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At least 19 recordsLinked to original sources

PubMed2006

A proteomic approach to study pea (Pisum sativum) responses to powdery mildew (Erysiphe pisi).

As a global approach to gain a better understanding of the mechanisms involved in pea resistance to Erysiphe pisi, changes in the leaf proteome of two pea genotypes differing in their resistance phenotype were analyzed by a combination of 2-DE and MALDI-TOF/TOF MS. Leaf proteins from control non-inoculated and inoculated susceptible (Messire) and resistant (JI2480) plants were resolved by 2-DE, with IEF in the 5-8 pH range and SDS-PAGE on 12% gels. CBB-stained gels revealed the existence of quantitative and qualitative differences between extracts from: (i) non-inoculated leaves of both genotypes (77 spots); (ii) inoculated and non-inoculated Messire leaves (19 spots); and (iii) inoculated and non-inoculated JI2480 leaves (12 spots). Some of the differential spots have been identified, after MALDI-TOF/TOF analysis and database searching, as proteins belonging to several functional categories, including photosynthesis and carbon metabolism, energy production, stress and defense, protein synthesis and degradation and signal transduction. Results are discussed in terms of constitutive and induced elements involved in pea resistance against Erysiphe pisi.

Ascomycota↗

PubMed1983

A microapparatus for liquid hydrogen fluoride solvolysis: sugar and amino sugar composition of Erysiphe graminis and Triticum aestivum cell walls.

The assembly and use of a simple and safe apparatus for HF solvolysis of microgram amounts of cell walls, polysaccharides, or glycoproteins are described. Using this apparatus the cell wall composition of Erysiphe graminis was compared with that of its wheat host. The HF solvolysis combined with TFA posthydrolysis considerably increased sugar yields compared with TFA hydrolysis alone, due mainly to increased yields of glucose from wheat, and glucosamine from Erysiphe, corresponding to cellulose and chitin, respectively. A potentially useful method for determining amounts of fungal hyphae in plant tissue is also provided.

Amino Sugars↗

PubMed1999

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.

Arabidopsis↗

PubMed1981

Hydroxyproline Enhancement as a Primary Event in the Successful Development of Erysiphe graminis in Wheat.

Host cell wall hydroxyproline enhancement was observed in the successful development of the parasite Erysiphe graminis DC. f. sp. tritici em Marchal (MS-1) on wheat (em Thell). Hydroxyproline enhancement, which was observed only in susceptible hosts, was detected as early as 25 hours after infection. This observation suggests that the increase in cell wall hydroxyproline is a primary event in the host-pathogen interaction of Erysiphe graminis in wheat.

Journal Article↗

PubMed2003

Evaluation of Erysiphe graminis f sp tritici field isolates for resistance to strobilurin fungicides with different SNP detection systems.

A single nucleotide polymorphism (snp) in the cytochrome b gene confers resistance to strobilurin fungicides in Erysiphe graminis DC fsp tritici Marchal. On the basis of this point mutation three different types of molecular markers have been developed. Cleaved amplified polymorphic sequences and allele-specific PCR were used to score resistant and sensitive isolates from specifically selected regional populations across Europe. The results of molecular tests were in total agreement with the resistance phenotypes revealed by in vivo tests. Serial dilutions of mixed samples (resistant/sensitive) delimited the detection for strobilurin-resistant alleles to a range of 10-50% for both marker classes. Due to these detection limits no mixture of mitochondria within individual isolates was found. Denaturing high performance chromatography was used to increase the detection sensitivity for the mutant allele. Although the detection limit was lowered to 5-10%, there was no evidence for the existence of mixed mitochondrial genotypes.

Drug Resistance, Fungal↗

PubMed1993

SINE-like properties of a highly repetitive element in the genome of the obligate parasitic fungus Erysiphe graminis f.sp. hordei.

The genomic organization of repetitive DNA in the obligate parasitic fungus Erysiphe graminis DC ex Mérat f.sp. hordei Em. Marchal was investigated using a cosmid library of the fungal genome. Three repetitive sequences were shown to be dispersed throughout the genome, and in a few cases they were found closely associated with long poly(dA) tracts. The most prevalent sequence is 903 bp long and accounts for at least 5% of the genome. Sequence analysis revealed features resembling mammalian Short INterspersed Elements (SINEs), namely the presence of a poly(dA) tail (33 bp), flanking direct repeats (13 bp), putative "A" and "B" blocks for RNA polymerase III binding; the corresponding transcript would be capable of forming a complex secondary structure.

Ascomycota↗

PubMed1994

Genetic analysis of DNA fingerprints and virulences in Erysiphe graminis f.sp. hordei.

A DNA probe, E9, which has been used extensively in population genetic studies of the barley powdery mildew pathogen, Erysiphe graminis f.sp. hordei, was shown to be homologous to dispersed sequences in the genome of this fungus. In a cross of the isolates CC52 and DH14, fragments with homology to E9 mapped to six clusters of loci. Avirulences matching five resistance genes in barley were controlled by single genes, in accordance with the gene-for-gene hypothesis, while avirulence matching a sixth resistance gene, Mla13, was controlled by two genes. A gene which controls the response to a fungicide, ethirimol, was not linked to any other gene. In all, seven linkage groups, comprising 22 loci, were detected. The results indicate that E9 can be used to identify members of a clone of E.g. f.sp. hordei, but should not be used for quantitative population genetic analysis.

Ascomycota↗

PubMed1996

Characterization of the transcript of a new class of retroposon-type repetitive element cloned from the powdery mildew fungus, Erysiphe graminis.

The putative master transcript of a novel class of repetitive element has been cloned from the fungus erysiphe graminis f.sp. hordei. Sequence analysis of the cDNA revealed that the element, designated Eg-R1, is a member of the retroposon superfamily with properties in common with SINEs and LINEs (short or long interspersed elements). SINE-like properties include the transcript size (approximately 700 bp), and the lack of major open reading frames. In contrast, the fact that the transcript is polyadenylated and is most probably transcribed by RNA polymerase II, suggests a functional relationship to LINEs. Except for a short, but striking, sequence identity to a published SINE from the same fungus, no similar sequence was found in database searches. A constitutively high transcript level is found throughout the asexual life cycle of the fungus. Small differences in band patterns of Southern blots were observed between two isolates of E. graminis f.sp. hordei, while the band patterns in an isolate of the very close relative E. graminis f.sp. tritici in general appear dissimilar. This may imply that the element is currently active. Recent dispersal is confirmed by the observation that an approximately 550 bp internal hinfI fragment is conserved in the majority of the copies in all three isolates. Approximately 50 copies are present in E. graminis f.sp. hordei.

Ascomycota↗

PubMed2002

Plant growth-promoting rhizobacteria-mediated induction of phenolics in pea ( Pisum sativum) after infection with Erysiphe pisi.

Qualitative and quantitative estimation of phenolic compounds was done through high performance liquid chromatography (HPLC) in different parts of pea ( Pisum sativum) after treatment with two plant growth-promoting rhizobacteria (PGPR), viz., Pseudomonas fluorescens (strain Pf4) and Pseudomonas aeruginosa (referred to here as Pag) and infection by Erysiphe pisi. The phenolic compounds detected were tannic, gallic, ferulic, and cinnamic acids on the basis of their retention time in HPLC. In all the treated plants, synthesis of phenolic compounds was enhanced. The induction of gallic, ferulic, and cinnamic acids was manyfold more than those in the control. Maximum accumulation of phenolic compounds was observed in plants raised from PGPR-treated seeds and infection with E. pisi. Under pathogenic stress, Pag performed better because a relatively higher amount of phenolics was induced compared with plants treated with Pf4.

Ascomycota↗

PubMed1997

Disparate sequence characteristics of the Erysiphe graminis f.sp. hordei glyceraldehyde-3-phosphate dehydrogenase gene.

The Erysiphe graminis f.sp. hordei (Egh) glyceraldehyde-3-phosphate dehydrogenase (gpd) gene was isolated and characterized. It contains typical promoter elements and has three introns, one of which is positioned in the 5' untranslated region of the gene. The deduced amino-acid sequence has 87% similarity to gpd genes from other Ascomycete fungi. This is at the same level as previously estimated among these fungi. Comparison at the DNA level reveal similarities of only around 70%, which is 10% lower than previously reported. In an evolutionary tree based on the sequences from 18 fungal gpd genes, Egh falls into the group of Ascomycetes located at a basal position. The regulatory region of the Egh gpd gene has no homology to corresponding sequences in other filamentous Ascomycetes. Codon usage was determined for the four characterized Egh genes (tub2, Egh7, Egh16 and gpd) and found to be similar for all four genes. The results of the codon-usage analysis suggest that Egh is more flexible than other fungi in the choice of nucleotides at the wobble position. Codon-usage preferences in Egh and barley genes indicate a level of difference which may be exploited to discriminate between fungal and plant genes in sequence mixtures. The Egh gpd promoter appears to be superior to that of the Egh beta-tubulin gene (tub2) for driving the E. coli beta-glucuronidase (GUS) gene in transformation experiments.

Ascomycota↗

PubMed1998

PCR cloning and detection of point mutations in the eburicol 14alpha-demethylase (CYP51) gene from Erysiphe graminis f. sp. hordei, a "recalcitrant" fungus.

Molecular studies of some micro-organisms are hampered by the difficulty of obtaining sufficient amounts of nucleic acids. A cloning strategy based on PCR has therefore been used to clone the eburicol 14alpha-demethylase (CYP51) gene of the obligate fungus Erysiphe graminis f. sp. hordei (Egh) using minute amounts of genomic DNA. The CYP51 gene encodes the enzymatic target of a major group of fungicides. Sequencing CYP51 from different Egh isolates revealed the occurrence of two alleles for this gene. An allele-specific PCR assay was developed to detect each CYP51 allele.

Alleles↗

PubMed1994

Genome manipulation in recalcitrant species: construction and characterization of a yeast artificial chromosome (YAC) library from Erysiphe graminis f. sp. hordei, an obligate fungal pathogen of barley.

Extraction of DNA from organisms where spores are the only source of pure material is a major problem. Methods are described which allow the isolation of high-M(r) DNA, from small quantities of Erysiphe graminis f. sp. hordei (Egh) conidia, suitable for cloning in yeast artificial chromosomes (YACs). A YAC library of 1500 clones was constructed in the vectors, pYAC4 and pYACRC. The average size of YAC inserts is 220 kb and range from 70 to 500 kb, providing ten haploid genome equivalents. Multicopy RFLP markers and an Egh-specific repetitive SINE element were used to characterize the library. The SINE element is effective in fingerprint analysis and contig assembly. Four out of five representative clones containing more than one YAC were mitotically unstable.

Ascomycota↗

PubMed1996

Isolation and characterization of two novel genes expressed in germinating conidia of the obligate biotroph Erysiphe graminis f.sp. hordei.

A cDNA library was constructed from germinating conidia of the obligate biotrophic fungus, Erysiphe graminis DC ex Mérat f.sp. hordei Em. Marchal (Egh). Subtractive hybridization and differential screening were carried out. Two cDNA clones, cEgh7 and cEgh16, which were highly expressed in germinating conidia, but not in ungerminated conidia, were selected for further characterization. The corresponding genomic sequences, gEgh7 and gEgh16, were isolated from a cosmid library and sequenced. The gEgh7 gene contains an open reading frame (ORF) that codes for a 249-amino-acid (aa), Pro-rich polypeptide with a repeated primary structure. Expression studies in planta indicated that gEgh7 may have a function in the development and maturation of conidia. The ORF of gEgh16 is interrupted by two introns of 91 and 119 bp. It encodes a 251-aa polypeptide of unknown function. This gene belongs to a multigene family and is expressed during all developmental stages of Egh in planta and may be associated with hyphal growth.

Amino Acid Sequence↗

PubMed2005

Isolation, cloning and expression analysis of EcPMA1, a putative plasma membrane H+ -ATPase transporter gene from the biotrophic pathogenic fungus Erysiphe cichoracearum.

Little is known at the molecular level about the transporters involved in nutrient transfer in the plant/powdery mildew interaction. A PCR-based approach was used to identify and isolate a partial-length cDNA coding for an isoform of the plasma membrane H+ -ATPase (EcPMA1) in the biotrophic pathogenic fungus Erysiphe cichoracearum. Southern analysis suggests that EcPMA1 exists as a single-copy gene. Sequence analysis indicated a high similarity of EcPMA1 to other fungal H+ -ATPases. Expression of EcPMA1 increases in infected Arabidopsis leaves as the disease progresses, correlating with the growth of the pathogen.

Amino Acid Sequence↗

PubMed2006

Detection and quantification of Erysiphe necator DNA in wine grapes and resultant must and juice.

Powdery mildew of grapevines is difficult to assess visually at the weighbridge, particularly in large consignments of machine-harvested fruit. To facilitate accurate methods for the detection and quantification of the disease in grape samples obtained from both the vineyard and winery, we developed a DNA probe for the pathogen Erysiphe necator. The E. necator-specific 450 bp DNA fragment pEnA1, targets highly repetitive sequences and was isolated from a partial genomic library. In screening for species specificity, clone pEnA1 was used in slot-blot hybridization and detected E. necator DNA from grapes and resultant must and juice, but not from clarified juice and wine. The detection threshold was approximately 50 pg of E. necator DNA per 100 ng total DNA of grape sample and was equivalent to 1-5% of a grape bunch visually affected by powdery mildew. Disease severity, expressed as the percentage of surface area of a bunch with powdery mildew, and E. necator DNA content were highly correlated, r2=0.955, P<0.001. The DNA-based hybridization assay has the potential to predict the severity of powdery mildew in grape samples from the vineyard and in must and juice samples at the winery. The DNA sequence of clone pEnA1 was used to design species-specific primers, the results maintaining the same specificity patterns observed in the initial hybridization assays. The PCR-based assay was sensitive enough to detect approximately 1 pg DNA, being equivalent to 1 conidium per sample. This is the first report to date of the detection of all known phenetic groups of E. necator DNA and of the quantification of DNA from grape samples at the winery. Accurate information on the extent of powdery mildew contamination of grape lots would enable wineries to make more informed decisions about the use of fruit and must.

Ascomycota↗

PubMed2000

Transgenic carrots with enhanced resistance against two major pathogens, Erysiphe heraclei and Alternaria dauci.

In vitro assay indicated that the human lysozyme has lytic activity against phytopathogenic fungi and bacteria. A human lysozyme gene was placed under control of the constitutive CaMV 35S promoter and the resulting expression plasmid was introduced into two cultivars (cv.) of carrot, Kurodagosun (K5) and Nantes Scarlet (NS), by Agrobacterium tumefaciens-mediated method. Seven and fourteen transgenic plants of cv. K5 and cv. NS were regenerated, respectively, and the obtained transgenic carrots of T0 generation was tested for disease resistance against Erysiphe heraclei, a pathogenic fungi causing powdery mildew. Among the tested lines, the transgenic plant No. 12-1 and 8-1 of cv. NS showed a fairly strong resistance against E. heraclei. The strong disease resistance was also confirmed in T1 generation. Disease resistance against another pathogen of leaf blight, Alternaria dauci, were also tested using T1 transgenic lines. Significant enhanced resistance was observed in the No. 12-1 of cv. NS. Accumulation of synthesized human lysozyme protein was observed in this line, a finding consistent with observed disease resistance.

Journal Article↗

PubMed2000

Stable transformation of erysiphe graminis an obligate biotrophic pathogen of barley.

Barley powdery mildew, Erysiphe graminis f.sp. hordei, is an obligate biotrophic pathogen and as such cannot complete its life cycle without a living host. The inability to transform this fungus and manipulate its genome has constrained research towards understanding its life cycle and pathogenicity. Here we describe an in planta transformation system based on delivery of DNA using a gold-particle gun and selection using benomyl or bialaphos. Using this method, we consistently obtained stable transformants with efficiencies comparable to other filamentous fungi. Stable expression of the beta-glucuronidase in E. graminis was demonstrated by co-transforming the uidA gene with the selectable markers.

Ascomycota↗

PubMed2001

Effect of calcium and calmodulin modulators on the development of Erysiphe pisi on pea leaves.

The effect of calcium and calmodulin modulators, viz., ethylene glycol bis (beta-amino ethyl ether)-N,N,N',N'-tetraacetic acid (EGTA), a calcium chelator; verapamil, a plasma membrane Ca2+ channel blocker; ruthenium red, an organelle Ca2+ channel blocker; and chlorpromazine, a calmodulin antagonist; on the development of Erysiphe pisi was studied by floating the inoculated leaves on the respective solutions of chemicals. All the modulators affected the development of E. pisi by inhibiting the colony diameter, number of secondary branches, number of hyphal cells per colony and number of haustoria. The calmodulin antagonist was more effective in reducing E. pisi development. The results suggest the possible involvement of calcium and calmodulin in the development of E. pisi on pea leaves.

Ascomycota↗