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A new partititvirus isolated from a Trichoderma harzianum strain (T673), collected in China, was characterized and annotated as Trichoderma harzianum partitivirus 2 (ThPV2). The genome of ThPV2 consists of a 1693 bp dsRNA1 encoding a... more
A new partititvirus isolated from a Trichoderma harzianum strain (T673), collected in China, was characterized and annotated as Trichoderma harzianum partitivirus 2 (ThPV2). The genome of ThPV2 consists of a 1693 bp dsRNA1 encoding a putative RNA-dependent RNA polymerase (RdRp) and a 1458 bp dsRNA2 encoding a hypothetical protein. In comparative studies employing the ThPV2-infected strain (T673) and a strain cured by ribavirin treatment (virus-free strain T673-F), we investigated biological effects of ThPV2 infection. While the growth rate of the virus-infected fungus differed little from that of the cured variant, higher mycelial density, conidiospore, and chlamydospore production were observed in the virus-infected strain T673. Furthermore, both the ThPV2-infected and the cured strain showed growth- and development-promoting activities in cucumber plants. In vitro confrontation tests showed that strains T673 and T673-F inhibited several important fungal pathogens and an oomycete p...
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Efficient plant protection is of fundamental importance in order to warrant food security. Here, we present arguments that a coordinate approach in plant protection is urgently required, taking advantage of a broad mix of measures,... more
Efficient plant protection is of fundamental importance in order to warrant food security. Here, we present arguments that a coordinate approach in plant protection is urgently required, taking advantage of a broad mix of measures, including modern synthetic chemistries, capable of protecting plants from adverse organisms.
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Antibiotic resistance is an increasing threat to human health. In the case of Aspergillus fumigatus , which is an environmental fungus that also causes life-threatening infections in humans, antimicrobial resistance is suggested to arise... more
Antibiotic resistance is an increasing threat to human health. In the case of Aspergillus fumigatus , which is an environmental fungus that also causes life-threatening infections in humans, antimicrobial resistance is suggested to arise from fungicide use in agriculture, as the chemicals used for plant protection are almost identical to the antifungals used clinically. However, removing azole fungicides from crop fields threatens the global food supply via a reduction in yield. In this study, we survey crop fields before and after fungicide application. We find a low overall azole resistance rate among agricultural isolates, as well as a lack of genomic and population impact following fungicide application, leading us to conclude azole use on crops does not significantly contribute to resistance in A. fumigatus .
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A growing world population requires an increase in the quality and quantity of food production. However, field losses due to biotic stresses are currently estimated to be between 10 and 20% worldwide. The risk of resistance and strict... more
A growing world population requires an increase in the quality and quantity of food production. However, field losses due to biotic stresses are currently estimated to be between 10 and 20% worldwide. The risk of resistance and strict pesticide legislation necessitate innovative agronomical practices to adequately protect crops in the future, such as the identification of new substances with novel modes of action. In the present study, liquid chromatography mass spectrometry was used to characterize Rheum rhabarbarum root extracts that were primarily composed of the stilbenes rhaponticin, desoxyrhaponticin, and resveratrol. Minor components were the flavonoids catechin, epicatechin gallate, and procyanidin B1. Specific polyphenolic mixtures inhibited mycelial growth of several phytopathogenic fungi and oomycetes. Foliar spray applications with fractions containing stilbenes and flavonoids inhibited spore germination of powdery mildew in Hordeum vulgare with indications of synergisti...
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Nitrate reductase (EC 1.6.6.2) has been isolated from 11 Sphagnum (Bryophyta) species and purified up to 500-fold by a procedure involving (NH4)2SO4 precipitation and blue sepharose affinity chromatography. The purest fraction showed a... more
Nitrate reductase (EC 1.6.6.2) has been isolated from 11 Sphagnum (Bryophyta) species and purified up to 500-fold by a procedure involving (NH4)2SO4 precipitation and blue sepharose affinity chromatography. The purest fraction showed a nitrate reductase activity of 7.7 nkat mg protein-1 with NADPH as the electron donor. Beside NADPH and NADH, FMNH2 and - after blue sepharose affinity purification of the enzyme - MVH2 were used as electron donors in different assays.
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Expression of the Aspergillus nidulans brlA gene induces a developmental pathway leading to the production of asexual spores. We have introduced mutations into brlA that are expected to disrupt either or both Cys2-His2 Zn(II) coordination... more
Expression of the Aspergillus nidulans brlA gene induces a developmental pathway leading to the production of asexual spores. We have introduced mutations into brlA that are expected to disrupt either or both Cys2-His2 Zn(II) coordination sites postulated to exist in the BrlA polypeptide. The resultant brlA alleles fail to induce either the asexual reproductive pathway or the expression of development-specific genes. These data support the hypothesis that brlA encodes a nucleic acid-binding protein whose activity requires each of two zinc fingers.
Research Interests: Genetics, Biology, Transcription Factors, Medicine, Biological Sciences, and 15 moreMolecular and cellular biology, Mutation, Animals, Zinc, Gene, Metalloproteins, Aspergillus Nidulans, Zinc Finger, Xenopus laevis, Amino Acid Sequence, Base Sequence, DNA binding proteins, Molecular Sequence Data, alleles, and Medical and Health Sciences
Many plant pathogenic fungi, such as the cereal pathogen Colletotrichum graminicola , differentiate highly specialized infection structures called appressoria, which send a penetration peg into the underlying plant cell. Appressoria have... more
Many plant pathogenic fungi, such as the cereal pathogen Colletotrichum graminicola , differentiate highly specialized infection structures called appressoria, which send a penetration peg into the underlying plant cell. Appressoria have been shown to generate enormous turgor pressure, but direct evidence for mechanical infection of plants by fungi is lacking. A microscopic method was developed that uses elastic optical waveguides to visualize and measure forces locally exerted by single appressoria. By this method, the force exerted by appressoria of C. graminicola was found to be about 17 micronewtons.
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Research Interests: Botany, Microbiology, Plant Biology, Biology, Medicine, and 15 morePeroxidase, Vigna, Haustorium, Vicia faba, Substrate Specificity, Western blot, Lignification, Enzyme activity, Cell Wall, Polyacrylamide Gel Electrophoresis, Extracellular, Planta, Phenylalanine ammonia lyase, Rust Fungi, and Host resistance
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When inoculated onto maize leaves at the onset of senescence, the hemibiotroph Colletotrichum graminicola causes green islands that are surrounded by senescing tissue. Taking advantage of green islands as indicators of sites of the... more
When inoculated onto maize leaves at the onset of senescence, the hemibiotroph Colletotrichum graminicola causes green islands that are surrounded by senescing tissue. Taking advantage of green islands as indicators of sites of the establishment of successful infection and of advanced high-performance liquid chromatography tandem mass spectrometry methodology, we analyzed changes in the patterns and levels of cytokinins (CK) at high spatial and analytical resolution. Twenty individual CK were detected in green islands. Levels of cis-zeatin-9-riboside and cis-zeatin-9-riboside-5′-monophosphate increased drastically, whereas that of the most prominent CK, cis-zeatin-O-glucoside, decreased. The fungus likely performed these conversions because corresponding activities were also detected in in vitro cultures amended with CK. We found no evidence that C. graminicola is able to synthesize CK entirely de novo in minimal medium but, after adding dimethylallyl diphosphate, a precursor of CK ...
Research Interests: Genetics, Microbiology, Photosynthesis, Plant Biology, Biology, and 15 moreMedicine, Plant diseases, Colletotrichum, Chlorophyll, Tandem Mass Spectrometry, Organophosphorus Compounds, Molecular Plant Microbe Interactions, High Pressure Liquid Chromatography, Zea mays, Host Pathogen Interactions, Culture Media, Cytokinins, Plant Leaves, Cytokinin, and zeatin
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SummaryThe hemibiotrophic maize pathogen Colletotrichum graminicola synthesizes one intracellular and three secreted siderophores. eGFP fusions with the key siderophore biosynthesis gene, SID1, encoding l‐ornithine‐N5‐monooxygenase,... more
SummaryThe hemibiotrophic maize pathogen Colletotrichum graminicola synthesizes one intracellular and three secreted siderophores. eGFP fusions with the key siderophore biosynthesis gene, SID1, encoding l‐ornithine‐N5‐monooxygenase, suggested that siderophore biosynthesis is rigorously downregulated specifically during biotrophic development. In order to investigate the role of siderophores during vegetative development and pathogenesis, SID1, which is required for synthesis of all siderophores, and the non‐ribosomal peptide synthetase gene NPS6, synthesizing secreted siderophores, were deleted. Mutant analyses revealed that siderophores are required for vegetative growth under iron‐limiting conditions, conidiation, ROS tolerance, and cell wall integrity. Δsid1 and Δnps6 mutants were hampered in formation of melanized appressoria and impaired in virulence. In agreement with biotrophy‐specific downregulation of siderophore biosynthesis, Δsid1 and Δnps6 strains were not affected in bi...
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Research Interests: Microbiology, Biology, Regulation, Multidisciplinary, Enzyme, and 5 moreHaustorium, Vicia faba, Pectin, Cell Wall, and Thallophyta
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Polyketide synthases (PKSs) and/or nonribosomal peptide synthetases (NRPSs) are central components of secondary metabolism in bacteria, plants, and fungi. In filamentous fungi, diverse PKSs and NRPSs participate in the biosynthesis of... more
Polyketide synthases (PKSs) and/or nonribosomal peptide synthetases (NRPSs) are central components of secondary metabolism in bacteria, plants, and fungi. In filamentous fungi, diverse PKSs and NRPSs participate in the biosynthesis of secondary metabolites such as pigments, antibiotics, siderophores, and mycotoxins. However, many secondary metabolites as well as the enzymes involved in their production are yet to be discovered. Both PKSs and NRPSs require activation by enzyme members of the 4′-phosphopantetheinyl transferase (PPTase) family. Here, we report the isolation and characterization of Aspergillus nidulans strains carrying conditional ( cfwA2 ) and null (Δ cfwA ) mutant alleles of the cfwA gene, encoding an essential PPTase. We identify the polyketides shamixanthone, emericellin, and dehydroaustinol as well as the sterols ergosterol, peroxiergosterol, and cerevisterol in extracts from A. nidulans large-scale cultures. The PPTase CfwA/NpgA was required for the production of ...
Research Interests: Biochemistry, Pigmentation, Biology, Medicine, Biological Sciences, and 15 morePhylogeny, Mutation, Phylogenetic analysis, Enzyme, Fermentation, Aspergillus Nidulans, Lysine, Filamentous fungi, Fatty Acid, Large Scale, Biological Factors, Ergosterol, Eukaryotic Cell, alleles, and Medical and Health Sciences
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The genus Colletotrichum harbors many plant pathogenic species, several of which cause significant yield losses in the field and post harvest. Typically, in order to infect their host plants, spores germinate, differentiate a pressurized... more
The genus Colletotrichum harbors many plant pathogenic species, several of which cause significant yield losses in the field and post harvest. Typically, in order to infect their host plants, spores germinate, differentiate a pressurized infection cell, and display a hemibiotrophic lifestyle after plant invasion. Several factors required for virulence or pathogenicity have been identified in different Colletotrichum species, and adaptation of cell wall biogenesis to distinct stages of pathogenesis has been identified as a major pre-requisite for the establishment of a compatible parasitic fungus–plant interaction. Here, we highlight aspects of fungal cell wall biogenesis during plant infection, with emphasis on the maize leaf anthracnose and stalk rot fungus, Colletotrichum graminicola.
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Fungal pathogenicity is governed by environmental factors, with nitrogen playing a key role in triggering pathogenic development. Spores germinating on the plant cuticle are exposed to a nitrogen-free environment, and reprograming of... more
Fungal pathogenicity is governed by environmental factors, with nitrogen playing a key role in triggering pathogenic development. Spores germinating on the plant cuticle are exposed to a nitrogen-free environment, and reprograming of nitrogen metabolism is required for bridging the time needed to gain access to the nitrogen sources of the host. Although degradation of endogenous purine bases efficiently generates ammonium and may allow the fungus to bridge the preinvasion nitrogen gap, the roles of the purine degradation pathway and of the key genes encoding allantoicase and urease are largely unknown in plant pathogenic fungi. To investigate the roles of the allantoicase and urease genes ALA1 and URE1 of the maize anthracnose fungus Colletotrichum graminicola in pathogenic development, we generated ALA1:eGFP and URE1:eGFP fusion strains as well as allantoicase- and urease-deficient mutants. Virulence assays, live cell, and differential interference contrast imaging, chemical comple...
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Plants producing antisense or double-stranded RNA molecules that target specific genes of eukaryotic pests or pathogens can become protected from their attack. This beneficial effect was also reported for plant-fungus interactions and is... more
Plants producing antisense or double-stranded RNA molecules that target specific genes of eukaryotic pests or pathogens can become protected from their attack. This beneficial effect was also reported for plant-fungus interactions and is believed to reflect uptake of the RNAs by the fungus via an as yet unknown mechanism, followed by target gene silencing. Here we report that wheat plants pre-infected with Barley stripe mosaic virus (BSMV) strains containing antisense sequences against target genes of the Fusarium head blight (FHB) fungus F. culmorum caused a reduction of corresponding transcript levels in the pathogen and reduced disease symptoms. Stable transgenic wheat plants carrying an RNAi hairpin construct against the β-1, 3-glucan synthase gene FcGls1 of F. culmorum or a triple combination of FcGls1 with two additional, pre-tested target genes also showed enhanced FHB resistance in leaf and spike inoculation assays under greenhouse and near-field conditions, respectively. Mi...
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Asian soybean rust (Phakopsora pachyrhizi) causes a devastating disease in soybean (Glycine max). We tested the hypothesis that the fungus generates high turgor pressure in its hyaline appressoria to mechanically pierce epidermal cells.... more
Asian soybean rust (Phakopsora pachyrhizi) causes a devastating disease in soybean (Glycine max). We tested the hypothesis that the fungus generates high turgor pressure in its hyaline appressoria to mechanically pierce epidermal cells. Turgor pressure was determined by a microscopic technique, called transmitted light double-beam interference Mach-Zehnder microscopy (MZM), which was developed in the 1960s as a forefront of live cell imaging. We revitalized some original microscopes and equipped them for modern image capturing. MZM data were corroborated by cytorrhysis experiments. Incipient cytorrhysis determined the turgor pressure in appressoria of P. pachyrhizi to be equivalent to 5.13 MPa. MZM data revealed that osmotically active sugar alcohols only accounted for 75% of this value. Despite having a lower turgor pressure, hyaline rust appressoria were able to penetrate non-biodegradable polytetrafluoroethylene (PTFE) membranes more efficiently than do melanized appressoria of t...
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β-1,3-Glucan and chitin are the most prominent polysaccharides of the fungal cell wall. Covalently linked, these polymers form a scaffold that determines the form and properties of vegetative and pathogenic hyphae. While the role of... more
β-1,3-Glucan and chitin are the most prominent polysaccharides of the fungal cell wall. Covalently linked, these polymers form a scaffold that determines the form and properties of vegetative and pathogenic hyphae. While the role of chitin in plant infection is well understood, the role of β-1,3-glucan is unknown. We functionally characterized the β-1,3-glucan synthase gene GLS1 of the maize (Zea mays) pathogen Colletotrichum graminicola, employing RNA interference (RNAi), GLS1 overexpression, live-cell imaging, and aniline blue fluorochrome staining. This hemibiotroph sequentially differentiates a melanized appressorium on the cuticle and biotrophic and necrotrophic hyphae in its host. Massive β-1,3-glucan contents were detected in cell walls of appressoria and necrotrophic hyphae. Unexpectedly, GLS1 expression and β-1,3-glucan contents were drastically reduced during biotrophic development. In appressoria of RNAi strains, downregulation of β-1,3-glucan synthesis increased cell wal...
Research Interests: Genetics, Microbiology, Plant Biology, Fluorescence Microscopy, Biology, and 15 moreMembrane Proteins, Medicine, RNA interference, Phylogeny, Plant diseases, Colletotrichum, Mutation, Disease resistance, Plant cell, Host Pathogen Interactions, Cell Wall, Biochemistry and cell biology, Gene expression profiling, reverse transcriptase polymerase chain reaction, and Hyphae
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The endophyte Piriformospora indica colonizes roots of a range of host plants and increases biomass production and resistance to fungal pathogens and, thus has been considered a biocontrol fungus. However, the field performance of this... more
The endophyte Piriformospora indica colonizes roots of a range of host plants and increases biomass production and resistance to fungal pathogens and, thus has been considered a biocontrol fungus. However, the field performance of this fungus has not yet been tested in temperate climates. Therefore, we evaluated the performance of this fungus in different substrata under greenhouse and practical field conditions. Roots of winter wheat were colonized efficiently, and biomass was particularly increased on poor substrata. In greenhouse experiments, symptom severity of a typical leaf (Blumeria graminis f. sp. tritici), stem base (Pseudocercosporella herpotrichoides), and root (Fusarium culmorum) pathogen was reduced significantly. However, in field experiments, symptoms caused by the leaf pathogen did not differ in Piriformospora indica-colonized compared with control plants. In the field, Pseudocercosporella herpotrichoides disease severity was significantly reduced in plants colonized...
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Research Interests: Biochemistry, Microbiology, Biology, Medicine, Biological Sciences, and 12 moreSaccharomyces cerevisiae, Phylogeny, Plant Physiology, Colletotrichum, Sucrose, Membrane transport proteins, Protein Sequence Analysis, Substrate Specificity, Hydrogen-Ion Concentration, Molecular Sequence Data, Cell Membrane, and Maltose
Acquisition of resistance to sterol demethylation inhibitors (DMIs) by populations of Venturia inaequalis was investigated using a microscopical method developed by C. Siebels and K. Mendgen. Microscopical analysis of conidiophore... more
Acquisition of resistance to sterol demethylation inhibitors (DMIs) by populations of Venturia inaequalis was investigated using a microscopical method developed by C. Siebels and K. Mendgen. Microscopical analysis of conidiophore formation enabled the earlier detection of resistance and a clearer distinction between DMI-resistant and DMI-sensitive populations than other in vivo methods commonly used to analyze inhibitory effects of fungicides. In addition, because observations were made on the level of individuals, quantitative measures of the composition of conidial populations were obtained. The development of DMI sensitivity was followed over a period of 3 years in control apple orchards that had never been treated with fungicides and in orchards with DMI history. The 50% effective dose values determined by microscopical evaluation of conidio-phore development for untreated populations revealed the baseline sensitivities of 0.3, 0.96, 0.09, 1.22, and 1.92 mg/liter for flusilazol...
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The hemibiotroph Colletotrichum graminicola is the causal agent of stem rot and leaf anthracnose on Zea mays. Following penetration of epidermal cells, the fungus enters a short biotrophic phase, followed by a destructive necrotrophic... more
The hemibiotroph Colletotrichum graminicola is the causal agent of stem rot and leaf anthracnose on Zea mays. Following penetration of epidermal cells, the fungus enters a short biotrophic phase, followed by a destructive necrotrophic phase of pathogenesis. During both phases, secreted fungal proteins are supposed to determine progress and success of the infection. To identify genes encoding such proteins, we constructed a yeast signal sequence trap (YSST) cDNA-library from RNA extracted from mycelium grown in vitro on corn cell walls and leaf extract. Of the 103 identified unigenes, 50 showed significant similarities to genes with a reported function, 25 sequences were similar to genes without a known function, and 28 sequences showed no similarity to entries in the databases. Macroarray hybridization and quantitative reverse-transcriptase polymerase chain reaction confirmed that most genes identified by the YSST screen are expressed in planta. Other than some genes that were const...
Research Interests: Genetics, Microbiology, Plant Biology, Biology, Plant Molecular Biology, and 15 moreMedicine, Quantitative analysis, In Vitro, Identification, Colletotrichum, Gene, Molecular Plant Microbe Interactions, Gen, Gramineae, Quantitative Analysis, Cell Wall, Database, cDNA library, Nucleotide sequence, and reverse transcriptase polymerase chain reaction
The ascomycete and causative agent of maize anthracnose and stem rot, Colletotrichum graminicola, differentiates melanized infection cells called appressoria that are indispensable for breaching the plant cell wall. High concentrations of... more
The ascomycete and causative agent of maize anthracnose and stem rot, Colletotrichum graminicola, differentiates melanized infection cells called appressoria that are indispensable for breaching the plant cell wall. High concentrations of osmolytes accumulate within the appressorium, and the internal turgor pressure of up to 5.4 MPa provides sufficient force to penetrate the leaf epidermis directly. In order to assess the function of melanin in C. graminicola appressoria, we identified and characterized the polyketide synthase 1 (CgPKS1) gene which displayed high similarity to fungal polyketide synthases (PKS) involved in synthesis of 1,3,6,8-tetrahydronaphthalene, the first intermediate in melanin biosynthesis. Cgpks1 albino mutants created by targeted gene disruption were unable to penetrate intact leaves and ruptured frequently but, surprisingly, were able to penetrate ultrathin polytetrafluoroethylene membranes mimicking the plant surface. Nonmelanized Cgpks1 appressoria were se...
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Typically, pathogenesis of the hemibiotroph Colletotrichum graminicola and defense responses of its host, Zea mays, are studied on young leaves. Equivalent studies have not been performed with leaves undergoing senescence, a situation... more
Typically, pathogenesis of the hemibiotroph Colletotrichum graminicola and defense responses of its host, Zea mays, are studied on young leaves. Equivalent studies have not been performed with leaves undergoing senescence, a situation that is relevant in the field. We discovered that, in contrast to anthracnose symptoms formed on young and mature leaves, green islands reminiscent of those known from obligate biotrophs were formed on senescing leaves. Microscopy revealed that the fungus grew in both symptoms from the epidermis towards the bundle sheath. In green islands, tissues remained intact for an extended time period. Imaging PAM (pulse-amplitude-modulation) fluorescence analyses revealed that photosynthesis is transiently maintained at green islands but declined in tissue surrounding the infection. In younger leaves however, photosynthesis was reduced only at infection sites. Support for the local modification of host physiology came from quantitative reverse transcription-poly...
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▪ Infection structures of phytopathogenic fungi are modified hyphae specialized for the invasion of plant tissues. Initial events are adhesion to the cuticle and directed growth of the germ tube on the plant surface. At the site of... more
▪ Infection structures of phytopathogenic fungi are modified hyphae specialized for the invasion of plant tissues. Initial events are adhesion to the cuticle and directed growth of the germ tube on the plant surface. At the site of penetration, appressoria are often formed that may have melanized walls and develop high turgor pressure to support the penetration process. The penetration hypha accumulates components of the cytoskeleton in the tip and secretes a variety of cell wall–degrading enzymes in a highly regulated fashion in order to penetrate the cuticle and the plant cell wall. This article reviews recent papers on the cytology, physiology, and molecular biology of the penetration process.
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Small Ras superfamily GTPases are highly conserved regulatory factors of fungal cell wall biosynthesis and morphogenesis. Previous experiments have shown that the Rho4-like protein of the maize anthracnose fungus Colletotrichum... more
Small Ras superfamily GTPases are highly conserved regulatory factors of fungal cell wall biosynthesis and morphogenesis. Previous experiments have shown that the Rho4-like protein of the maize anthracnose fungus Colletotrichum graminicola, formerly erroneously annotated as a Rho1 protein, physically interacts with the β-1,3-glucan synthase Gls1 (Lange et al., 2014; Curr. Genet. 60:343–350). Here, we show that Rho4 is required for β-1,3-glucan synthesis. Accordingly, Δrho4 strains formed distorted vegetative hyphae with swellings, and exhibited strongly reduced rates of hyphal growth and defects in asexual sporulation. Moreover, on host cuticles, conidia of Δrho4 strains formed long hyphae with hyphopodia, rather than short germ tubes with appressoria. Hyphopodia of Δrho4 strains exhibited penetration defects and often germinated laterally, indicative of cell wall weaknesses. In planta differentiated infection hyphae of Δrho4 strains were fringy, and anthracnose disease symptoms cau...