B, 2B, 3B, 7B, 3D, 4D, and 6D. A physical map of the 2P chromosome was constructed with 82 STS markers, consisting of nine bins with 34 markers on 2PS and eight bins with 48 markers on 2PL. The BC₁F₂ populations of seven wheat-A. cristatum 2P translocation lines (2PT-3, 2PT-4, 2PT-5, 2PT-6, 2PT-8, 2PT-9, and 2PT-10) were developed by self-pollination, tested with powdery mildew and genotyped with 2P-specific STS markers. From these results, the gene(s) conferring powdery mildew resistance was(were) located on 2PL bin FL 0.66–0.86 and 19 2P-specific markers were identified in this bin. Moreover, two new powdery mildew-resistant translocation lines (2PT-4 and 2PT-5) with small 2PL chromosome segments were obtained. The newly developed wheat lines with powdery mildew resistance and the closely linked molecular markers will be valuable for wheat disease breeding in the future.

     

抗白粉病小偃麦异代换系的细胞学和RAPD鉴定

利用细胞学和RAPD方法,对从长穗偃麦草与普通小麦复合杂交后代中选育的抗白粉病小麦种质系山农87074－526和山农87074－551进行了鉴定。结果表明,两种质系的根尖细胞染色体数目均为2n=42,花粉母细胞减数分裂中期I（PMC MI）染色体构型为2n=21Ⅱ;二者杂交F1 PMC MI染色体构型亦为2n=21Ⅱ,两种质系分别与小麦中国春的杂种F1 PMC MI染色体构型均为2n=20Ⅱ 2I,说明两种质系为相同的双体异代系。在苗期和成株期两种质系对白粉病15号菌种均表现免疫,其白粉病抗性为显性,并且来自长穗偃麦草,抗白粉病基因位于它们所含的偃麦草染色体上。从80个随机引物中,筛选出2个引物OPE13和OPH15能在两种质系中稳定地扩增出长穗偃麦草亲本的特异DNA片段。     

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.