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Pierre J. G. M. De Wit - One of the best experts on this subject based on the ideXlab platform.

  • A survey on occurrence of Cladosporium fulvum identifies race 0 and race 2 in tomato-growing areas of Argentina
    Plant disease, 2015
    Co-Authors: Roció Medina, Pierre J. G. M. De Wit, Silvina Marianela Yanil López, Mario Emilio Ernesto Franco, Cristina Rollan, Blanca Lía Ronco, Mario Carlos Nazareno Saparrat, Pedro Alberto Balatti
    Abstract:

    The presence of Cladosporium fulvum (syn. Passalora fulva), causal agent of tomato leaf mold, was confirmed in the two main greenhouse-production areas for tomato in Argentina. Using both morphological characters and internal transcribed spacer sequencing, we confirmed the presence of physiological races of this pathogen. A diagnostic multiplex polymerase chain reaction (PCR) was also developed, using primers derived from C. fulvum avirulence (Avr) genes. In all, 20 isolates of Cladosporium spp. were obtained as monospore cultures and 12 were identified as C. fulvum. By this method, we showed that, of these 12 isolates, 5 were race 0 (carrying functional Avr2, Avr4, Avr4E, and Avr9 genes) and 7 were race 2 (lacking the Avr2 gene). Race identity was confirmed by testing their virulence on a set of tomato differentials carrying different Cf resistance genes. All Avr genes could be amplified in single or multiplex PCR using DNA isolated from in vitro grown monospore cultures but only three Avr could be amplified when genomic DNA was isolated from C. fulvum-infected necrotic leaf tissue.

  • A survey on occurrence of Cladosporium fulvum identifies race 0 and race 2 in tomato-growing areas of Argentina
    'Scientific Societies', 2015
    Co-Authors: Medina Rocío, Pierre J. G. M. De Wit, López, Silvina Marianela Yanil, Franco, Mario Emilio Ernesto, Rollán, María Cristina, Saparrat, Mario Carlos Nazareno, Ronco, Blanca L., Balatti, Pedro Alberto
    Abstract:

    The presence of Cladosporium fulvum (syn. Passalora fulva), causal agent of tomato leaf mold, was confirmed in the two main greenhouseproduction areas for tomato in Argentina. Using both morphological characters and internal transcribed spacer sequencing, we confirmed the presence of physiological races of this pathogen. A diagnostic multiplex polymerase chain reaction (PCR) was also developed, using primers derived from C. fulvum avirulence (Avr) genes. In all, 20 isolates of Cladosporium spp. were obtained as monospore cultures and 12 were identified as C. fulvum. By this method, we showed that, of these 12 isolates, 5 were race 0 (carrying functional Avr2, Avr4, Avr4E, and Avr9 genes) and 7 were race 2 (lacking the Avr2 gene). Race identity was confirmed by testing their virulence on a set of tomato differentials carrying different Cf resistance genes. All Avr genes could be amplified in single or multiplex PCR using DNA isolated from in vitro grown monospore cultures but only three Avr could be amplified when genomic DNA was isolated from C. fulvum-infected necrotic leaf tissue.Fil: Medina, Rocio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Investigación y Desarrollo en Fermentaciones Industriales. Universidad Nacional de la Plata. Facultad de Ciencias Exactas. Centro de Investigación y Desarrollo en Fermentaciones Industriales; Argentina. Universidad Nacional de la Plata. Facultad de Ciencias Agrarias y Forestales. Departamento de Ciencias Biológicas. Centro de Investigaciones de Fitopatología. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigaciones de Fitopatología; ArgentinaFil: López, Silvina Marianela Yanil. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Fisiología Vegetal. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Instituto de Fisiología Vegetal; ArgentinaFil: Franco, Mario Emilio Ernesto. Universidad Nacional de la Plata. Facultad de Ciencias Agrarias y Forestales. Departamento de Ciencias Biológicas. Centro de Investigaciones de Fitopatología. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigaciones de Fitopatología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Rollán, María Cristina. Universidad Nacional de la Plata. Facultad de Ciencias Agrarias y Forestales. Departamento de Ciencias Biológicas. Centro de Investigaciones de Fitopatología. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigaciones de Fitopatología; ArgentinaFil: Ronco, Blanca L.. Universidad Nacional de la Plata. Facultad de Ciencias Agrarias y Forestales. Departamento de Ciencias Biológicas. Centro de Investigaciones de Fitopatología. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigaciones de Fitopatología; ArgentinaFil: Saparrat, Mario Carlos Nazareno. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Fisiología Vegetal. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Instituto de Fisiología Vegetal; ArgentinaFil: Wit, Pierre J. G. M. De. Wageningen University; Países BajosFil: Balatti, Pedro Alberto. Universidad Nacional de la Plata. Facultad de Ciencias Agrarias y Forestales. Departamento de Ciencias Biológicas. Centro de Investigaciones de Fitopatología. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigaciones de Fitopatología; Argentin

  • The Genomes of the Fungal Plant Pathogens Cladosporium fulvum and Dothistroma septosporum Reveal Adaptation to Different Hosts and Lifestyles But Also Signatures of Common Ancestry
    PLoS Genetics, 2012
    Co-Authors: Pierre J. G. M. De Wit, Ate Van Der Burgt, Bilal Okmen, Ioannis Stergiopoulos, Kamel A. Abd-elsalam, Andrea L. Aerts, Ali H. Bahkali, Henriek G. Beenen, Pranav Chettri, Murray P. Cox
    Abstract:

    We sequenced and compared the genomes of the Dothideomycete fungal plant pathogens Cladosporium fulvum (Cfu) (syn. Passalora fulva) and Dothistroma septosporum (Dse) that are closely related phylogenetically, but have different lifestyles and hosts. Although both fungi grow extracellularly in close contact with host mesophyll cells, Cfu is a biotroph infecting tomato, while Dse is a hemibiotroph infecting pine. The genomes of these fungi have a similar set of genes (70% of gene content in both genomes are homologs), but differ significantly in size (Cfu >61.1-Mb; Dse 31.2-Mb), which is mainly due to the difference in repeat content (47.2% in Cfu versus 3.2% in Dse). Recent adaptation to different lifestyles and hosts is suggested by diverged sets of genes. Cfu contains an alpha-tomatinase gene that we predict might be required for detoxification of tomatine, while this gene is absent in Dse. Many genes encoding secreted proteins are unique to each species and the repeat-rich areas in Cfu are enriched for these species-specific genes. In contrast, conserved genes suggest common host ancestry. Homologs of Cfu effector genes, including Ecp2 and Avr4, are present in Dse and induce a Cf-Ecp2- and Cf-4-mediated hypersensitive response, respectively. Strikingly, genes involved in production of the toxin dothistromin, a likely virulence factor for Dse, are conserved in Cfu, but their expression differs markedly with essentially no expression by Cfu in planta. Likewise, Cfu has a carbohydrate-degrading enzyme catalog that is more similar to that of necrotrophs or hemibiotrophs and a larger pectinolytic gene arsenal than Dse, but many of these genes are not expressed in planta or are pseudogenized. Overall, comparison of their genomes suggests that these closely related plant pathogens had a common ancestral host but since adapted to different hosts and lifestyles by a combination of differentiated gene content, pseudogenization, and gene regulation.

  • The Genomes of the Fungal Plant Pathogens Cladosporium fulvum and Dothistroma septosporum Reveal Adaptation to Different Hosts and Lifestyles But Also Signatures of Common Ancestry
    eScholarship University of California, 2012
    Co-Authors: Pierre J. G. M. De Wit, Burgt, Ate Van Der, Okmen Bilal, Stergiopoulos Ioannis, Abd-elsalam, Kamel A., Aerts, Andrea L., Bahkali, Ali H., Beenen, Henriek G., Chettri Oranav, Cos, Murray P.
    Abstract:

    We sequenced and compared the genomes of the Dothideomycete fungal plant pathogens Cladosporium fulvum (Cfu) (syn. Passalora fulva) and Dothistroma septosporum (Dse) that are closely related phylogenetically, but have different lifestyles and hosts. Although both fungi grow extracellularly in close contact with host mesophyll cells, Cfu is a biotroph infecting tomato, while Dse is a hemibiotroph infecting pine. The genomes of these fungi have a similar set of genes (70percent of gene content in both genomes are homologs), but differ significantly in size (Cfu >61.1-Mb; Dse 31.2-Mb), which is mainly due to the difference in repeat content (47.2percent in Cfu versus 3.2percent in Dse). Recent adaptation to different lifestyles and hosts is suggested by diverged sets of genes. Cfu contains an tomatinase gene that we predict might be required for detoxification of tomatine, while this gene is absent in Dse. Many genes encoding secreted proteins are unique to each species and the repeat-rich areas in Cfu are enriched for these species-specific genes. In contrast, conserved genes suggest common host ancestry. Homologs of Cfu effector genes, including Ecp2 and Avr4, are present in Dse and induce a Cf-Ecp2- and Cf-4-mediated hypersensitive response, respectively. Strikingly, genes involved in production of the toxin dothistromin, a likely virulence factor for Dse, are conserved in Cfu, but their expression differs markedly with essentially no expression by Cfu in planta. Likewise, Cfu has a carbohydrate-degrading enzyme catalog that is more similar to that of necrotrophs or hemibiotrophs and a larger pectinolytic gene arsenal than Dse, but many of these genes are not expressed in planta or are pseudogenized. Overall, comparison of their genomes suggests that these closely related plant pathogens had a common ancestral host but since adapted to different hosts and lifestyles by a combination of differentiated gene content, pseudogenization, and gene regulation

  • The Cladosporium fulvum virulence protein Avr2 inhibits host proteases required for basal defense
    The Plant cell, 2008
    Co-Authors: H. Peter Van Esse, Pierre J. G. M. De Wit, John W. Van 't Klooster, Melvin D. Bolton, Koste A. Yadeta, Peter Van Baarlen, Sjef Boeren, Jacques Vervoort, Bart P. H. J. Thomma
    Abstract:

    Cladosporium fulvum (syn. Passalora fulva) is a biotrophic fungal pathogen that causes leaf mold of tomato (Solanum lycopersicum). During growth in the apoplast, the fungus establishes disease by secreting effector proteins, 10 of which have been characterized. We have previously shown that the Avr2 effector interacts with the apoplastic tomato Cys protease Rcr3, which is required for Cf-2-mediated immunity. We now show that Avr2 is a genuine virulence factor of C. fulvum. Heterologous expression of Avr2 in Arabidopsis thaliana causes enhanced susceptibility toward extracellular fungal pathogens, including Botrytis cinerea and Verticillium dahliae, and microarray analysis showed that Avr2 expression triggers a global transcriptome reflecting pathogen challenge. Cys protease activity profiling showed that Avr2 inhibits multiple extracellular Arabidopsis Cys proteases. In tomato, Avr2 expression caused enhanced susceptibility toward Avr2-defective C. fulvum strains and also toward B. cinerea and V. dahliae. Cys protease activity profiling in tomato revealed that, in this plant also, Avr2 inhibits multiple extracellular Cys proteases, including Rcr3 and its close relative Pip1. Finally, silencing of Avr2 significantly compromised C. fulvum virulence on tomato. We conclude that Avr2 is a genuine virulence factor of C. fulvum that inhibits several Cys proteases required for plant basal defense.

Bart P. H. J. Thomma - One of the best experts on this subject based on the ideXlab platform.

  • The Cladosporium fulvum virulence protein Avr2 inhibits host proteases required for basal defense
    The Plant cell, 2008
    Co-Authors: H. Peter Van Esse, Pierre J. G. M. De Wit, John W. Van 't Klooster, Melvin D. Bolton, Koste A. Yadeta, Peter Van Baarlen, Sjef Boeren, Jacques Vervoort, Bart P. H. J. Thomma
    Abstract:

    Cladosporium fulvum (syn. Passalora fulva) is a biotrophic fungal pathogen that causes leaf mold of tomato (Solanum lycopersicum). During growth in the apoplast, the fungus establishes disease by secreting effector proteins, 10 of which have been characterized. We have previously shown that the Avr2 effector interacts with the apoplastic tomato Cys protease Rcr3, which is required for Cf-2-mediated immunity. We now show that Avr2 is a genuine virulence factor of C. fulvum. Heterologous expression of Avr2 in Arabidopsis thaliana causes enhanced susceptibility toward extracellular fungal pathogens, including Botrytis cinerea and Verticillium dahliae, and microarray analysis showed that Avr2 expression triggers a global transcriptome reflecting pathogen challenge. Cys protease activity profiling showed that Avr2 inhibits multiple extracellular Arabidopsis Cys proteases. In tomato, Avr2 expression caused enhanced susceptibility toward Avr2-defective C. fulvum strains and also toward B. cinerea and V. dahliae. Cys protease activity profiling in tomato revealed that, in this plant also, Avr2 inhibits multiple extracellular Cys proteases, including Rcr3 and its close relative Pip1. Finally, silencing of Avr2 significantly compromised C. fulvum virulence on tomato. We conclude that Avr2 is a genuine virulence factor of C. fulvum that inhibits several Cys proteases required for plant basal defense.

  • The chitin-binding Cladosporium fulvum effector protein Avr4 is a virulence factor.
    Molecular plant-microbe interactions : MPMI, 2007
    Co-Authors: H. Peter Van Esse, Pierre J. G. M. De Wit, Ioannis Stergiopoulos, Melvin D. Bolton, Bart P. H. J. Thomma
    Abstract:

    The biotrophic fungal pathogen Cladosporium fulvum (syn. Passalora fulva) is the causal agent of tomato leaf mold. The Avr4 protein belongs to a set of effectors that is secreted by C. fulvum during infection and is thought to play a role in pathogen virulence. Previous studies have shown that Avr4 binds to chitin present in fungal cell walls and that, through this binding, Avr4 can protect these cell walls against hydrolysis by plant chitinases. In this study, we demonstrate that Avr4 expression in Arabidopsis results in increased virulence of several fungal pathogens with exposed chitin in their cell walls, whereas the virulence of a bacterium and an oomycete remained unaltered. Heterologous expression of Avr4 in tomato increased the virulence of Fusarium oxysporum f. sp. lycopersici. Through tomato GeneChip analyses, we demonstrate that Avr4 expression in tomato results in the induced expression of only a few genes. Finally, we demonstrate that silencing of the Avr4 gene in C. fulvum decreases its virulence on tomato. This is the first report on the intrinsic function of a fungal avirulence protein that has a counter-defensive activity required for full virulence of the pathogen.

  • Affinity-tags are removed from Cladosporium fulvum effector proteins expressed in the tomato leaf apoplast
    Journal of experimental botany, 2006
    Co-Authors: H. Peter Van Esse, Bart P. H. J. Thomma, John W. Van 't Klooster, Pierre J. G. M. De Wit
    Abstract:

    Cladosporium fulvum (syn. Passalora fulva) is a biotrophic fungal pathogen that causes leaf mould on tomato (Solanum esculentum). The fungus grows exclusively in the tomato leaf apoplast where it secretes several small (

  • Cladosporium fulvum (syn. Passalora fulva), a highly specialized plant pathogen as a model for functional studies on plant pathogenic Mycosphaerellaceae
    Molecular plant pathology, 2005
    Co-Authors: Bart P. H. J. Thomma, H. Peter Van Esse, Pedro W. Crous, Pierre J. G. M. De Wit
    Abstract:

    SUMMARY Taxonomy: Cladosporium fulvum is an asexual fungus for which no sexual stage is currently known. Molecular data, however, support C. fulvum as a member of the Mycosphaerellaceae, clustering with other taxa having Mycosphaerella teleomorphs . C. fulvum has recently been placed in the anamorph genus Passalora as P. fulva . Its taxonomic disposition is supported by its DNA phylogeny, as well as the distinct scars on its conidial hila, which are typical of Passalora , and unlike Cladosporium s.s. , which has teleomorphs that reside in Davidiella , and not Mycosphaerella . Host range and disease symptoms: The presently known sole host of C. fulvum is tomato (members of the genus Lycopersicon ). C. fulvum is mainly a foliar pathogen. Disease symptoms are most obvious on the abaxial side of the leaf and include patches of white mould that turn brown upon sporulation. Due to stomatal clogging, curling of leaves and wilting can occur, leading to defoliation. C. fulvum as a model pathogen: The interaction between C. fulvum and tomato is governed by a gene-for-gene relationship. A total of eight Avr and Ecp genes, and for four of these also the corresponding plant Cf genes, have been cloned. Obtaining conclusive evidence for gene-for-gene relationships is complicated by the poor availability of genetic tools for most Mycosphaerellaceae ‐ plant interactions. Newly developed tools, including Agrobacterium -mediated transformation and RNAi, added to the genome sequence of its host tomato, which will be available within a few years, render C. fulvum attractive as a model species for plant pathogenic Mycosphaerellaceae. Useful websites: http://www.sgn.cornell.edu /help/about / index.html; http://cogeme.ex.ac.uk

  • Affinity-tags are removed from Cladosporium fulvum effector proteins expressed in the tomato leaf apoplast
    2005
    Co-Authors: Peter H. Van Esse, Bart P. H. J. Thomma, John W. Van ’t Klooster
    Abstract:

    Cladosporium fulvum (syn. Passalora fulva) is a bio-trophic fungal pathogen that causes leaf mould on tomato (Solanum esculentum). The fungus grows exclusively in the tomato leaf apoplast where it secretes several small (<15 kDa) cysteine-rich proteins that are thought to play a role in disease establishment. To investigate the role of these proteins, and to identify their in planta targets, a targeted proteomics approach was undertaken. C. fulvum proteins were expressed as recombinant fusion proteins carrying various affinity-tags at either their C- or N-terminus. Although these fusion proteins were correctly expressed and secreted into the leaf apoplast, detection of affinity-tagged C. fulvum proteins failed, and affinity purification did not result in the recovery of these proteins. However, when using C. fulvum effector protein-specific antibodies, specific signals were obtained for the different pro-teins. It is concluded that the stability of the in planta expressed recombinant fusion proteins is insufficient, which results in removal of the affinity-tag from the fusion proteins, irrespective of the C- or N-terminal fusion or the nature of the affinity-tag. Similar phenom-ena were observed when the fusion proteins were expressed in other Solanaceous species, but not when expressed in Arabidopsis thaliana

H. Peter Van Esse - One of the best experts on this subject based on the ideXlab platform.

  • The Cladosporium fulvum virulence protein Avr2 inhibits host proteases required for basal defense
    The Plant cell, 2008
    Co-Authors: H. Peter Van Esse, Pierre J. G. M. De Wit, John W. Van 't Klooster, Melvin D. Bolton, Koste A. Yadeta, Peter Van Baarlen, Sjef Boeren, Jacques Vervoort, Bart P. H. J. Thomma
    Abstract:

    Cladosporium fulvum (syn. Passalora fulva) is a biotrophic fungal pathogen that causes leaf mold of tomato (Solanum lycopersicum). During growth in the apoplast, the fungus establishes disease by secreting effector proteins, 10 of which have been characterized. We have previously shown that the Avr2 effector interacts with the apoplastic tomato Cys protease Rcr3, which is required for Cf-2-mediated immunity. We now show that Avr2 is a genuine virulence factor of C. fulvum. Heterologous expression of Avr2 in Arabidopsis thaliana causes enhanced susceptibility toward extracellular fungal pathogens, including Botrytis cinerea and Verticillium dahliae, and microarray analysis showed that Avr2 expression triggers a global transcriptome reflecting pathogen challenge. Cys protease activity profiling showed that Avr2 inhibits multiple extracellular Arabidopsis Cys proteases. In tomato, Avr2 expression caused enhanced susceptibility toward Avr2-defective C. fulvum strains and also toward B. cinerea and V. dahliae. Cys protease activity profiling in tomato revealed that, in this plant also, Avr2 inhibits multiple extracellular Cys proteases, including Rcr3 and its close relative Pip1. Finally, silencing of Avr2 significantly compromised C. fulvum virulence on tomato. We conclude that Avr2 is a genuine virulence factor of C. fulvum that inhibits several Cys proteases required for plant basal defense.

  • Cladosporium fulvum effector proteins and their role in pathogen virulence
    2008
    Co-Authors: H. Peter Van Esse
    Abstract:

    Cladosporium fulvum (syn. Passalora fulva) is a biotrophic fungal pathogen that causes leaf mould of tomato (Solanum esculentum). Chapter 1 is a “pathogen profile” describing the biology of the pathogen. During growth in the leaf apoplast, the intercellular space surrounding the mesophyll cells, the fungus secretes effector proteins that are thought to play a role in disease establishment. Eight of these effectors have been characterized in detail. For most of these effectors, cognate C. fulvum (Cf) resistance loci have been identified in tomato that mediate an immune response upon recognition of (the activity of) the cognate effector. In chapter 2, a targeted proteomics approach to investigate the role of these effector proteins and to identify possible in planta targets is described. C. fulvum proteins were expressed as recombinant fusion proteins carrying various affinity–tags at either their C– or N–terminus. Although these fusion proteins were correctly expressed and secreted into the leaf apoplast, detection of affinity–tagged C. fulvum proteins failed and affinity–purification did not result in the recovery of these proteins. However, when using C. fulvum effector protein–specific antibodies, specific signals were obtained for the different proteins. It was therefore concluded that the stability of the in planta expressed recombinant fusion proteins is insufficient, which resulted in removal of the affinity–tag from the fusion proteins, irrespective of C– or N–terminal fusion or the nature of the affinity–tag. Similar observations were made when the fusion proteins were expressed in other Solanaceous species, but not when expressed in Arabidopsis thaliana. Previous studies have demonstrated that Avr4 binds to chitin present in fungal cell walls, and that this binding by Avr4 can protect these cell walls against hydrolysis by plant chitinases. In chapter 3 it is described that Avr4–expression in Arabidopsis results in increased virulence of several fungal pathogens with exposed chitin in their cell walls, whereas the virulence of a bacterium and an oomycete remained unaltered. Heterologous expression of Avr4 in tomato increased the virulence of Fusarium oxysporum f. sp. lycopersici. Tomato GeneChip analysis was used to demonstrate that Avr4–expression in tomato results in the induced expression of only a handful of genes. Finally, silencing of the Avr4 gene in C. fulvum decreased fungal virulence on tomato. In conclusion, chapter 3 is the first report on the intrinsic function of a fungal avirulence protein that displays self-defense activity which is required for full pathogen virulence. In chapter 4, a study on the intrinsic biological function of Avr2 is presented. The Avr2 effector interacts with the apoplastic tomato cysteine protease Rcr3, which is required for Cf–2–mediated immunity. In this chapter it is demonstrated that Avr2 is a genuine virulence factor of C. fulvum. Heterologous expression of Avr2 in Arabidopsis resulted in enhanced susceptibility towards a number of extracellular fungal pathogens that include Botrytis cinerea and Verticillium dahliae, and microarray analysis of unchallenged Arabidopsis plants showed that Avr2 expression triggered a global transcription profile that is reminiscent of pathogen challenge. Cysteine protease activity profiling revealed that Avr2 inhibits multiple extracellular Arabidopsis cysteine proteases. In tomato, Avr2 expression resulted in enhanced susceptibility not only towards natural Avr2–defective C. fulvum strains, but also towards Botrytis cinerea and Verticillium dahliae. Cysteine protease activity profiling in tomato revealed that Avr2 inhibits multiple extracellular cysteine proteases including Rcr3 and its close relative PIP1. Finally, silencing of the Avr2 gene in C. fulvum significantly compromised fungal virulence on tomato. This all shows that Avr2 is a genuine virulence factor of C. fulvum that inhibits several cysteine proteases required for plant basal defense in tomato. Chapter 5 describes the discovery and characterization of a novel effector protein of C. fulvum, Ecp6. To discover novel C. fulvum effectors that might play a role in virulence, two–dimensional polyacrylamide gel electrophoresis (2D–PAGE) was used to visualize proteins secreted during C. fulvum–tomato interactions. Three novel C. fulvum proteins were identified; CfPhiA, Ecp6, and Ecp7. CfPhiA shows homology to proteins found on fungal sporogenous cells called phialides, while Ecp6 contains lysine motifs (LysM domains), which are recognized as carbohydrate–binding modules. Finally, Ecp7 encodes a small, cysteine–rich protein with no homology to known proteins. Heterologous expression of Ecp6 significantly increased the virulence of the vascular pathogen Fusarium oxysporum on tomato. Furthermore, by RNAi–mediated gene silencing it was demonstrated that Ecp6 is instrumental for C. fulvum virulence on tomato. Hardly any allelic variation was observed in the Ecp6 coding region of a worldwide collection of C. fulvum strains. Although none of the C. fulvum effectors identified so far have obvious orthologs in other organisms, conserved Ecp6 orthologs were identified in various fungal species. Homology based modelling suggests that the LysM domains of C. fulvum Ecp6 may be involved in chitin binding. Chapter 6 presents global transcriptional profiling study to compare transcriptional changes in tomato during compatible and incompatible interactions with the foliar pathogenic fungus Cladosporium fulvum and the soil–borne vascular pathogenic fungus Verticillium dahliae. Although both pathogens colonize different host tissues, they display significant commonalities in their infection strategies as they both penetrate natural openings and grow strictly extracellular without the formation of haustoria. Furthermore, in incompatible interactions with both pathogens resistance is conveyed by extracellular transmembrane receptors that belong to the class of receptor–like proteins. For each of the two pathogens, the transcriptomes of the compatible and incompatible interaction largely overlapped. However, the C. fulvum–induced transcriptomes showed little overlap with the V. dahliae–induced transcriptomes, as most genes were uniquely regulated by one of the two pathogens. This also applied to both incompatible interactions, despite defense activation by the same type of resistance protein. Remarkably, of the relatively small subset of genes that was regulated by both pathogens a large portion showed an inverse regulation; induced by one pathogen and repressed by the other. With pathway reconstruction, interacting networks of tomato genes implicated in photorespiration, hypoxia and glycoxylate metabolism were identified that were repressed upon infection with C. fulvum and induced by V. dahliae. Similarly, auxin signaling was differentially affected by the two pathogens. In chapter 7, the general discussion, the implications are of the data that are presented in this thesis are discussed for the use of C. fulvum as a model, and for fungal pathogens in general. Furthermore, the use of heterologous expression systems to study fungal effectors is briefly discussed. In several of the chapters presented in this thesis, the use of microarrays has been instrumental to investigate the biology of C. fulvum and the role of specific effectors secreted by the pathogen. Therefore, an overview of the currently available in silico tools for reconstruction of cellular pathways based on plant gene expression datasets is presented.

  • The chitin-binding Cladosporium fulvum effector protein Avr4 is a virulence factor.
    Molecular plant-microbe interactions : MPMI, 2007
    Co-Authors: H. Peter Van Esse, Pierre J. G. M. De Wit, Ioannis Stergiopoulos, Melvin D. Bolton, Bart P. H. J. Thomma
    Abstract:

    The biotrophic fungal pathogen Cladosporium fulvum (syn. Passalora fulva) is the causal agent of tomato leaf mold. The Avr4 protein belongs to a set of effectors that is secreted by C. fulvum during infection and is thought to play a role in pathogen virulence. Previous studies have shown that Avr4 binds to chitin present in fungal cell walls and that, through this binding, Avr4 can protect these cell walls against hydrolysis by plant chitinases. In this study, we demonstrate that Avr4 expression in Arabidopsis results in increased virulence of several fungal pathogens with exposed chitin in their cell walls, whereas the virulence of a bacterium and an oomycete remained unaltered. Heterologous expression of Avr4 in tomato increased the virulence of Fusarium oxysporum f. sp. lycopersici. Through tomato GeneChip analyses, we demonstrate that Avr4 expression in tomato results in the induced expression of only a few genes. Finally, we demonstrate that silencing of the Avr4 gene in C. fulvum decreases its virulence on tomato. This is the first report on the intrinsic function of a fungal avirulence protein that has a counter-defensive activity required for full virulence of the pathogen.

  • Affinity-tags are removed from Cladosporium fulvum effector proteins expressed in the tomato leaf apoplast
    Journal of experimental botany, 2006
    Co-Authors: H. Peter Van Esse, Bart P. H. J. Thomma, John W. Van 't Klooster, Pierre J. G. M. De Wit
    Abstract:

    Cladosporium fulvum (syn. Passalora fulva) is a biotrophic fungal pathogen that causes leaf mould on tomato (Solanum esculentum). The fungus grows exclusively in the tomato leaf apoplast where it secretes several small (

  • Cladosporium fulvum (syn. Passalora fulva), a highly specialized plant pathogen as a model for functional studies on plant pathogenic Mycosphaerellaceae
    Molecular plant pathology, 2005
    Co-Authors: Bart P. H. J. Thomma, H. Peter Van Esse, Pedro W. Crous, Pierre J. G. M. De Wit
    Abstract:

    SUMMARY Taxonomy: Cladosporium fulvum is an asexual fungus for which no sexual stage is currently known. Molecular data, however, support C. fulvum as a member of the Mycosphaerellaceae, clustering with other taxa having Mycosphaerella teleomorphs . C. fulvum has recently been placed in the anamorph genus Passalora as P. fulva . Its taxonomic disposition is supported by its DNA phylogeny, as well as the distinct scars on its conidial hila, which are typical of Passalora , and unlike Cladosporium s.s. , which has teleomorphs that reside in Davidiella , and not Mycosphaerella . Host range and disease symptoms: The presently known sole host of C. fulvum is tomato (members of the genus Lycopersicon ). C. fulvum is mainly a foliar pathogen. Disease symptoms are most obvious on the abaxial side of the leaf and include patches of white mould that turn brown upon sporulation. Due to stomatal clogging, curling of leaves and wilting can occur, leading to defoliation. C. fulvum as a model pathogen: The interaction between C. fulvum and tomato is governed by a gene-for-gene relationship. A total of eight Avr and Ecp genes, and for four of these also the corresponding plant Cf genes, have been cloned. Obtaining conclusive evidence for gene-for-gene relationships is complicated by the poor availability of genetic tools for most Mycosphaerellaceae ‐ plant interactions. Newly developed tools, including Agrobacterium -mediated transformation and RNAi, added to the genome sequence of its host tomato, which will be available within a few years, render C. fulvum attractive as a model species for plant pathogenic Mycosphaerellaceae. Useful websites: http://www.sgn.cornell.edu /help/about / index.html; http://cogeme.ex.ac.uk

Pedro Alberto Balatti - One of the best experts on this subject based on the ideXlab platform.

  • A survey on occurrence of Cladosporium fulvum identifies race 0 and race 2 in tomato-growing areas of Argentina
    Plant disease, 2015
    Co-Authors: Roció Medina, Pierre J. G. M. De Wit, Silvina Marianela Yanil López, Mario Emilio Ernesto Franco, Cristina Rollan, Blanca Lía Ronco, Mario Carlos Nazareno Saparrat, Pedro Alberto Balatti
    Abstract:

    The presence of Cladosporium fulvum (syn. Passalora fulva), causal agent of tomato leaf mold, was confirmed in the two main greenhouse-production areas for tomato in Argentina. Using both morphological characters and internal transcribed spacer sequencing, we confirmed the presence of physiological races of this pathogen. A diagnostic multiplex polymerase chain reaction (PCR) was also developed, using primers derived from C. fulvum avirulence (Avr) genes. In all, 20 isolates of Cladosporium spp. were obtained as monospore cultures and 12 were identified as C. fulvum. By this method, we showed that, of these 12 isolates, 5 were race 0 (carrying functional Avr2, Avr4, Avr4E, and Avr9 genes) and 7 were race 2 (lacking the Avr2 gene). Race identity was confirmed by testing their virulence on a set of tomato differentials carrying different Cf resistance genes. All Avr genes could be amplified in single or multiplex PCR using DNA isolated from in vitro grown monospore cultures but only three Avr could be amplified when genomic DNA was isolated from C. fulvum-infected necrotic leaf tissue.

  • Identification of Races 0 and 2 of Cladosporium fulvum (syn Passalora fulva) on Tomato in the Cinturón Hortícola de La Plata, Argentina
    Plant disease, 2013
    Co-Authors: C. Rollan, Roció Medina, Silvina Marianela Yanil López, Mario Carlos Nazareno Saparrat, V. Protto, J. Vera Bahima, L. Ronco, Pedro Alberto Balatti
    Abstract:

    Surveys aimed at evaluating the incidence and severity of a new disease that developed in greenhouses cultivated with tomato (Solanum lycopersicum L.) were performed during 2009 and 2010 in greenhouses of the cultivars Elpida (Enza Zaden) and Colibri (Clause) in an area of tomato production known as the Cinturon Horticola de La Plata (the “horticultural belt of La Plata”). The disease had a 100% prevalence and 90% incidence within the ten 250 m2 greenhouses that were monitored in 2009, 2010, and 2011. In two consecutive assays, severity was 40%. The wide distribution of the disease suggests that the tomato hybrids under use lack resistance genes. The upper surface of diseased leaves had pale green to yellowish, 1- to 1.5-cm spots with undefined margins that progressed to a yellowish brown color, while on the lower side they had pale brown to brown sporulation of fungal conidiophores and conidia. Monosporic fungal cultures were obtained by needle transfer of conidia from sporulating areas of leaves (n = 20...

John W. Van 't Klooster - One of the best experts on this subject based on the ideXlab platform.

  • The Cladosporium fulvum virulence protein Avr2 inhibits host proteases required for basal defense
    The Plant cell, 2008
    Co-Authors: H. Peter Van Esse, Pierre J. G. M. De Wit, John W. Van 't Klooster, Melvin D. Bolton, Koste A. Yadeta, Peter Van Baarlen, Sjef Boeren, Jacques Vervoort, Bart P. H. J. Thomma
    Abstract:

    Cladosporium fulvum (syn. Passalora fulva) is a biotrophic fungal pathogen that causes leaf mold of tomato (Solanum lycopersicum). During growth in the apoplast, the fungus establishes disease by secreting effector proteins, 10 of which have been characterized. We have previously shown that the Avr2 effector interacts with the apoplastic tomato Cys protease Rcr3, which is required for Cf-2-mediated immunity. We now show that Avr2 is a genuine virulence factor of C. fulvum. Heterologous expression of Avr2 in Arabidopsis thaliana causes enhanced susceptibility toward extracellular fungal pathogens, including Botrytis cinerea and Verticillium dahliae, and microarray analysis showed that Avr2 expression triggers a global transcriptome reflecting pathogen challenge. Cys protease activity profiling showed that Avr2 inhibits multiple extracellular Arabidopsis Cys proteases. In tomato, Avr2 expression caused enhanced susceptibility toward Avr2-defective C. fulvum strains and also toward B. cinerea and V. dahliae. Cys protease activity profiling in tomato revealed that, in this plant also, Avr2 inhibits multiple extracellular Cys proteases, including Rcr3 and its close relative Pip1. Finally, silencing of Avr2 significantly compromised C. fulvum virulence on tomato. We conclude that Avr2 is a genuine virulence factor of C. fulvum that inhibits several Cys proteases required for plant basal defense.

  • Affinity-tags are removed from Cladosporium fulvum effector proteins expressed in the tomato leaf apoplast
    Journal of experimental botany, 2006
    Co-Authors: H. Peter Van Esse, Bart P. H. J. Thomma, John W. Van 't Klooster, Pierre J. G. M. De Wit
    Abstract:

    Cladosporium fulvum (syn. Passalora fulva) is a biotrophic fungal pathogen that causes leaf mould on tomato (Solanum esculentum). The fungus grows exclusively in the tomato leaf apoplast where it secretes several small (