Necrotrophic Plant Pathogen Research Articles

Membrane protein Bcsdr2 mediates biofilm integrity, hyphal growth and virulence of Botrytis cinerea

Grey mould caused by Botrytis cinerea is a devastating disease responsible for large losses to agricultural production, and B. cinerea is a necrotrophic model fungal plant pathogen. Membrane proteins are important targets of fungicides and hotspots in the research and development of fungicide products. Wuyiencin affects the permeability and pathogenicity of B. cinerea, parallel reaction monitoring revealed the association of membrane protein Bcsdr2, and the bacteriostatic mechanism of wuyiencin was elucidated. In the present work, we generated and characterised ΔBcsdr2 deletion and complemented mutant B. cinerea strains. The ΔBcsdr2 deletion mutants exhibited biofilm loss and dissolution, and their functional activity was illustrated by reduced necrotic colonisation on strawberry and grape fruits. Targeted deletion of Bcsdr2 also blocked several phenotypic defects in aspects of mycelial growth, conidiation and virulence. All phenotypic defects were restored by targeted gene complementation. The roles of Bcsdr2 in biofilms and pathogenicity were also supported by quantitative real-time RT-PCR results showing that phosphatidylserine decarboxylase synthesis gene Bcpsd and chitin synthase gene BcCHSVII were downregulated in the early stages of infection for the ΔBcsdr2 strain. The results suggest that Bcsdr2 plays important roles in regulating various cellular processes in B. cinerea.Key points• The mechanism of wuyiencin inhibits B. cinerea is closely associated with membrane proteins.• Wuyiencin can downregulate the expression of the membrane protein Bcsdr2 in B. cinerea.• Bcsdr2 is involved in regulating B. cinerea virulence, growth and development.

Genome-wide analysis of the PG gene family in Penicillium expansum: Expression during the infection stage in pear fruits (Pyrus bretschneideri)

Penicillium expansum is a necrotrophic plant pathogen that has a broad range of fruit hosts and is responsible for blue mold rot. This rot has a significant impact on the fruit industry, causing economic losses during storage, transport, and sale. Polygalacturonase (PGs) enzymes are essential for pathogenic fungi to break down pectin, as they hydrolyze the polygalacturonic acid chain along the oxygen bridge. This enzyme has been identified in decayed tissue and is linked to multiple soft rot diseases. This study encompasses a thorough examination of the PGs gene family within the genome of P. expansum, utilizing a genome-wide analysis approach. As a result, a total of seven PG genes were successfully found. These genes were phylogenetically divided into four distinct clades. The gene structural characteristics of each group exhibited a fundamental level of conservation. Furthermore, the gene architectures and motif compositions exhibited robust evidence in favor of the credibility of the phylogenetic relationship. Motif analysis reveals that most of the motifs predicted had Glycosyl hydrolases family 28 conserved domains (PF00295). Analysis of RNA-seq data and the PG genes identified by RT-qPCR were mostly upregulated. Hence, comprehending the characteristics of PePGs holds significant scientific importance and will offer comprehensive insights into the PG superfamily in P. expansum and will make a valuable contribution to the ongoing validation of functional genes.

Evaluation of cell death-inducing activity of Monilinia spp. effectors in several plants using a modified TRV expression system.

Brown rot is the most important fungal disease affecting stone fruit and it is mainly caused by Monilinia fructicola, M. laxa and M. fructigena. Monilinia spp. are necrotrophic plant pathogens with the ability to induce plant cell death by the secretion of different phytotoxic molecules, including proteins or metabolites that are collectively referred to as necrotrophic effectors (NEs). We exploited the genomes of M. fructicola, M. laxa and M. fructigena to identify their common group of secreted effector proteins and tested the ability of a selected set of effectors to induce cell death in Nicotiana benthamiana, Solanum lycopersicum and Prunus spp. leaves. Fourteen candidate effector genes of M. fructicola, which displayed high expression during infection, were transiently expressed in plants by agroinfiltration using a modified Tobacco Rattle Virus (TRV)-based expression system. Some, but not all, effectors triggered leaf discoloration or cell death in N. benthamiana and S. lycopersicum, which are non-hosts for Monilinia and in Prunus spp., which are the natural hosts. The effector MFRU_030g00190 induced cell death in almost all Prunus genotypes tested, but not in the Solanaceous plants, while MFRU_014g02060, which is an ortholog to BcNep1, caused necrosis in all plant species tested. This method provides opportunities for screening Prunus germplasm with Monilinia effector proteins, to serve as a tool for identifying genetic loci that confer susceptibility to brown rot disease.

Bacteriophage cocktail for biocontrol of soft rot disease caused by Pectobacterium species in Chinese cabbage.

Pectobacterium spp. are necrotrophic plant pathogens that cause the soft rot disease in Chinese cabbage, resulting in severe yield loss. The use of conventional antimicrobial agents, copper-based bactericides, and antibiotics has encountered several limitations, such as bioaccumulation on plants and microbial resistance. Bacteriophages (phages) are considered promising alternative antimicrobial agents against diverse phytopathogens. In this study, we isolated and characterized two virulent phages (phiPccP-2 and phiPccP-3) to develop a phage cocktail. Morphological and genomic analyses revealed that two phages belonged to the Tevenvirinae and Mccorquodalevirinae subfamilies, respectively. The phiPccP-2 and phiPccP-3 phages, which have a broad host range, were stable at various environmental conditions, such as various pHs and temperatures and exposure to ultraviolet light. The phage cocktail developed using these two lytic phages inhibited the emergence of phage-resistant bacteria compared to single-phage treatments in in vitro challenge assays. The phage cocktail treatment effectively prevented the development of soft rot symptom in matured Chinese cabbage leaves. Additionally, the phage cocktail comprising three phages (phiPccP-1, phiPccP-2, and phiPccP-3) showed superior biocontrol efficacy against the mixture of Pectobacterium strains in Chinese cabbage seedlings. These results suggest that developing phage cocktails is an effective approach for biocontrol of soft rot disease caused by Pectobacterium strains in crops compared to single-phage treatments. KEY POINTS: •Two newly isolated Pectobacterium phages, phiPccP-2 and phiPccP-3, infected diverse Pectobacterium species and effectively inhibited the emergence of phage-resistant bacteria. •Genomic and physiological analyses suggested that both phiPccP-2 and phiPccP-3 are lytic phages and that their lytic activities are stable in the environmental conditions under which Chinese cabbage grows. •Treatment using a phage cocktail comprising phiPccP-2 and phiPccP-3 efficiently suppressed soft rot disease in detached mature leaves and seedlings of Chinese cabbage, indicating the applicability of the phage cocktail as an alternative antimicrobial agent.

Plant hypersensitive induced reaction protein facilitates cell death induced by secreted xylanase associated with the pathogenicity of Sclerotinia sclerotiorum.

Necrotrophic fungal plant pathogens employ cell death-inducing proteins (CDIPs) to facilitate infection. However, the specific CDIPs and their mechanisms in pathogenic processes of Sclerotinia sclerotiorum, a necrotrophic pathogen that causes disease in many economically important crop species, have not yet been clearly defined. This study found that S. sclerotiorum secretes SsXyl2, a glycosyl hydrolase family 11 xylanase, at the late stage of hyphal infection. SsXyl2 targets the apoplast of host plants to induce cell death independent of xylanase activity. Targeted disruption of SsXyl2 leads to serious impairment of virulence, which can be recovered by a catalytically impaired SsXyl2 variant, thus supporting the critical role of cell death-inducing activity of SsXyl2 in establishing successful colonization of S. sclerotiorum. Remarkably, infection by S. sclerotiorum induces the accumulation of Nicotiana benthamiana hypersensitive-induced reaction protein 2 (NbHIR2). NbHIR2 interacts with SsXyl2 at the plasma membrane and promotes its localization to the cell membrane and cell death-inducing activity. Furthermore, gene-edited mutants of NbHIR2 displayed increased resistance to the wild-type strain of S. sclerotiorum, but not to the SsXyl2-deletion strain. Hence, SsXyl2 acts as a CDIP that manipulates host cell physiology by interacting with hypersensitive induced reaction protein to facilitate colonization by S. sclerotiorum. These findings provide valuable insights into the pathogenic mechanisms of CDIPs in necrotrophic pathogens and lead to a more promising approach for breeding resistant crops against S. sclerotiorum.

An efficient targeted gene deletion approach for Cochliobolus heterostrophus using Agrobacterium tumefaciens-mediated transformation

Cochliobolus heterostrophus is a plant pathogenic fungus of southern corn leaf blight, which has been regarded as a model necrotrophic plant pathogen. Many methods have been developed to knock out targeted genes in C. heterostrophus, of which the most widely-used one is protoplast-mediated transformation. However, there are several problems of this method associated with protoplast preparation, DNA product, time consumption, or high cost. In this study, a highly efficient target gene deletion approach in C. heterostrophus was established and optimized, based on Agrobacterium tumefaciens-mediated transformation (ATMT); the transformation efficiency of this approach was 85–88 transformants per 105 conidia, and the homologous recombination efficiency was approximately 68.3%. Furthermore, six gene knockout mutants of C. heterostrophus were obtained using this ATMT method. The phenotypes of this fungus altered in the mutant strains, and the virulence of the mutants significantly reduced compared to of the wild type strain. Taken together, this ATMT system established in this study can be used as a genetic manipulation tool for C. heterostrophus, to better understand the functions of genes and its relation to virulence.

Genome Resource of Sclerotinia nivalis Strain SnTB1, a Plant Pathogen Isolated from American Ginseng

Sclerotinia fungi are necrotrophic plant pathogens that can infect a wide range of plant species and persist in natural conditions. Sclerotinia nivalis is a recently reported species in Sclerotinia and can cause white rot on many plant species. We obtained the whole genome sequence of S. nivalis strain SnTB1 assembled from Oxford Nanopore reads and further polished with Illumina reads. This assembly consists of 32 contigs with a total length of 50.37 Mb, a contig N50 of 2.74 Mb, a GC content of 39.53%, and a repeat element content of 20.68%. In total, 13,276 candidate protein-coding genes were identified and functionally annotated, including those encoding glycometabolic enzymes, proteolytic enzymes, and potential effectors. This is the first available reference genome resource for S. nivalis. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

The Botrytis cinerea transglycosylase BcCrh4 is a cell death-inducing protein with cell death-promoting and -suppressing domains.

Botrytis cinerea is a necrotrophic fungal plant pathogen that causes grey mould and rot diseases in many crops. Here, we show that the B. cinerea BcCrh4 transglycosylase is secreted during plant infection and induces plant cell death and pattern-triggered immunity (PTI), fulfilling the characteristics of a cell death-inducing protein (CDIP). The CDIP activity of BcCrh4 is independent of the transglycosylase enzymatic activity, it takes place in the apoplast and does not involve the receptor-like kinases BAK1 and SOBIR1. During saprophytic growth, BcCrh4 is localized in the endoplasmic reticulumand in vacuoles, but during plant infection, it accumulates in infection cushions (ICs) and is then secreted to the apoplast. Two domains within the BcCrh4 protein determine the CDIP activities: a 20aa domain at the N' end activates intense cell death and PTI, while a stretch of 52aa in the middle of the protein induces a weaker response and suppresses the activity of the 20aa N' domain. Deletion of bccrh4 affected fungal development and IC formation in particular, resulting in reduced virulence. Collectively, our findings demonstrate that BcCrh4 is required for fungal development and pathogenicity, and hint at a dual mechanism that balances the virulence activity of this, and potentially other CDIPs.

The secondary metabolites profiling of the phytopathogenic fungus Sclerotinia Sclerotiorum

Sclerotinia sclerotiorum is a necrotrophic plant pathogen causing more than 60 different disease symptoms in approximately 400 plants globally. Hence, due to this distinctive characteristic, S. sclerotiorum has been the subject of various research to comprehend its pathogenicity mechanism, including virulent genes, proteins, and metabolites. Likewise, the genomic annotation of S. sclerotiorum uncovered its remarkable potential for producing secondary metabolites, of which genome mining has additionally prompted the disclosure of these uncharacterized metabolic pathways, which might aid the pathogenicity process. To comprehend the secondary metabolites secreted by S. sclerotiorum that might be involved in its pathogenicity, a secondary metabolite-level investigation of this plant pathogen was performed. Profiling and characterizing these secondary metabolites produced during in vitro germination would increase the current knowledge of this pathogen. In this study, S. sclerotiorum secondary metabolites profile examination was conducted, utilizing the Ultra-High Resolution Qq-Time-Of-Flight mass spectrometer (UHR-QqTOF). Proficient data analysis and verification with the genomic pathways of S. sclerotiorum gave an unequivocal metabolome profile of this pathogen. Two hundred and thirty secondary metabolites were identified in all three biological replicates, and their bodily functions were identified.

The Genetic Requirements for HiVir-Mediated Onion Necrosis by Pantoea ananatis, a Necrotrophic Plant Pathogen.

Pantoea ananatis is an unusual bacterial pathogen that lacks typical virulence determinants yet causes extensive necrosis in onion foliage and bulb tissues. The onion necrosis phenotype is dependent on the expression of the phosphonate toxin, pantaphos, which is synthesized by putative enzymes encoded by the HiVir (high virulence) gene cluster. The genetic contributions of individual hvr genes in HiVir-mediated onion necrosis remain largely unknown, except for the first gene, hvrA (phosphoenolpyruvate mutase, pepM), whose deletion resulted in the loss of onion pathogenicity. In this study, using gene-deletion mutation and complementation, we report that, of the ten remaining genes, hvrB to hvrF are also strictly required for the HiVir-mediated onion necrosis and in-planta bacterial growth, whereas hvrG to hvrJ partially contributed to these phenotypes. As the HiVir gene cluster is a common genetic feature shared among the onion-pathogenic P. ananatis strains that could serve as a useful diagnostic marker of onion pathogenicity, we sought to understand the genetic basis of HiVir-positive yet phenotypically deviant (non-pathogenic) strains. We identified and genetically characterized inactivating single nucleotide polymorphisms in the essential hvr genes of six phenotypically deviant P. ananatis strains. Finally, inoculation of cell-free spent medium of the isopropylthio-β-galactoside (IPTG)-inducible promoter (Ptac)-driven HiVir strain caused P.ananatis-characteristic red onion scale necrosis as well as cell death symptoms in tobacco. Co-inoculation of the spent medium with essential hvr mutant strains restored in-planta populations of the strains to the wild-type level, suggesting that necrotic tissues are important for the proliferation of P. ananatis in onion. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Botrytis cinerea

Chen, Zhang et al. introduce the necrotrophic fungal plant pathogen Botrytis cinerea more commonly known as gray mold.

Understanding bud rot development, caused by Botrytis cinerea, on cannabis (Cannabis sativa L.) plants grown under greenhouse conditions

Botrytis cinerea is a widespread necrotrophic plant pathogen that causes diseases on >1000 plant species, including vegetables and ornamental greenhouse crops. On cannabis ( Cannabis sativ a L.), the pathogen is responsible for causing “bud rot”, a major disease affecting the inflorescences (compound flowers), as well as seedling damping-off and leaf blight under certain conditions. During greenhouse cultivation, Botrytis cinerea can destroy cannabis inflorescences rapidly under optimal relative humidity conditions (>70%) and moderate temperatures (17–24 °C). Little is currently known about the host–pathogen interactions of Botrytis cinerea on cannabis. Information gleaned from other hosts can provide valuable insights for comparative purposes to understand disease development, epidemiology, and pathogenicity of Botrytis cinerea on cannabis crops. This review describes the pathogenesis and host responses to Botrytis infection and assesses potential mechanisms involved in disease resistance. The effects of microclimatic and other environmental conditions on disease development, strategies for early disease detection using prediction models, and the application of biological control agents that can prevent Botrytis cinerea development on cannabis are discussed. Other potential disease management approaches to reduce the impact of Botrytis bud rot are also reviewed. Numerous opportunities for conducting additional research to better understand the cannabis– Botrytis cinerea interaction are identified.

Membrane Protein Bcest Is Involved in Hyphal Growth, Virulence and Stress Tolerance of Botrytis cinerea.

Botrytis cinerea is a necrotrophic model fungal plant pathogen that causes grey mould, a devastating disease responsible for large losses in the agriculture sector. As important targets of fungicides, membrane proteins are hot spots in the research and development of fungicide products. We previously found that membrane protein Bcest may be closely related to the pathogenicity of Botrytis cinerea. Herein, we further explored its function. We generated and characterised ΔBcest deletion mutants of B. cinerea and constructed complemented strains. The ΔBcest deletion mutants exhibited reduced conidia germination and germ tube elongation. The functional activity of ΔBcest deletion mutants was investigated by reduced necrotic colonisation of B. cinerea on grapevine fruits and leaves. Targeted deletion of Bcest also blocked several phenotypic defects in aspects of mycelial growth, conidiation and virulence. All phenotypic defects were restored by targeted-gene complementation. The role of Bcest in pathogenicity was also supported by reverse-transcriptase real-time quantitative PCR results indicating that melanin synthesis gene Bcpks13 and virulence factor Bccdc14 were significantly downregulated in the early infection stage of the ΔBcest strain. Taken together, these results suggest that Bcest plays important roles in the regulation of various cellular processes in B. cinerea.

Variable Tandem Glycine-Rich Repeats Contribute to Cell Death-Inducing Activity of a Glycosylphosphatidylinositol-Anchored Cell Wall Protein That Is Associated with the Pathogenicity of Sclerotinia sclerotiorum.

Glycosylphosphatidylinositol (GPI) anchoring of proteins is a conserved posttranslational modification in eukaryotes. GPI-anchored proteins are widely distributed in fungal plant pathogens, but the specific roles of the GPI-anchored proteins in the pathogenicity of Sclerotinia sclerotiorum, a devastating necrotrophic plant pathogen with a worldwide distribution, remain largely unknown. This research addresses SsGSR1, which encodes an S. sclerotiorum glycine- and serine-rich protein named SsGsr1 with an N-terminal secretory signal and a C-terminal GPI-anchor signal. SsGsr1 is located at the cell wall of hyphae, and deletion of SsGSR1 leads to abnormal cell wall architecture and impaired cell wall integrity of hyphae. The transcription levels of SsGSR1 were maximal in the initial stage of infection, and SsGSR1-deletion strains showed impaired virulence in multiple hosts, indicating that SsGSR1 is critical for the pathogenicity. Interestingly, SsGsr1 targeted the apoplast of host plants to induce cell death that relies on the glycine-rich 11-amino-acid repeats arranged in tandem. The homologs of SsGsr1 in Sclerotinia, Botrytis, and Monilinia species contain fewer repeat units and have lost their cell death activity. Moreover, allelic variants of SsGSR1 exist in field isolates of S. sclerotiorum from rapeseed, and one of the variants lacking one repeat unit results in a protein that exhibits loss of function relative to the cell death-inducing activity and the virulence of S. sclerotiorum. Taken together, our results demonstrate that a variation in tandem repeats provides the functional diversity of GPI-anchored cell wall protein that, in S. sclerotiorum and other necrotrophic pathogens, allows successful colonization of the host plants. IMPORTANCE Sclerotinia sclerotiorum is an economically important necrotrophic plant pathogen and mainly applies cell wall-degrading enzymes and oxalic acid to kill plant cells before colonization. In this research, we characterized a glycosylphosphatidylinositol (GPI)-anchored cell wall protein named SsGsr1, which is critical for the cell wall architecture and the pathogenicity of S. sclerotiorum. Additionally, SsGsr1 induces rapid cell death of host plants that is dependent on glycine-rich tandem repeats. Interestingly, the number of repeat units varies among homologs and alleles of SsGsr1, and such a variation creates alterations in the cell death-inducing activity and the role in pathogenicity. This work advances our understanding of the variation of tandem repeats in accelerating the evolution of a GPI-anchored cell wall protein associated with the pathogenicity of necrotrophic fungal pathogens and prepares the way toward a fuller understanding of the interaction between S. sclerotiorum and host plants.

Quantification of Botrytis cinerea Growth in Arabidopsis thaliana.

Yield losses attributed to plant pathogens pose a serious threat to plant productivity and food security. Botrytis cinerea is one of the most devastating plant pathogens, infecting a wide array of plant species; it has also been established as a model organism to study plant-pathogen interactions. In this context, development of different assays to follow the relative success of B. cinerea infections is required. Here, we describe two methods to quantify B. cinerea development in Arabidopsis thaliana genotypes through measurements of lesion development and quantification of fungal genomic DNA in infected tissues. This provides two independent techniques that are useful in assessing the susceptibility or tolerance of different Arabidopsis genotypes to B. cinerea. Key features Protocol for the propagation of the necrotrophic plant pathogen fungus Botrytis cinerea and spore production. Two methods of Arabidopsis thaliana infection with the pathogen using droplet and spray inoculation. Two readouts, either by measuring lesion size or by the quantification of fungal DNA using quantitative PCR. The two methods are applicable across plant species susceptible the B. cinerea. Graphical overview A simplified overview of the droplet and spray infection methods used for the determination of B. cinerea growth in different Arabidopsis genotypes.

The AP2/ERF transcription factor ORA59 regulates ethylene-induced phytoalexin synthesis through modulation of an acyltransferase gene expression.

The gaseous ethylene (ET) and the oxylipin-derived jasmonic acid (JA) in plants jointly regulate an arsenal of pathogen responsive genes involved in defending against necrotrophic pathogens. The APETALA2 (AP2)/ETHYLENE RESPONSE FACTOR (ERF) transcription factor ORA59 is a major positive regulator of the ET/JA-mediated defense pathway in Arabidopsis thaliana. The Arabidopsis agmatine coumaroyltransferase (AtACT) catalyzes the formation of hydroxycinnamic acid amides (HCAAs) which are effective toxic antimicrobial substances known as phytoalexins and play an important role in plant defense response. However, induction and regulation of AtACT gene expression and HCAAs synthesis in plants remain less understood. Through gene coexpression network analysis, we identified a list of GCC-box cis-element containing genes that were coexpressed with ORA59 under diverse biotic stress conditions and might be potential downstream targets of this AP2/ERF-domain transcription factor. Particularly, ORA59 directly binds to AtACT gene promoter via the GCC-boxes and activates AtACT gene expression. The ET precursor 1-aminocyclopropane-1-carboxylic acid (ACC)-treatment significantly induces AtACT gene expression. Both ORA59 and members of the class II TGA transcription factors are indispensable for ACC-induced AtACT expression. Interestingly, the expression of AtACT is also subject to the signaling crosstalk of the salicylic acid- and ET/JA-mediated defense response pathways. In addition, we found that genes of the phenylpropanoid metabolism pathway were specifically induced by Botrytis cinerea. Taking together, these evidence suggest that the ET/JA signaling pathway activate the expression of AtACT to increase antimicrobial HCAAs production through the transcription factor ORA59 in response to the infection of necrotrophic plant pathogens.

Mycoviral gene integration converts a plant pathogenic fungus into a biocontrol agent.

Mycovirus-infected fungi can suffer from poor growth, attenuated pigmentation, and virulence. However, the molecular mechanisms of how mycoviruses confer these symptoms remain poorly understood. Here, we report a mycovirus Stemphylium lycopersici alternavirus 1 (SlAV1) isolated from a necrotrophic plant pathogen Stemphylium lycopersici that causes altered colony pigmentation and hypovirulence by specifically interfering host biosynthesis of Altersolanol A, a polyketide phytotoxin. SlAV1 significantly down-regulates a fungal polyketide synthase (PKS1), the core enzyme of Altersolanol A biosynthesis. PKS1 deletion mutants do not accumulate Altersolanol A and lose pathogenicity to tomato and lettuce. Transgenic expression of SlAV1 open-reading frame 3 (ORF3) in S. lycopersici inhibits fungal PKS1 expression and Altersolanol A accumulation, leading to symptoms like SlAV1-infected fungal strains. Multiple plant species sprayed with mycelial suspension of S. lycopersici or S. vesicarium strains integrating and expressing ORF3 display enhanced resistance against virulent strains, converting the pathogenic fungi into biocontrol agents. Hence, our study not only proves inhibiting a key enzyme of host phytotoxin biosynthesis as a molecular mechanism underlying SlAV1-mediated hypovirulence of Stemphylium spp., but also demonstrates the potential of mycovirus-gene integrated fungi as a potential biocontrol agent to protect plants from fungal diseases.

Botrytis cinerea Transcription Factor BcXyr1 Regulates (Hemi-)Cellulase Production and Fungal Virulence.

Botrytis cinerea is an agriculturally notorious plant-pathogenic fungus with a broad host range. During plant colonization, B. cinerea secretes a wide range of plant-cell-wall-degrading enzymes (PCWDEs) that help in macerating the plant tissue, but their role in pathogenicity has been unclear. Here, we report on the identification of a transcription factor, BcXyr1, that regulates the production of (hemi-)cellulases and is necessary for fungal virulence. Deletion of the bcxyr1 gene led to impaired spore germination and reduced fungal virulence and reactive oxygen species (ROS) production in planta. Secreted proteins collected from the bcxyr1 deletion strain displayed a weaker cell-death-inducing effect than the wild-type secretome when infiltrated to Nicotiana benthamiana leaves. Transcriptome sequencing (RNA-seq) analysis revealed 41 genes with reduced expression in the Δbcxyr1 mutant compared with those in the wild-type strain, of which half encode secreted proteins that are particularly enriched in carbohydrate-active enzyme (CAZyme)-encoding genes. Among them, we identified a novel putative expansin-like protein that was necessary for fungal virulence, supporting the involvement of BcXyr1 in the regulation of extracellular virulence factors. IMPORTANCE PCWDEs are considered important components of the virulence arsenal of necrotrophic plant pathogens. However, despite intensive research, the role of PCWDEs in the pathogenicity of necrotrophic phytopathogenic fungi remains ambiguous. Here, we demonstrate that the transcription factor BcXyr1 regulates the expression of a specific set of secreted PCWDE-encoding genes and that it is essential for fungal virulence. Furthermore, we identified a BcXyr1-regulated expansin-like gene that is required for fungal virulence. Our findings provide strong evidence for the importance of PCWDEs in the pathogenicity of B. cinerea and highlight specific PCWDEs that might be more important than others.

Automated quantitative measurement of yellow halos suggests activity of necrotrophic effectors in Septoria tritici blotch.

Many necrotrophic plant pathogens utilize host selective toxins or necrotrophic effectors during the infection process. We hypothesized that the chlorotic yellow halos frequently observed around necrotic lesions caused by the wheat pathogen Zymoseptoria tritici may result from the activity of necrotrophic effectors interacting with the products of toxin sensitivity genes. As an initial step towards testing this hypothesis, we developed an automated image analysis (AIA) workflow that could quantify the degree of yellow halo formation occurring in wheat leaves naturally infected by a highly diverse pathogen population under field conditions. This AIA based on statistical learning was applied to more than 10,000 naturally infected leaves collected from 335 wheat cultivars grown in a replicated field experiment. We measured a high heritability (h2 = 0.71) for the degree of yellow halo formation, suggesting that this quantitative trait has a significant genetic component. Using genome wide association mapping, we identified six chromosome segments significantly associated with the yellow halo phenotype. Most of these segments contained candidate genes associated with targets of necrotrophic effectors in other necrotrophic pathogens. Our findings conform with the hypothesis that toxin sensitivity genes may account for a significant fraction of the observed variation in quantitative resistance to Septoria tritici blotch.

Defense Surveillance System at the Interface: Response of Rice Towards Rhizoctonia solani During Sheath Blight Infection.

Sheath blight of rice caused by necrotrophic plant pathogen Rhizoctonia solani is one of the most common fungal diseases of rice leading to significant yield loss. Among the defense responses exhibited by the host plants towards fungal infections, those functional within the apoplast contribute significantly. Here, we have studied apoplastic defense response of rice towards R. solani during sheath blight infection. The transcriptome of R. solani-infected rice plants was compared with that of uninfected rice, to identify the set of defense genes that undergo differential expression and code for proteins with a predicted N-terminal signal peptide. Significant changes in the stress-responsive, molecular signal perception, protein modification, and metabolic process pathways represented by a group of differentially expressed genes were observed. Our data also revealed two secreted protease inhibitors from rice that exhibit increased expression during R. solani infection and induce disease resistance when expressed in Nicotiana benthamiana. [Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.