ATR2C ala2 from Arabidopsis-infecting downy mildew requires 4 TIR-NLR immune receptors for full recognition.

Arabidopsis Col-0 RPP2A and RPP2B confer recognition of Arabidopsis downy mildew (Hyaloperonospora arabidopsidis [Hpa]) isolate Cala2, but the identity of the recognized ATR2Cala2 effector was unknown. To reveal ATR2Cala2, an F2 population was generated from a cross between Hpa-Cala2 and Hpa-Noks1. We identified ATR2Cala2 as a non-canonical RxLR-type effector that carries a signal peptide, a dEER motif, and WY domains but no RxLR motif. Recognition of ATR2Cala2 and its effector function were verified by biolistic bombardment, ectopic expression and Hpa infection. ATR2Cala2 is recognized in accession Col-0 but not in Ler-0 in which RPP2A and RPP2B are absent. In ATR2Emoy2 and ATR2Noks1 alleles, a frameshift results in an early stop codon. RPP2A and RPP2B are essential for the recognition of ATR2Cala2. Stable and transient expression of ATR2Cala2 under 35S promoter in Arabidopsis and Nicotiana benthamiana enhances disease susceptibility. Two additional Col-0 TIR-NLR (TNL) genes (RPP2C and RPP2D) adjacent to RPP2A and RPP2B are quantitatively required for full resistance to Hpa-Cala2. We compared RPP2 haplotypes in multiple Arabidopsis accessions and showed that all four genes are present in all ATR2Cala2-recognizing accessions.

Decoding the Arsenal: Protist Effectors and Their Impact on Photosynthetic Hosts.

Interactions between various microbial pathogens including viruses, bacteria, fungi, oomycetes, and their plant hosts have traditionally been the focus of phytopathology. In recent years, a significant and growing interest in the study of eukaryotic microorganisms not classified among fungi or oomycetes has emerged. Many of these protists establish complex interactions with photosynthetic hosts, and understanding these interactions is crucial in understanding the dynamics of these parasites within traditional and emerging types of farming, including marine aquaculture. Many phytopathogenic protists are biotrophs with complex polyphasic life cycles, which makes them difficult or impossible to culture, a fact reflected in a wide gap in the availability of comprehensive genomic data when compared to fungal and oomycete plant pathogens. Furthermore, our ability to use available genomic resources for these protists is limited by the broad taxonomic distance that these organisms span, which makes comparisons with other genomic datasets difficult. The current rapid progress in genomics and computational tools for the prediction of protein functions and interactions is revolutionizing the landscape in plant pathology. This is also opening novel possibilities, specifically for a deeper understanding of protist effectors. Tools like AlphaFold2 enable structure-based function prediction of effector candidates with divergent protein sequences. In turn, this allows us to ask better biological questions and, coupled with innovative experimental strategies, will lead into a new era of effector research, especially for protists, to expand our knowledge on these elusive pathogens and their interactions with photosynthetic hosts. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Glyoxalase I-4 functions downstream of NAC72 to modulate downy mildew resistance in grapevine.

Glyoxalase I (GLYI) is part of the glyoxalase system; its major function is the detoxification of α-ketoaldehydes, including the potent and cytotoxic methylglyoxal (MG). Methylglyoxal disrupts mitochondrial respiration and increases production of reactive oxygen species (ROS), which also increase during pathogen infection of plant tissues; however, there have been few studies relating the glyoxalase system to the plant pathogen response. We used the promoter of VvGLYI-4 to screen the upstream transcription factors and report a NAC (NAM/ATAF/CUC) domain-containing transcription factor VvNAC72 in grapevine, which is localized to the nucleus. Our results show that VvNAC72 expression is induced by downy mildew, Plasmopara viticola, while the transcript level of VvGLYI-4 decreases. Further analysis revealed that VvNAC72 can bind directly to the promoter region of VvGLYI-4 via the CACGTG element, leading to inhibition of VvGLYI-4 transcription. Stable overexpression of VvNAC72 in grapevine and tobacco showed a decreased expression level of VvGLYI-4 and increased content of MG and ROS, as well as stronger resistance to pathogen stress. Taken together, these results demonstrate that grapevine VvNAC72 negatively modulates detoxification of MG through repression of VvGLYI-4, and finally enhances resistance to downy mildew, at least in part, via the modulation of MG-associated ROS homeostasis through a salicylic acid-mediated defense pathway.

The Mitogen-Activated Protein Kinase PlMAPK2 Is Involved in Zoosporogenesis and Pathogenicity of Peronophythoralitchii.

As an evolutionarily conserved pathway, mitogen-activated protein kinase (MAPK) cascades function as the key signal transducers that convey information by protein phosphorylation. Here we identified PlMAPK2 as one of 14 predicted MAPKs encoding genes in the plant pathogenic oomycete Peronophythora litchii. PlMAPK2 is conserved in P.litchii and Phytophthora species. We found that PlMAPK2 was up-regulated in sporangium, zoospore, cyst, cyst germination and early stage of infection. We generated PlMAPK2 knockout mutants using the CRISPR/Cas9 method. Compared with wild-type strain, the PlMAPK2 mutants showed no significant difference in vegetative growth, oospore production and sensitivity to various abiotic stresses. However, the sporangium release was severely impaired. We further found that the cleavage of the cytoplasm into uninucleate zoospores was disrupted in the PlMAPK2 mutants, and this developmental phenotype was accompanied by reduction in the transcription levels of PlMAD1 and PlMYB1 genes. Meanwhile, the PlMAPK2 mutants exhibited lower laccase activity and reduced virulence to lychee leaves. Overall, this study identified a MAPK that is critical for zoosporogenesis by regulating the sporangial cleavage and pathogenicity of P.litchii, likely by regulating laccase activity.

The oligomeric states of elicitins affect the hypersensitive response and resistance in tobacco.

Successful plant defence against microbial pathogens is based on early recognition and fast activation of inducible responses. Key mechanisms include detection of microbe-associated molecular patterns by membrane-localized pattern recognition receptors that induce a basal resistance response. A well-described model of such responses to pathogens involves the interactions between Solanaceae plants and proteinaceous elicitors secreted by oomycetes, called elicitins. It has been hypothesized that the formation of oligomeric structures by elicitins could be involved in their recognition and activation of defensive transduction cascades. In this study, we tested this hypothesis using several approaches, and we observed differences in tobacco plant responses induced by the elicitin ß-cryptogein (ß-CRY) and its homodimer, ß-CRYDIM. We also found that the C-terminal domain of elicitins of other ELI (true-elicitin) clades plays a significant role in stabilization of their oligomeric structure and restraint in the cell wall. In addition, covalently cross-linking ß-CRYDIM impaired the formation of signalling complexes, thereby reducing its capacity to elicit the hypersensitive response and resistance in the host plant, with no significant changes in pathogenesis-related protein expression. By revealing the details of the effects of ß-CRY dimerization on recognition and defence responses in tobacco, our results shed light on the poorly understood role of elicitins' oligomeric structures in the interactions between oomycetes and plants.

Plasmopara viticola effector PvRXLR111 stabilizes VvWRKY40 to promote virulence.

Plasmopara viticola, the causal organism of grapevine downy mildew, secretes a vast array of effectors to manipulate host immunity. Previously, several cell death-inducing PvRXLR effectors have been identified, but their functions and host targets are poorly understood. Here, we investigated the role of PvRXLR111, a cell death-inducing RXLR effector, in manipulating plant immunity. When coexpressed with other PvRXLR effectors, PvRXLR111-induced cell death was prevented. Transient expression of PvRXLR111 in Nicotiana benthamiana suppressed bacterial flagellin peptide flg22-elicited immune responses and enhanced Phytophthora capsici infection. PvRXLR111 induction in Arabidopsis increased susceptibility to Hyaloperonospora arabidopsidis. PvRXLR111 expression in Pseudomonas syringae promoted bacterial colonization. By immunoprecipitation-mass spectrometry analysis, yeast two-hybrid, pull-down, and bimolecular fluorescence complementation assays, it was shown that PvRXLR111 interacted with Vitis vinifera putative WRKY transcription factor 40 (VvWRKY40), which increased VvWRKY40 stability. Transient expression of VvWRKY40 in N. benthamiana inhibited flg22-induced reactive oxygen species burst and enhanced P. capsici infection and silencing NbWRKY40 attenuated P. capsici colonization. These results suggest VvWRKY40 functions as a negative regulator in plant immunity and that PvRXLR111 suppresses host immunity by stabilizing VvWRKY40.

Plasmopara viticola effector PvRXLR53 suppresses innate immunity in Nicotiana benthamiana.

Plasmopara viticola, the casual oomycete of grapevine downy mildew, could cause yield loss and compromise berry quantity. Previously, we have identified several PvRXLR effectors that could suppress plant immunity to promote infection and disease development. In this study, the role of effector, PvRXLR53, in plant-microbe interaction was investigated. PvRXLR53 has several orthologs in other oomycetes and contains a functional signal peptide. Expression level of PvRXLR53 was already detected upon inoculation, further induced in the early stage after P. viticola inoculation and decreased to low level in the late infection stage in grapevine (Vitis vinifera 'Cabernet Sauvignon'). PvRXLR53 is localized in both nucleus and cytoplasm. When transiently expressed in Nicotiana benthamiana, PvRXLR53 suppressed oomycete elicitor INF1-triggered programmed cell death and defense gene expression, and Phytophthora capsici-induced reactive oxygen species production (ROS) and eventually resistance to P. capsici. In summary, these findings suggest that P. viticola secretes PvRXLR53 to suppress host immunity from the very early stage of infection.

Comparative transcriptome analysis between a resistant and a susceptible Chinese cabbage in response to Hyaloperonospora brassicae.

Downy mildew caused by Hyaloperonosporabrassicae (H. brassicae) leads to up to 90% of the crop yield loss in Chinese cabbage in China. A transcriptome analysis was carried out between a resistant line (13-13, R) and a susceptible line (15-14, S) of Chinese cabbage in response to H. brassicae. The NOISeq method was used to find differentially expressed genes (DEGs) between these two groups and GO and KEGG were carried out to find R genes related to downy mildew response of Chinese cabbage. qRT-PCR was carried out to verify the reliability of RNA-seq expression data. A total of 3,055 DEGs were screened out from 41,020 genes and clustered into 6 groups with distinct expression patterns. A total of 87 candidate DEGs were identified by functional annotation based on GO and KEGG analysis. These candidate genes are involved in plant-pathogen interaction pathway, among which 54 and 33 DEGs were categorized into plant-pathogen interaction proteins and transcription factors, respectively. Proteins encoded by these genes have been reported to play an important role in the pattern-triggered immunity (PTI) and effector-triggered immunity (ETI) processes of disease responses in some model plants, such as Arabidopsis, rice, tobacco, and tomato. However, little is known about the mechanisms of these genes in resistance to downy mildew in Chinese cabbage. Our findings are useful for further characterization of these candidate genes and helpful in breeding resistant strains.

Oregano essential oil vapour prevents Plasmopara viticola infection in grapevine (Vitis Vinifera) and primes plant immunity mechanisms.

The reduction of synthetic fungicides in agriculture is necessary to guarantee a sustainable production that protects the environment and consumers' health. Downy mildew caused by the oomycete Plasmopara viticola is the major pathogen in viticulture worldwide and responsible for up to 60% of pesticide treatments. Alternatives to reduce fungicides are thus utterly needed to ensure sustainable vineyard-ecosystems, consumer health and public acceptance. Essential oils (EOs) are amongst the most promising natural plant protection alternatives and have shown their antibacterial, antiviral and antifungal properties on several agricultural crops. However, the efficiency of EOs highly depends on timing, application method and the molecular interactions between the host, the pathogen and EO. Despite proven EO efficiency, the underlying processes are still not understood and remain a black box. The objectives of the present study were: a) to evaluate whether a continuous fumigation of a particular EO can control downy mildew in order to circumvent the drawbacks of direct application, b) to decipher molecular mechanisms that could be triggered in the host and the pathogen by EO application and c) to try to differentiate whether essential oils directly repress the oomycete or act as plant resistance primers. To achieve this a custom-made climatic chamber was constructed that enabled a continuous fumigation of potted vines with different EOs during long-term experiments. The grapevine (Vitis vinifera) cv Chasselas was chosen in reason of its high susceptibility to Plasmopara viticola. Grapevine cuttings were infected with P. viticola and subsequently exposed to continuous fumigation of different EOs at different concentrations, during 2 application time spans (24 hours and 10 days). Experiments were stopped when infection symptoms were clearly observed on the leaves of the control plants. Plant physiology (photosynthesis and growth rate parameters) were recorded and leaves were sampled at different time points for subsequent RNA extraction and transcriptomics analysis. Strikingly, the Oregano vulgare EO vapour treatment during 24h post-infection proved to be sufficient to reduce downy mildew development by 95%. Total RNA was extracted from leaves of 24h and 10d treatments and used for whole transcriptome shotgun sequencing (RNA-seq). Sequenced reads were then mapped onto the V. vinifera and P. viticola genomes. Less than 1% of reads could be mapped onto the P. viticola genome from treated samples, whereas up to 30% reads from the controls mapped onto the P. viticola genome, thereby confirming the visual observation of P. viticola absence in the treated plants. On average, 80% of reads could be mapped onto the V. vinifera genome for differential expression analysis, which yielded 4800 modulated genes. Transcriptomic data clearly showed that the treatment triggered the plant's innate immune system with genes involved in salicylic, jasmonic acid and ethylene synthesis and signaling, activating Pathogenesis-Related-proteins as well as phytoalexin synthesis. These results elucidate EO-host-pathogen interactions for the first time and indicate that the antifungal efficiency of EO is mainly due to the triggering of resistance pathways inside the host plants. This is of major importance for the production and research on biopesticides, plant stimulation products and for resistance-breeding strategies.

Plasmopara viticola effector PvRXLR131 suppresses plant immunity by targeting plant receptor-like kinase inhibitor BKI1.

The grapevine downy mildew pathogen Plasmopara viticola secretes a set of RXLR effectors (PvRXLRs) to overcome host immunity and facilitate infection, but how these effectors function is unclear. Here, the biological function of PvRXLR131 was investigated via heterologous expression. Constitutive expression of PvRXLR131 in Colletotrichum gloeosporioides significantly enhanced its pathogenicity on grapevine leaves. Constitutive expression of PvRXLR131 in Arabidopsis promoted Pseudomonas syringae DC3000 and P. syringae DC3000 (hrcC- ) growth as well as suppressed defence-related callose deposition. Transient expression of PvRXLR131 in Nicotiana benthamiana leaves could also suppress different elicitor-triggered cell death and inhibit plant resistance to Phytophthora capsici. Further analysis revealed that PvRXLR131 interacted with host Vitis vinifera BRI1 kinase inhibitor 1 (VvBKI1), and its homologues in N. benthamiana (NbBKI1) and Arabidopsis (AtBKI1). Moreover, bimolecular fluorescence complementation analysis revealed that PvRXLR131 interacted with VvBKI1 in the plasma membrane. Deletion assays showed that the C-terminus of PvRXLR131 was responsible for the interaction and mutation assays showed that phosphorylation of a conserved tyrosine residue in BKI1s disrupted the interaction. BKI1 was a receptor inhibitor of growth- and defence-related brassinosteroid (BR) and ERECTA (ER) signalling. When silencing of NbBKI1 in N. benthamiana, the virulence function of PvRXLR131 was eliminated, demonstrating that the effector activity is mediated by BKI1. Moreover, PvRXLR131-transgenic plants displayed BKI1-overexpression dwarf phenotypes and suppressed BR and ER signalling. These physiological and genetic data clearly demonstrate that BKI1 is a virulence target of PvRXLR131. We propose that P. viticola secretes PvRXLR131 to target BKI1 as a strategy for promoting infection.

Molecular characterization of Phytophthora palmivora responsible for bud rot disease of oil palm in Colombia.

Bud rot disease is a damaging disease of oil palm in Colombia. The pathogen responsible for this disease is a species of oomyctes, Phytophthora palmivora which is also the causal pathogen of several tropical crop diseases such as fruit rot and stem canker of cocoa, rubber, durian and jackfruit. No outbreaks of bud rot have been reported in oil palm in Malaysia or other Southeast Asian countries, despite this particular species being present in the region. Analysis of the genomic sequences of several genetic markers; the internal transcribe spacer regions (ITS) of the ribosomal RNA gene cluster, beta-tubulin gene, translation elongation factor 1 alpha gene (EF-1α), cytochrome c oxidase subunit I & II (COXI and COXII) gene cluster along with amplified fragment length polymorphism (AFLP) analyses have been carried out to investigate the genetic diversity and variation of P. palmivora isolates from around the world and from different hosts in comparison to Colombian oil palm isolates, as one of the steps in understanding why this species of oomycetes causes devastating damage to oil palm in Latin America but not in other regions. Phylogenetic analyses of these regions showed that the Colombian oil palm isolates were not separated from Malaysian isolates. AFLP analysis and a new marker PPHPAV, targeting an unclassified hypothetical protein, was found to be able to differentiate Malaysian and Colombian isolates and showed a clear clade separations. Despite this, pathogenicity studies did not show any significant differences in the level of aggressiveness of different isolates against oil palm in glasshouse tests.

Two grapevine metacaspase genes mediate ETI-like cell death in grapevine defence against infection of Plasmopara viticola.

Metacaspase, as hypersensitive response (HR) executors, has been identified in many plant species. Previously, the entire gene family of metacaspase has been uncovered, but there are still questions that remain unclear regarding HR-regulating gene members. In this study, based on metacaspase expression during different grapevine genotypes interacting with Plasmopara viticola, we identified MC2 and MC5 as candidates involved in HR. We overexpressed both metacaspases as GFP fusions in tobacco BY-2 cells to address subcellular localization and cellular functions. We found MC2 located at the ER, while MC5 was nucleocytoplasmic. In these overexpressor lines, cell death elicited by the bacterial protein harpin, is significantly enhanced, indicating MC2 and MC5 mediated defence-related programmed cell death (PCD). This effect was mitigated, when the membrane-located NADPH oxidase was inhibited by the specific inhibitor diphenylene iodonium, or when cells were complemented with methyl jasmonate, a crucial signal of basal immunity. Both findings are consistent with a role of MC2 and MC5 in cell death-related immunity. Using a dual-luciferase reporter system in grapevine cells we demonstrated both MC2 and MC5 promoter alleles from V. rupestris were more responsive to harpin than those from V. vinifera cv 'Müller-Thurgau', while they were not induced by MeJA as signal linked with basal immunity. These findings support a model, where MC2 and MC5 act specifically as executors of the HR.

Phytophthora infestans small phospholipase D-like proteins elicit plant cell death and promote virulence.

The successful invasion of host tissue by (hemi-)biotrophic plant pathogens is dependent on modifications of the host plasma membrane to facilitate the two-way transfer of proteins and other compounds. Haustorium formation and the establishment of extrahaustorial membranes are probably dependent on a variety of enzymes that modify membranes in a coordinated fashion. Phospholipases, enzymes that hydrolyse phospholipids, have been implicated as virulence factors in several pathogens. The oomycete Phytophthora infestans is a hemibiotrophic pathogen that causes potato late blight. It possesses different classes of phospholipase D (PLD) proteins, including small PLD-like proteins with and without signal peptide (sPLD-likes and PLD-likes, respectively). Here, we studied the role of sPLD-like-1, sPLD-like-12 and PLD-like-1 in the infection process. They are expressed in expanding lesions on potato leaves and during in vitro growth, with the highest transcript levels in germinating cysts. When expressed in planta in the presence of the silencing suppressor P19, all three elicited a local cell death response that was visible at the microscopic level as autofluorescence and strongly boosted in the presence of calcium. Moreover, inoculation of leaves expressing the small PLD-like genes resulted in increased lesion growth and greater numbers of sporangia, but this was abolished when mutated PLD-like genes were expressed with non-functional PLD catalytic motifs. These results show that the three small PLD-likes are catalytically active and suggest that their enzymatic activity is required for the promotion of virulence, possibly by executing membrane modifications to support the growth of P. infestans in the host.

Sunflower resistance to multiple downy mildew pathotypes revealed by recognition of conserved effectors of the oomycete Plasmopara halstedii.

Over the last 40 years, new sunflower downy mildew isolates (Plasmopara halstedii) have overcome major gene resistances in sunflower, requiring the identification of additional and possibly more durable broad-spectrum resistances. Here, 354 RXLR effectors defined in silico from our new genomic data were classified in a network of 40 connected components sharing conserved protein domains. Among 205 RXLR effector genes encoding conserved proteins in 17 P. halstedii pathotypes of varying virulence, we selected 30 effectors that were expressed during plant infection as potentially essential genes to target broad-spectrum resistance in sunflower. The transient expression of the 30 core effectors in sunflower and in Nicotiana benthamiana leaves revealed a wide diversity of targeted subcellular compartments, including organelles not so far shown to be targeted by oomycete effectors such as chloroplasts and processing bodies. More than half of the 30 core effectors were able to suppress pattern-triggered immunity in N. benthamiana, and five of these induced hypersensitive responses (HR) in sunflower broad-spectrum resistant lines. HR triggered by PhRXLRC01 co-segregated with Pl22 resistance in F3 populations and both traits localized in 1.7 Mb on chromosome 13 of the sunflower genome. Pl22 resistance was physically mapped on the sunflower genome recently sequenced, unlike all the other downy mildew resistances published so far. PhRXLRC01 and Pl22 are proposed as an avirulence/resistance gene couple not previously described in sunflower. Core effector recognition is a successful strategy to accelerate broad-spectrum resistance gene identification in complex crop genomes such as sunflower.

NLR Mutations Suppressing Immune Hybrid Incompatibility and Their Effects on Disease Resistance.

Genetic divergence between populations can lead to reproductive isolation. Hybrid incompatibilities (HI) represent intermediate points along a continuum toward speciation. In plants, genetic variation in disease resistance (R) genes underlies several cases of HI. The progeny of a cross between Arabidopsis (Arabidopsis thaliana) accessions Landsberg erecta (Ler, Poland) and Kashmir2 (Kas2, central Asia) exhibits immune-related HI. This incompatibility is due to a genetic interaction between a cluster of eight TNL (TOLL/INTERLEUKIN1 RECEPTOR-NUCLEOTIDE BINDING-LEU RICH REPEAT) RPP1 (RECOGNITION OF PERONOSPORA PARASITICA1)-like genes (R1-R8) from Ler and central Asian alleles of a Strubbelig-family receptor-like kinase (SRF3) from Kas2. In characterizing mutants altered in Ler/Kas2 HI, we mapped multiple mutations to the RPP1-like Ler locus. Analysis of these suppressor of Ler/Kas2 incompatibility (sulki) mutants reveals complex, additive and epistatic interactions underlying RPP1-like Ler locus activity. The effects of these mutations were measured on basal defense, global gene expression, primary metabolism, and disease resistance to a local Hyaloperonospora arabidopsidis isolate (Hpa Gw) collected from Gorzów (Gw), where the Landsberg accession originated. Gene expression sectors and metabolic hallmarks identified for HI are both dependent and independent of RPP1-like Ler members. We establish that mutations suppressing immune-related Ler/Kas2 HI do not compromise resistance to Hpa Gw. QTL mapping analysis of Hpa Gw resistance point to RPP7 as the causal locus. This work provides insight into the complex genetic architecture of the RPP1-like Ler locus and immune-related HI in Arabidopsis and into the contributions of RPP1-like genes to HI and defense.

Two genes encoding GH10 xylanases are essential for the virulence of the oomycete plant pathogen Phytophthora parasitica.

Plant cell walls are pivotal battlegrounds between microbial pathogens and their hosts. To penetrate the cell wall and thereby to facilitate infection, microbial pathogens are equipped with a wide array of cell wall-degrading enzymes to depolymerize the polysaccharides in the cell wall. However, many of these enzymes and their role in the pathogenesis of microbial pathogens are not characterized, especially those from Oomycetes. In this study, we analyzed the function of four putative endo-beta-1,4-xylanase-encoding genes (ppxyn1-ppxyn4) from Phytophthora parasitica, an oomycete plant pathogen known to cause severe disease in a wide variety of plant species. All four genes belong to the glycoside hydrolase family 10 (GH10). Recombinant proteins of ppxyn1, ppxyn2, and ppxyn4 obtained from the yeast Pichia pastoris showed degrading activities toward birch wood xylan, but they behaved differently in terms of the conditions for optimal activity, thermostability, and durability. Quantitative RT-PCR revealed upregulated expression of all four genes, especially ppxyn1 and ppxyn2, during plant infection. In contrast, ppxyn3 was highly expressed in cysts and its close homolog, ppxyn4, in germinating cysts. To uncover the role of ppxyn1 and ppxyn2 in the pathogenesis of P. parasitica, we generated silencing transformants for these two genes by double-stranded RNA-mediated gene silencing. Silencing ppxyn1 and ppxyn2 reduced the virulence of P. parasitica toward tobacco (Nicotiana benthamiana) and tomato plants. These results demonstrate the crucial role of xylanase-encoding ppxyn1 and ppxyn2 in the infection process of P. parasitica.

Conserved RxLR Effectors From Oomycetes Hyaloperonospora arabidopsidis and Phytophthora sojae Suppress PAMP- and Effector-Triggered Immunity in Diverse Plants.

Effector proteins are exported to the interior of host cells by diverse plant pathogens. Many oomycete pathogens maintain large families of candidate effector genes, encoding proteins with a secretory leader followed by an RxLR motif. Although most of these genes are very divergent between oomycete species, several genes are conserved between Phytophthora species and Hyaloperonospora arabidopsidis, suggesting that they play important roles in pathogenicity. We describe a pair of conserved effector candidates, HaRxL23 and PsAvh73, from H. arabidopsidis and P. sojae respectively. We show that HaRxL23 is expressed early during infection of Arabidopsis. HaRxL23 triggers an ecotype-specific defense response in Arabidopsis, suggesting that it is recognized by a host surveillance protein. HaRxL23 and PsAvh73 can suppress pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) in Nicotiana benthamiana and effector-triggered immunity (ETI) in soybean. Transgenic Arabidopsis constitutively expressing HaRxL23 or PsAvh73 exhibit suppression of PTI and enhancement of bacterial and oomycete virulence. Together, our experiments demonstrate that these conserved oomycete RxLR effectors suppress PTI and ETI across diverse plant species.

Immunodetection of fungal and oomycete pathogens: established and emerging threats to human health, animal welfare and global food security.

Filamentous fungi (moulds), yeast-like fungi, and oomycetes cause life-threatening infections of humans and animals and are a major constraint to global food security, constituting a significant economic burden to both agriculture and medicine. As well as causing localized or systemic infections, certain species are potent producers of allergens and toxins that exacerbate respiratory diseases or cause cancer and organ damage. We review the pathogenic and toxigenic organisms that are etiologic agents of both animal and plant diseases or that have recently emerged as serious pathogens of immunocompromised individuals. The use of hybridoma and phage display technologies and their success in generating monoclonal antibodies for the detection and control of fungal and oomycete pathogens are explored. Monoclonal antibodies hold enormous potential for the development of rapid and specific tests for the diagnosis of human mycoses, however, unlike plant pathology, their use in medical mycology remains to be fully exploited.

Fungal endopolygalacturonases are recognized as microbe-associated molecular patterns by the arabidopsis receptor-like protein RESPONSIVENESS TO BOTRYTIS POLYGALACTURONASES1.

Plants perceive microbial invaders using pattern recognition receptors that recognize microbe-associated molecular patterns. In this study, we identified RESPONSIVENESS TO BOTRYTIS POLYGALACTURONASES1 (RBPG1), an Arabidopsis (Arabidopsis thaliana) leucine-rich repeat receptor-like protein, AtRLP42, that recognizes fungal endopolygalacturonases (PGs) and acts as a novel microbe-associated molecular pattern receptor. RBPG1 recognizes several PGs from the plant pathogen Botrytis cinerea as well as one from the saprotroph Aspergillus niger. Infiltration of B. cinerea PGs into Arabidopsis accession Columbia induced a necrotic response, whereas accession Brno (Br-0) showed no symptoms. A map-based cloning strategy, combined with comparative and functional genomics, led to the identification of the Columbia RBPG1 gene and showed that this gene is essential for the responsiveness of Arabidopsis to the PGs. Transformation of RBPG1 into accession Br-0 resulted in a gain of PG responsiveness. Transgenic Br-0 plants expressing RBPG1 were equally susceptible as the recipient Br-0 to the necrotroph B. cinerea and to the biotroph Hyaloperonospora arabidopsidis. Pretreating leaves of the transgenic plants with a PG resulted in increased resistance to H. arabidopsidis. Coimmunoprecipitation experiments demonstrated that RBPG1 and PG form a complex in Nicotiana benthamiana, which also involves the Arabidopsis leucine-rich repeat receptor-like protein SOBIR1 (for SUPPRESSOR OF BIR1). sobir1 mutant plants did not induce necrosis in response to PGs and were compromised in PG-induced resistance to H. arabidopsidis.

Pathogenesis-related protein 4b interacts with leucine-rich repeat protein 1 to suppress PR4b-triggered cell death and defense response in pepper.

To control defense and cell-death signaling, plants contain an abundance of pathogen recognition receptors such as leucine-rich repeat (LRR) proteins. Here we show that pepper (Capsicum annuum) LRR1 interacts with the pepper pathogenesis-related (PR) protein 4b, PR4b, in yeast and in planta. PR4b is synthesized in the endoplasmic reticulum, interacts with LRR1 in the plasma membrane, and is secreted to the apoplast via the plasma membrane. Binding of PR4b to LRR1 requires the chitin-binding domain of PR4b. Purified PR4b protein inhibits spore germination and mycelial growth of plant fungal pathogens. Transient expression of PR4b triggers hypersensitive cell death. This cell death is compromised by co-expression of LRR1 as a negative regulator in Nicotiana benthamiana leaves. LRR1/PR4b silencing in pepper and PR4b over-expression in Arabidopsis thaliana demonstrated that LRR1 and PR4b are necessary for defense responses to Pseudomonas syringae pv. tomato and Hyaloperonospora arabidopsidis (Hpa) infection. The mutant of the PR4b Arabidopsis ortholog, pr4, showed enhanced susceptibility to Hpa infection. Together, our results suggest that PR4b functions as a positive modulator of plant cell death and defense responses. However, the activity of PR4b is suppressed by interaction with LRR1.