Genetic characterization of five powdery mildew disease resistance loci in Arabidopsis thaliana

This paper reports on six Arabidopsis accessions that show resistance to a wild isolate of the powdery mildew pathogen, Erysiphe cichoracearum . Resistance at 7 days post-inoculation in these accessions was characterized by limited fungal growth and sporadic development of chlorotic or necrotic lesions at inoculation sites. Three accessions, Wa-1, Kas-1 and SI-0, were highly resistant, while the other accessions permitted some fungal growth and conidiation. Papilla formation was a frequent host response; however, cell death appeared to be neither a rapid nor a common response to infection. To determine the genetic basis of resistance, segregation analyses of progeny from crosses between each of the resistant accessions and Columbia ( gl1 ), which is susceptible to the powdery mildew pathogen, were performed. For all accessions except SI-0, resistance was conferred by a single locus. SI-0 was unique in that two unlinked loci controlled the disease reaction phenotype. In accessions Wa-1, Kas-1, Stw-0 and Su-0, powdery mildew resistance was encoded by a semi-dominant allele. However, susceptibility was dominant to resistance in accessions Te-0 and SI-0. Mapping studies revealed that powdery mildew resistances in Kas-1, Wa-1, Te-0, Su-0 and Stw-0 were controlled by five independent loci. This study suggests that the Arabidopsis powdery mildew disease will be a suitable model system in which to investigate powdery mildew diseases.     

Involvement of lipid transfer proteins in resistance against a non-host powdery mildew in Arabidopsis thaliana

Non-specific lipid transfer proteins (LTPs) are involved in the transport of lipophilic compounds to the cuticular surface in epidermal cells and in the defence against pathogens. The role of glycophosphatidylinositol (GPI)-anchored LTPs (LTPGs) in resistance against non-host mildews in Arabidopsis thaliana was investigated using reverse genetics. Loss of either LTPG1, LTPG2, LTPG5 or LTPG6 increased the susceptibility to penetration of the epidermal cell wall by Blumeria graminis f. sp. hordei (Bgh). However, no impact on pre-penetration defence against another non-host mildew, Erysiphe pisi (Ep), was observed. LTPG1 was localized to papillae at the sites of Bgh penetration. This study shows that, in addition to the previously known functions, LTPGs contribute to pre-invasive defence against certain non-host powdery mildew pathogens.     

A novel receptor‐like kinase involved in fungal pathogen defence in Arabidopsis thaliana

Plants are under constant attack from a variety of disease‐causing organisms. Lacking an adaptive immune system, plants repel pathogen attack via an array of pathogen recognition machinery. Receptor‐like kinases (RLKs) are involved in the recognition of pathogen‐associated molecular patterns (PAMPs) and activate resistance pathways against broad classes of pathogens. We have identified powdery mildew‐resistant kinase 1, an Arabidopsis gene encoding an RLK that is highly induced by chitin at early time points and localizes to the plasma membrane. Knockout mutants in pmrk1 are more susceptible to both Golovinomyces cichoracearum and Plectosphaerella cucumerina. Our data show that PMRK1 is essential in early stages of defence against fungi and provide evidence that PMRK1 may be unique to chitin‐induced signalling pathways. The results of this study indicate that PMRK1 is a critical component of plant innate immunity against fungal pathogens.     

Quantitative trait loci analysis of powdery mildew disease resistance in the Arabidopsis thaliana accession kashmir-1.

Powdery mildew diseases are economically important diseases, caused by obligate biotrophic fungi of the Erysiphales. To understand the complex inheritance of resistance to the powdery mildew disease in the model plant Arabidopsis thaliana, quantitative trait loci analysis was performed using a set of recombinant inbred lines derived from a cross between the resistant accession Kashmir-1 and the susceptible accession Columbia glabrous1. We identified and mapped three independent powdery mildew quantitative disease resistance loci, which act additively to confer disease resistance. The locus with the strongest effect on resistance was mapped to a 500-kbp interval on chromosome III.     

Methyl jasmonate alters polyamine metabolism and induces systemic protection against powdery mildew infection in barley seedlings

Treatment of the first leaves of barley (Hordeum vulgare L. cv. Golden Promise) seedlings with methyl jasmonate (MJ) led to small, but significant increases in levels of free putrescine and spermine 1 d later and to significant increases in levels of free putrescine, spermidine and spermine by 4 d following treatment. MJ-treated first leaves also exhibited significant increases in the amounts of soluble conjugates of putrescine and spermidine 1, 2 and 4 d after treatment. In second leaves of plants where the first leaves had been treated with MJ, no significant changes in levels of free polyamines were observed, but significant increases in levels of soluble conjugates of putrescine and spermidine were detected. These changes were accompanied by increased activities of soluble ornithine decarboxylase (ODC), soluble and particulate arginine decarboxylase (ADC), and S-adenosylmethionine decarboxylase (AdoMetDC), in first and second leaves following treatment of the first leaves with MJ. Activities of soluble and particulate diamine oxidase (DAO) were also higher in first and second leaves following treatment of the first leaves with MJ. Treatment of the first leaves with MJ led to a significant reduction in powdery mildew (Blumeria graminis f. sp. hordei) infection on the second leaves and also resulted in significant increases in activities of the plant defence-related enzymes, phenylalanine ammonia lyase (PAL) and peroxidase.     

Cross-talk between wound signalling pathways determines local versus systemic gene expression in Arabidopsis thaliana

Plants react to mechanical damage by activating a set of genes, the products of which are thought to serve defensive functions. In solanaceous plants, cell wall-derived oligosaccharides and the plant hormones jasmonic acid and ethylene participate in the signalling network for wound-induced expression of proteinase inhibitors and other defence-related genes, both in the locally damaged and in the systemic non-damaged leaves. Here we show that in Arabidopsis thaliana, these signalling components interact in novel ways to activate distinct responses. In damaged tissues, oligosaccharides induce the expression of a specific set of wound-responsive genes while repressing jasmonic acid-responsive genes that are activated in the systemic tissues. The oligosaccharide-mediated repression of the jasmonic acid-dependent signalling pathway is exerted through the production and perception of ethylene in the locally damaged tissue. This cross-talk between separate wound signalling pathways thus allows the set up of different responses in the damaged and the systemic tissues of plants reacting to injury.     

Identification and utilization of a sow thistle powdery mildew as a poorly adapted pathogen to dissect post-invasion non-host resistance mechanisms in Arabidopsis

To better dissect non-host resistance against haustorium-forming powdery mildew pathogens, a sow thistle powdery mildew isolate designated Golovinomyces cichoracearum UMSG1 that has largely overcome penetration resistance but is invariably stopped by post-invasion non-host resistance of Arabidopsis thaliana was identified. The post-invasion non-host resistance is mainly manifested as the formation of a callosic encasement of the haustorial complex (EHC) and hypersensitive response (HR), which appears to be controlled by both salicylic acid (SA)-dependent and SA-independent defence pathways, as supported by the susceptibility of the pad4/sid2 double mutant to the pathogen. While the broad-spectrum resistance protein RPW8.2 enhances post-penetration resistance against G. cichoracearum UCSC1, a well-adapted powdery mildew pathogen, RPW8.2, is dispensable for post-penetration resistance against G. cichoracearum UMSG1, and its specific targeting to the extrahaustorial membrane is physically blocked by the EHC, resulting in HR cell death. Taken together, the present work suggests an evolutionary scenario for the Arabidopsis-powdery mildew interaction: EHC formation is a conserved subcellular defence evolved in plants against haustorial invasion; well-adapted powdery mildew has evolved the ability to suppress EHC formation for parasitic growth and reproduction; RPW8.2 has evolved to enhance EHC formation, thereby conferring haustorium-targeted, broad-spectrum resistance at the post-invasion stage.     

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.     

Elevated CO2 induces physiological, biochemical and structural changes in leaves of Arabidopsis thaliana

Leaves of Arabidopsis thaliana grown under elevated or ambient CO2 (700 or 370 micromol mol(-1), respectively) were examined for physiological, biochemical and structural changes. Stomatal characters, carbohydrate and mineral nutrient concentrations, leaf ultrastructure and plant hormone content were investigated using atomic absorption spectrophotometry, transmission electron microscopy and enzyme-linked immunosorbent assay (ELISA). Elevated CO2 reduced the stomatal density and stomatal index of leaves, and also reduced stomatal conductance and transpiration rate. Elevated CO2 increased chloroplast number, width and profile area, and starch grain size and number, but reduced the number of grana thylakoid membranes. Under elevated CO2, the concentrations of carbohydrates and plant hormones, with the exception of abscisic acid, increased whereas mineral nutrient concentrations declined. These results suggest that the changes in chloroplast ultrastructure may primarily be a consequence of increased starch accumulation. Accelerated A. thaliana growth and development in elevated CO2 could in part be attributed to increased foliar concentrations of plant hormones. The reductions in mineral nutrient concentrations may be a result of dilution by increased concentrations of carbohydrates and also of decreases in stomatal conductance and transpiration rate.     

A mutation in a coproporphyrinogen III oxidase gene confers growth inhibition, enhanced powdery mildew resistance and powdery mildew-induced cell death in Arabidopsis

Key message

A gene encoding a coproporphyrinogen III oxidase mediates disease resistance in plants by the salicylic acid pathway.

Abstract

A number of genes that regulate powdery mildew resistance have been identified in Arabidopsis, such as ENHANCED DISEASE RESISTANCE 1 to 3 (EDR1 to 3). To further study the molecular interactions between the powdery mildew pathogen and Arabidopsis, we isolated and characterized a mutant that exhibited enhanced resistance to powdery mildew. The mutant also showed dramatic powdery mildew-induced cell death as well as growth defects and early senescence in the absence of pathogens. We identified the affected gene by map-based cloning and found that the gene encodes a coproporphyrinogen III oxidase, a key enzyme in the tetrapyrrole biosynthesis pathway, previously known as LESION INITIATION 2 (LIN2). Therefore, we designated the mutant lin2-2. Further studies revealed that the lin2-2 mutant also displayed enhanced resistance to Hyaloperonospora arabidopsidis (H.a.) Noco2. Genetic analysis showed that the lin2-2-mediated disease resistance and spontaneous cell death were dependent on PHYTOALEXIN DEFICIENT 4 (PAD4), SALICYLIC ACID INDUCTION-DEFICIENT 2 (SID2), and NONEXPRESSOR OF PATHOGENESIS-RELATED GENES 1 (NPR1), which are all involved in salicylic acid signaling. Furthermore, the relative expression levels of defense-related genes were induced after powdery mildew infection in the lin2-2 mutant. These data indicated that LIN2 plays an important role in cell death control and defense responses in plants.     

Local and systemic production of nitric oxide in tomato responses to powdery mildew infection

Various genetic and physiological aspects of resistance of Lycopersicon spp. to Oidium neolycopersici have been reported, but limited information is available on the molecular background of the plant–pathogen interaction. This article reports the changes in nitric oxide (NO) production in three Lycopersicon spp. genotypes which show different levels of resistance to tomato powdery mildew. NO production was determined in plant leaf extracts of L. esculentum cv. Amateur (susceptible), L. chmielewskii (moderately resistant) and L. hirsutum f. glabratum (highly resistant) by the oxyhaemoglobin method during 216 h post-inoculation. A specific, two-phase increase in NO production was observed in the extracts of infected leaves of moderately and highly resistant genotypes. Moreover, transmission of a systemic response throughout the plant was observed as an increase in NO production within tissues of uninoculated leaves. The results suggest that arginine-dependent enzyme activity was probably the main source of NO in tomato tissues, which was inhibited by competitive reversible and irreversible inhibitors of animal NO synthase, but not by a plant nitrate reductase inhibitor. In resistant tomato genotypes, increased NO production was localized in infected tissues by confocal laser scanning microscopy using the fluorescent probe 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate. NO production observed in the extracts from pathogen conidia, together with elevated NO production localized in developing pathogen hyphae, demonstrates a complex role of NO in plant–pathogen interactions. Our results are discussed with regard to a possible role of increased NO production in pathogens during pathogenesis, as well as local and systemic plant defence mechanisms.     

Characterizing the pathotype structure of barley powdery mildew and effectiveness of resistance genes to this pathogen in Kazakhstan

Background

Powdery mildew of barley is a wind-borne and obligate biotrophic pathogen, which ranks among the most widespread barley pathogens worldwide. However, purposeful research towards studying the structure of the barley powdery mildew populations, of their virulence and of effectiveness of certain resistance genes against the infection was not conducted in Kazakhstan till present time. This paper is the first to describe characteristics of the pathotype structure of Blumeria graminis f.sp. hordei (Bgh) population and effectiveness of resistance genes in two regions of barley cultivation in the republic.

Results

One hundred and seven isolates of Bgh were obtained from seven populations occurring on cultivated barley at two geographically locations in Kazakhstan during 2015 and 2016. Their virulence frequency was determined on 17 differential lines Pallas. All isolates were virulent on the resistance gene Mla8 and avirulent for the resistance genes Mla9, Mla1 + MlaAl2, Mla6 + Mla14, Mla13 + MlRu3, Mla7 + MlNo3, Mla10 + MlDu2, Mla13 + MlRu3 and Mlo-5. The frequencies of isolates overcoming the genes Mla3, Mla22, Mlat Mlg + MlCP and Mla12 + MlEm2 were 0.0–33.33%, and frequencies of isolates overcoming the genes Mlra, Mlk, MlLa and Mlh ranged from 10.0 to 78.6%. Based on reactions of differential lines possessing the genes Mla22, Mlra, Mlk, Mlat, MlLa and Mlh, pathotypes were identified. In total, 23 pathotypes with virulence complexity ranging from 1 to 6 were identified. During both years in all populations of South Kazakhstan and Zhambyl regions pathotypes 24 and 64 mainly prevailed.

Conclusions

Obtained data suggest that low similarity of populations Bgh in Kazakhstan to European, African, Australian and South-East Asian populations. The present study provides a foundation for future studies on the pathogenic variability within of Bgh populations in Kazakhstan and addresses the knowledge gap on the virulence structure of Bgh in Central Asia. Complete effectiveness of the resistance genes, for which no corresponding virulence was found, will allow Kazakhstanean breeders to access many modern barley cultivars that those possessing the resistance effectiveness genes.

     

Comparison of Erysiphe cichoracearum and E. cruciferarum and a survey of 360 Arabidopsis thaliana accessions for resistance to these two powdery mildew pathogens

In previous work, UEA1 and UCSC1, two geographically distinct, powdery mildew isolates, were recognized for their ability to infect Arabidopsis thaliana. We have clarified the identity of these isolates by determining their host ranges, reexamining their morphology, and comparing their DNA sequences for the 5.8S ribosomal RNA and two flanking internal transcribed spacer sequences. These experiments confirm that UEA1 is a member of Erysiphe cruciferarum and that UCSC1 belongs to E. cichoracearum. Interactions of the two Erysiphe isolates with 360 A. thaliana accessions were examined to provide a comprehensive profile of naturally occurring powdery mildew resistance in this weedy species. The majority of A. thaliana accessions (213) were susceptible to both isolates. Among the accessions exhibiting some degree of resistance, most (84) responded differentially to UEA1 and UCSC1 and the remainder were resistant to both isolates. Notably, resistance to UCSC1 cosegregated with RPW7, a locus previously demonstrated to confer resistance to UEA1 in Ms-0 x Landsberg (erecta) crosses. With this large collection of resistant accessions, questions about species specificity, genetic diversity and the evolution of resistance to powdery mildews can be addressed.