Microsatellite analysis of Icelandic populations of the poplar fungal pathogen Melampsora larici-populina shows evidence of repeated colonization events

The basidiomycete Melampsora larici-populina causes foliar rust on Populus species from the sections Aigeiros and Tacamahaca, causing reduction in biomass production and economic losses. In the present study, samples of Icelandic M. larici-populina were collected for analysis of genetic diversity and population structure. A total of 439 isolates, collected at 15 locations, and analysed using 22 microsatellite markers were compared to data from French M. larici-populina populations. Twenty-one of the loci analysed were polymorphic, with an average of 3.4 alleles per locus. The mean observed and expected heterozygosities for all populations were 0.35 and 0.38. Evidence was found for a substructure within the Icelandic population with three subpopulations being the most likely scenario with low levels of gene flow. The population structure seen here is most likely shaped by both isolation and genetic drift as well as repeated events of colonization. In the future it can therefore be expected that regional poplar rust genotypes in Iceland change by two different modes; on one hand by transport of spores within the country and on the other hand by repeated colonization events. The results reported here underline the importance of closely monitoring the development of fungal diseases in Iceland, and to carefully select for resistance in Icelandic plant breeding programs.     

Low genetic diversity of Rhynchosporium commune in Iran,a secondary centre of barley origin

Rhynchosporium commune is a destructive pathogen of barley, causing leaf scald. Previous microsatellite studies used Syria as a representative of cultivated barley's centre of origin, the Fertile Crescent. These suggested that R. commune and Hordeum vulgare (cultivated barley) did not co‐evolve in the host's centre of origin. The present study compares R. commune populations from Syria with those from Iran, which represents a secondary centre of origin for barley at the eastern edge of the Iranian Plateau. Results from this study also suggest that R. commune and barley did not co‐evolve in the centre of origin of cultivated barley. This was evidenced by the low pathogen genetic diversity in Iran, which was even lower than in Syria, indicating that the pathogen may have been introduced recently into Iran, perhaps through infected barley seed. Hierarchical analyses of molecular variance revealed that most genetic diversity in Iran and Syria is distributed within populations, with only 14% among populations. Analyses of multilocus association, genotype diversity and mating type frequency suggest that Iranian populations reproduce predominantly asexually. The presence of both mating types on barley and uncultivated grasses suggest a potential for sexual reproduction. Rhynchosporium commune was also found on Hordeum murinum subsp. glaucum, H. vulgare subsp. spontaneum, Lolium multiflorum and, for the first time, on Avena sativa. The variety of wild grasses that can be infected with R. commune in Iran raises concerns of these grasses acting as evolutionary breeding grounds and sources of inoculum.     

How can research on pathogen population biology suggest disease management strategies? The example of barley scald (Rhynchosporium commune)

Barley scald fungus, Rhynchosporium commune, belongs to a host‐specialized species complex infecting grasses. Coalescent analyses of several genes indicate that R. commune originated c. 2500 years ago, after the domestication of barley. Phylogeographical analyses identified a diversity hotspot in Scandinavia, indicating an origin in northern Europe. After its emergence, R. commune became distributed globally on infected seeds, eventually invading the Fertile Crescent and infecting wild barley progenitors. Analyses of gene flow identified historical routes of migration out of Scandinavia and indicate a high degree of modern gene flow among South Africa, Australia and New Zealand. About 76% of global species diversity is found within barley fields, with all field populations showing a genetic structure consistent with sexual recombination. High levels of regional gene flow suggest wind‐dispersed ascospores and movement of infected seed. Quantitative traits, including pathogen aggressiveness, thermal sensitivity and fungicide resistance, showed high heritability and high levels of diversity within nine globally representative populations. Field experiments provided evidence for a fitness cost associated with complex virulence and a trade‐off between pathogenic and saprophytic fitness. All findings indicate that global R. commune populations have significant potential to evolve rapidly in response to environmental changes, including deployment of resistant cultivars, fungicide applications and global warming. Specific recommendations to improve management of barley scald include: (i) focus resistance breeding efforts in northern Europe, which offers the best location to screen germplasm and may provide useful sources of scald resistance; (ii) improve seed treatment, certification and quarantine programmes to limit long distance pathogen movement; (iii) manage barley stubble to decrease pathogen population size, limit production of sexual inoculum and reduce the pathogen's evolutionary potential.     

The Hordeum vulgare subsp. spontaneum–Blumeria graminis f. sp. hordei pathosystem: its position in resistance research and breeding applications

Barley (Hordeum vulgare L.) is an important cereal crop and powdery mildew, caused by Blumeria graminis f. sp. hordei, is one of the most serious diseases that occurs on barley throughout the world. In the Middle East, which is the centre of diversity for barley and its pathogens, the wild barley–powdery mildew pathosystem co-evolves resulting in many specific resistances in the host as well as corresponding virulences in the pathogen. Many specific resistances have been used in European breeding programmes and a centre of pathogen diversity has arisen also especially in central Europe. This short review briefly summarizes the use of host resistances derived from wild barley and land races including the durable resistance gene mlo. The use of powdery mildew pathogenicity for studying new and unknown specific resistances and for identifying resistances in commercial varieties is described. However, highly heterogeneous wild barley is also characterized as a valuable source of minor genes for powdery mildew resistance. These might be exploited by barley breeders especially for winter barley improvement where the non-specific resistance gene mlo cannot be used.     

High levels of genetic and genotypic diversity in field populations of the barley pathogen Ramularia collo-cygni

The ascomycete pathogen Ramularia collo-cygni causes Ramularia leaf spot (RLS) on barley. Although R. collo-cygni is considerd an emerging disease of barley, little is known about genetic diversity or population genetic structure of this pathogen. We applied a set of polymorphic AFLP (Amplified Fragment Length Polymorphism) markers to investigate population genetic structure in two Northern European populations of R. collo-cygni. The distribution of AFLP alleles revealed low levels of population subdivision and high levels of genetic diversity at both locations. Our analyses included 87 isolates and of these 84 showed a unique genotype pattern. The genetic structure of populations in Scotland and Denmark is highly similar and we find no evidence of population sub-division. An analysis of molecular variance was used to show that 86 % of the variance is attributable to within field genetic variance. In spite of the high levels of genetic and genotypic diversity in the R. collo-cygni populations, we find significant evidence of linkage disequilibrium among the AFLP alleles using a multilocus analysis. We propose that the high levels of genotypic diversity and the lack of population differentiation result from considerable levels of gene flow between populations most likely mediated by seed borne dispersal of inoculum.     

Assessment of the loose smut fungi (Ustilago nuda and U. tritici) in tissues of barley and wheat by fluorescence microscopy and real-time PCR

Loose smut fungi of barley and wheat (Ustilago nuda and U. tritici, respectively) colonize the plant without causing obvious disease symptoms before heading. The availability of diagnostic methods to detect and follow the growth of these pathogens in the plant would therefore be highly advantageous for both resistance breeding and the development of effective seed treatments. Using seed lots of barley and wheat highly infected with loose smut, we studied the early establishment of the loose smut pathogens in the plant by fluorescence microscopy. In hand-cut sections stained with the fluorochrome Blankophor?, fungal hyphae were observed to invade the shoot apical meristem and leaf primordia during the first days after the onset of germination. At the first node stage the ear and leaf primordia were generally extensively colonized. Hyphae of U. nuda were also regularly observed in high density in the nodes. A protocol was developed for the specific amplification of U. nuda and U. tritici DNA extracted from infected plant tissue. PCR screening of U nuda in seedlings from infected and healthy seed lots was compared to ELISA, microscopy and ultimately head infection of mature plants derived from tillers of the tested seedlings. The results indicated that a prediction of loose smut infection by real-time PCR is possible at the second leaf stage, and that the assay is equally suited for use with spring and winter varieties of barley and wheat.     

Monitoring für Getreidekrankheiten in Bayern – mehrjährige Auswertungen über die Bedeutung der einzelnen Schaderreger

Since the year 1991, up-to-date data about the actual infestation situations of the most important fungal diseases have been made available to the official advisory service and to the farmers by means of the “Monitoring for cereal diseases”. The classification of infestation is taken according to the scientifically elaborated thresholds of the Wheat Crop System Bayern and the Barley Crop System Bayern, respectively. The extensive data material allowed an evaluation over several years for the various cultivated areas. The least importance in winter wheat had eyespot, brown rust und yellow rust. Most exceedings of the thresholds were caused by Septoria tritici and Drechslera tritici-repentis. On about one third of the examined locations, exceedings of the threshold could already be noted before the growth stage “flag leaf just visible”. According to the infestation situation, in more than half of the cases one treatment would have been appropriate. In the past five years, the net blotch caused by Drechslera teres has turned out to be the most important disease in winter barley. With some difference in importance followed Rhynchosporium leaf blotch, powdery mildew and brown rust. At almost 80% of the locations, threshold exceedings were registered until the growth stage “flag life ligule just visible” (BBCH 39). In the majority of cases, the infestation situation makes a single treatment in winter barley appearing reasonable. In the years 2001–2004 the main pathogen in spring barley had been powdery mildew, followed by Drechslera teres and Rhynchosporium secalis. The exceedings of the thresholds had been spread over a broad range of growth stages. In the past five years the infestation of fungal diseases in triticale has been low. However since 2005 an increase of infestation, especially of powdery mildew, has been registered. The evaluation of the results from the “Monitoring for cereal diseases” over several years allows to estimate the risk of the different fungi in the regions. From that, conclusions about the fungicide intensities required in the average of cases can be drawn.     

What types of powdery mildew can infect wheat-barley introgression lines?

This work is a detailed study of the infection of fungal biotrophic pathogens causing powdery mildew diseases on introgression lines originating from the intergeneric hybridisation between wheat and barley (Triticum aestivum L. × Hordeum vulgare L.). Powdery mildew fungi are among the most widespread biotrophic pathogens of plants also and infect dicot and monocot species. Most powdery mildew species are strictly host specific. They colonize only a narrow range of species or one particular host species. The intergeneric hybridisation between wheat and barley could result in expansions of host ranges of the barley powdery mildew. Our experiments covered natural infections in the field and artificial infections under greenhouse conditions. Formae speciales of powdery mildew were identified on the basis of the sequencing results of ribosomal internal transcribed spacer sequences (rDNA-ITS). We identified Blumeria graminis f.sp. tritici isolate 14 (HM484334) on the wheat parent and all wheat-barley introgression lines and B. g. f. sp. hordei isolate MUMH1723 (AB 273556) on the barley parent, respectively. The wheat-barley introgression lines were inoculated with barley powdery mildew under greenhouse conditions. According to our results the added barley chromosomes (or segments) do not cause host range expansion of barley powdery mildew.     

Correlation between sensitivity of barley to Pyrenophora teres toxins and susceptibility to the fungus

Detached leaves of 25 barleys, ranging from highly susceptible to highly resistant to Pyrenophora teres f. teres and Pyrenophora teres f. maculata, were tested for their reaction to three phytotoxins isolated from cultures of the fungus: toxin A [ L, L - N -(2-amino-2-carboxyethyl)aspartic acid], toxin C (aspergillomarasmine A) and toxin B (anhydroaspergillomarasmine A). 0.75 m M toxin A caused mainly dark yellow chlorotic symptoms but little necrosis, whereas leaves treated with 0.25 m M toxin C developed distinct necrotic symptoms and zones of light yellow chlorosis. Toxin B is only weakly toxic, and toxin B and control solutions containing aspartic acid in the concentration of 0.75 m M did not cause any symptoms. The best differentiation between the barleys was obtained by scoring chlorosis after 120 h, and the optimal toxin concentrations for this differentiation were 0.75 m M toxin A and 0.25 m M toxin C, respectively. Results with different toxin concentrations inducing distinct variation in symptom expression indicate that the two toxins have different potencies as phytotoxins. The reaction of the barleys to toxins A and C correlated well with their reaction to infection by P. teres f. teres and P. teres f. maculata, suggesting that toxins A and C may be used to select resistant barley lines in the early stages of a breeding programme.     

Population genetic structure of four regional populations of the barley pathogen Pyrenophora teres f. maculata in Iran is characterized by high genetic diversity and sexual recombination

The leaf spot form of the barley disease net blotch, caused by the fungus Pyrenophora teres f. maculata (PTM), is an increasingly important foliar disease of barley. Studies of population genetic structure and reproductive mode are necessary to make predictions of the evolutionary potential of the pathogen. Sources of resistance to PTM have been found in Iranian landraces, which may have the potential to improve plant breeding efforts. However, little is known about the population genetic structure of this fungus in Iran. In this study, we analysed the frequency of the mating type genes to assess the potential for sexual mating of PTM collected from four provinces—Khuzestan, Hamadan, Golestan, and East Azerbaijan—and we investigated the population genetic structure using seven simple sequence repeat markers. High genotype diversity, linkage equilibrium, and equal ratios of mating types frequencies in the PTM populations at Khuzestan and Hamadan support the occurrence of sexual reproduction in these populations, while in Golestan and East Azerbaijan populations, significant gametic disequilibrium and relatively low genotype diversity suggest a higher incidence of clonality or different demographic histories. Unequal mating type frequencies in Golestan confirm a predominance of asexual reproduction. Finally, we found significant evidence for strong population structure with most of the genetic variation represented within regional populations (89%). Overall, our study provides evidence for high genetic variation in Iranian PTM populations, which may be the basis for rapid adaptive evolution in this pathosystem. This highlights the need for integrated efforts to control the disease.     

Population structure of South African and Australian Pyrenophora teres isolates

There are two recognized forms of the disease net blotch of barley: the net form caused by Pyrenophora teres f. teres (PTT) and the spot form caused by P. teres f. maculata (PTM). In this study, amplified fragment length polymorphism analysis was used to investigate the genetic diversity and population structure of 60 PTT and 64 PTM isolates collected across Australia (66 isolates) and in the south‐western Cape of South Africa (58 isolates). For comparison, P. tritici‐repentis, Exserohilum rostratum and Bipolaris sorokiniana samples were also included in the analyses. Both distance‐ and model‐based cluster analyses separated the PTT and PTM isolates into two strongly divergent genetic groups. Significant variation was observed both among the South African and Australian populations of PTT and PTM and among sampling locations for the PTT samples. Results suggest that sexual reproduction between the two forms is unlikely and that reproduction within the PTT and PTM groups occurs mainly asexually.     

Genetic structure of <Emphasis Type="Italic">Pyrenophora teres</Emphasis> f. <Emphasis Type="Italic">teres</Emphasis> and <Emphasis Type="Italic">P. teres</Emphasis> f. <Emphasis Type="Italic">maculata</Emphasis> populations from western Canada

Infection by Pyrenophora teres f. teres (Ptt) or P. teres f. maculata (Ptm), the causal agents of the net and spot forms of net blotch of barley, respectively, can result in significant yield losses. The genetic structure of a collection of 128 Ptt and 92 Ptm isolates from the western Canadian provinces of Alberta (55 Ptt, 27 Ptm), Saskatchewan (58 Ptt, 46 Ptm) and Manitoba (15 Ptt, 19 Ptm) were analyzed by simple sequence repeat (SSR) marker analysis. Thirteen SSR loci were examined and found to be polymorphic within both Ptt and Ptm populations. In total, 110 distinct alleles were identified, with 19 of these shared between Ptt and Ptm, 75 specific to Ptt, and 16 specific to Ptm. Genotypic diversity was relatively high, with a clonal fraction of approximately 10 % within Ptt and Ptm populations. Significant genetic differentiation (PhiPT = 0.230, P = 0.001) was found among all populations; 77 % of genetic variation occurred within populations and 23 % between populations. Lower, but still significant genetic differentiation (PhiPT = 0.038, P = 0.001) was detected in Ptt, with 96 % of genetic variation occurring within populations. No significant genetic differentiation (PhiPT = 0.010, P = 0.177) was observed among Ptm populations. Isolates clustered in two distinct groups conforming to Ptt or Ptm, with no intermediate cluster. The high number of haplotypes observed, combined with an equal mating type ratio for both forms of the fungus, suggests that P. teres goes through regular cycles of sexual recombination in western Canada.     

Control of Stagonospora nodorum and Septoria tritici in Wheat by Pre-treatment with Drechslera teres, a Non-host Pathogen

A field study is described which explored the possibility of controlling Stagonospora nodorum and Septoria tritici on wheat using a barley pathogen, Drechslera teres. Pre-treatment of wheat cv. Hussar flag leaves with D. teres resulted in a significant reduction in disease caused by S. nodorum and S. tritici, resulting in a significant increase in grain yield. When cv. Brigadier leaves were treated with D. teres prior to inoculation with S. nodorum there was an initial increase in disease expression whilst D. teres had no effect on symptoms produced by S. tritici on cv. Brigadier. There was significantly less disease on leaves of cvs. Hussar and Brigadier pre-treated with D. teres prior to inoculation with an equal mixture of S. nodorum and S. tritici compared to plants pre-treated with water. It is concluded that D. teres and other non-host pathogens show potential as biological control agents for S. nodorum and S. tritici.     

An integrated multivariate approach to net blotch of barley: Virulence quantification,pathotyping and a breeding strategy for disease resistance

Knowledge of pathotype diversity and virulence in local populations of Pyrenophora teres is a prerequisite to screening for durable resistance to net blotch. The current study aimed to quantify the virulence level of Moroccan isolates, identify and designate existing pathotypes, and select resistant genotypes. We developed a method for virulence quantification of P. teres isolates based on a conversion of infection responses into frequencies for use in correspondence analysis. Coordinates of the first axis of this analysis had a virulence spectrum and ranked isolates from virulent to avirulent. Mixed model analysis was also devised for virulence quantification. Coordinates of the first dimension of correspondence analysis were linearly correlated to BLUPs (Best Linear Unbiased Predictors) of the mixed model. A genotype by genotype by environment model (GGE) coupled with cluster analysis differentiated P. teres isolates into ten and nine pathotypes for net- and spot-forms respectively. Populations of these two forms were dissimilar in terms of classes of virulence. For P. teres f. maculata, avirulent, moderately virulent and highly virulent isolates represented one-third of the population, whereas 90% of P. teres f. teres population was composed of avirulent to moderately avirulent isolates. Barley differential sets were subsequently reduced to two new sets that simplified pathotyping through a key code based on resistant or susceptible reactions. Dendrograms of cluster analysis based on GGE analysis depicted the stability of a genotype’s reactions across all isolates, and using only resistant cultivars as sources of resistance to control net blotch disease would, based on this analysis, fail to control all pathotypes. Therefore, we propose an alternative breeding strategy to control net blotch effectively.     

Classification and parasitic specialization of blast fungi

Pyricularia oryzae (Magnaporthe oryzae), a causal agent of blast diseases on staple gramineous crops, is a model organism listed as the most important economically and scientifically of the top 10 fungal pathogens by fungal molecular pathologists. Although we are now in an era of genome-enabled analysis, we need to understand the history of the pathogen’s taxonomy, classification, and parasitic specialization in addition to recent research advances. In this review, we focus on these rather fundamental topics. First, the history of classification, including the discovery of its sexual stage and designation, is overviewed. Based on recent results of phylogenetic analysis of Magnaporthaceae isolates, blast fungi are suggested to constitute a robust population that is not congeneric with Magnaporthe salvinii, the type species of Magnaporthe. Second, genetic mechanisms involved in its parasitic specialization into host-specific subgroups and races are outlined. Implications of recent molecular data for resistance breeding are discussed.     

Incidence and diversity of the fungal genera Aspergillus and Penicillium in Portuguese almonds and chestnuts

Almonds (Prunus dulcis (Miller) D.A. Webb) and European (sweet) chestnuts (Castanea sativa Miller) are of great economic and social impact in Mediterranean countries, and in some areas they constitute the main income of rural populations. Despite all efforts to control fungal contamination, toxigenic fungi are ubiquitous in nature and occur regularly in worldwide food supplies, and these nuts are no exception. This work aimed to provide knowledge on the general mycobiota of Portuguese almonds and chestnuts, and its evolution from field to the end of storage. For this matter, 45 field chestnut samples and 36 almond samples (30 field samples and six storage samples) were collected in Trás-os-Montes, Portugal. All fungi belonging to genus Aspergillus were isolated and identified to the section level. Fungi representative of other genera were identified to the genus level. In the field, chestnuts were mainly contaminated with the genera Fusarium, Cladosporium, Alternaria and Penicillium, and the genus Aspergillus was only rarely found, whereas almonds were more contaminated with Aspergillus. In almonds, Aspergillus incidence increased significantly from field to the end of storage, but diversity decreased, with potentially toxigenic isolates belonging to sections Flavi and Nigri becoming more significant and widespread throughout storage. These fungi were determined to be moderately associated, which can be indicative of mycotoxin co-contamination problems if adequate storage conditions are not secured.     

Genetic structure of a Pyrenophora teres f. teres population over time in an Australian barley field as revealed by Diversity Arrays Technology markers

Net form of net blotch caused by Pyrenophora teres f. teres (Ptt) is a major foliar disease of barley (Hordeum vulgare) worldwide. Knowledge of the evolution of Ptt pathogen populations is important for development of durable host-plant resistance. This study was conducted to investigate changes in genetic structure of a Ptt population within a barley field during three cropping years. The susceptible barley cultivar Henley was inoculated with Ptt isolate NB050. Leaf samples were collected during the years 2013–15 and 174 single spore Ptt isolates stored. Genotyping using Diversity Arrays Technology markers identified that 25% of isolates were clones of the inoculated isolate and 75% of isolates were multilocus genotypes (MLGs) differing from the original inoculated genotype. The novel genotypes probably originated from a combination of windborne spores from neighbouring fields, infected seed and sexual recombination in the field. The rapid change in the genotypic composition of the Ptt population in this study suggests adaptive potential of novel genotypes and demonstrates the need for barley breeders to use multiple sources of host-plant resistance to safeguard against resistance being overcome.     

Morphological and molecular analysis of <Emphasis Type="Italic">Paratrichodorus teres</Emphasis> (Hooper 1962) (Nematoda: Trichodoridae): a groundwork for discussion on the phylogeny and pathogenicity of <Emphasis Type="Italic">Paratrichodorus</Emphasis> species

Due to its ability to transmit plant viruses, Paratrichodorus teres (Hooper in Nematologica, 7, 273–280, 1962) is recognized as an economically important trichodorid species. Morphological and molecular analyses (18S and 28S rDNA) were performed, and 10 new plant hosts are reported for Polish P. teres populations. Major morphological features and the measurements obtained for the investigated specimens were within the wide ranges indicated for this species. However, a more detailed comparative analysis of Polish and Iranian P. teres showed significant morphological differences, particularly, in the shape and the structure of the walls of pars proximalis vaginae and the shape of the rectum.Phylogenetic study based on the 18S rDNA data suggests positioning of the Polish P. teres sequences within a cluster of sequences originating from the Netherlands. A comparison of the 28S rDNA fragment from Polish populations with the only P. teres 28S rDNA sequence available (from Iran) in GenBank revealed a sequence variability of 9.3%. The variation across these two representatives was higher than in the case of many other pairs of Trichodoridae species. The results obtained on the Polish P. teres specimens are discussed in the framework of the species taxonomy and phylogenetic relationships.     

Genetic variation of single nucleotide polymorphisms identified at the mating type locus correlates with form-specific disease phenotype in the barley net blotch fungus Pyrenophora teres

Mating-type (MAT) locus-specific single nucleotide polymorphisms (SNPs) have been shown to be sufficient for conventional PCR-based differentiation of Pyrenophora teres f. teres (Ptt) and P. teres f. maculata (Ptm), the cause of the net and spot form, respectively, of barley net blotch. Here, we report the cloning and characterization of the MAT locus from 10 California isolates that cause atypical blotch symptoms on barley. Analysis of the full-length nucleotide sequences of one MAT1-1 (1,993?bp) and nine MAT1-2 (2,149 or 2,161?bp) idiomorphs revealed high (98?C99?%) similarity to those of Ptt isolates. However, distinct SNP patterns were identified in the newly cloned MAT idiomorphs. Two new MAT1-2-specific SNPs were found to be conserved in one Australia and eight California isolates that all cause similar atypical blotch symptoms. Phylogenetic analysis indicated that all 10 California isolates form a separate branch (or clade) within the Ptt group, except for one that appears to be ancestral to both Ptt and Ptm. PCR primers designed based on the identified SNP patterns were used successfully to differentiate each atypical isolate from highly virulent forms. This study extends our previous work and, taken together, the results demonstrate that the genetic variation at the MAT locus correlates with variation in the form-specific disease phenotype, and that MAT-specific SNPs can serve as reliable and convenient markers for subspecies level differentiation in P. teres, an economically important plant-pathogenic ascomycete (http://en.wikipedia.org/wiki/Single_nucleotide_polymorphism).     

Susceptibility-inducing factor (suppressor) from Blumeria graminis f. sp. hordei has no effect on the primary infection of the fungus

When fungal germlings, after forming haustoria of Blumeria graminis f. sp. hordei (B. graminis), were removed from the surfaces of barley coleoptiles by cellulose acetate, followed by challenge inoculation with the non-pathogen Erysiphe pisi, they infected the nonhost barley coleoptile cells. This phenomenon was not observed on the coleoptile surface when the fungal germlings of B. graminis were removed before the formation of haustoria. Also, when the surface was inoculated with the pathogen of barley B. graminis as a challenger, after removing the fungal germlings of inducer post haustorial formation, the penetration efficiency of the fungi increased significantly compared with that of the control. Furthermore, when we extracted the crude-susceptibility inducing factor (suppressor) from coleoptiles before and after the formation of haustoria of B. graminis, suppressor activity against infection with E. pisi was observed only in the extract of barley coleoptiles that included haustoria of B. graminis about 18 h or later after inoculation. Surprisingly, however, the extract did not increase the penetration efficiency of B. graminis significantly. Thus, we hypothesize that the suppressor extracted from barley coleoptiles in which B. graminis had formed haustoria has no effect on increasing the penetration efficiency of the primary infection from the appressorium of B. graminis but has an effect on the infection with non-pathogen E. pisi.