Phytophthora infestans RXLR effector Pi23014 targets host RNA-binding protein NbRBP3a to suppress plant immunity.

Phytophthora infestans is a destructive oomycete that causes the late blight of potato and tomato worldwide. It secretes numerous small proteins called effectors in order to manipulate host cell components and suppress plant immunity. Identifying the targets of these effectors is crucial for understanding P. infestans pathogenesis and host plant immunity. In this study, we show that the virulence RXLR effector Pi23014 of P. infestans targets the host nucleus and chloroplasts. By using a liquid chromatogrpahy-tandem mass spectrometry assay and co-immunoprecipitation assasys, we show that it interacts with NbRBP3a, a putative glycine-rich RNA-binding protein. We confirmed the co-localization of Pi23014 and NbRBP3a within the nucleus, by using bimolecular fluorescence complementation. Reverse transcription-quantitative PCR assays showed that the expression of NbRBP3a was induced in Nicotiana benthamiana during P. infestans infection and the expression of marker genes for multiple defence pathways were significantly down-regulated in NbRBP3-silenced plants compared with GFP-silenced plants. Agrobacterium tumefaciens-mediated transient overexpression of NbRBP3a significantly enhanced plant resistance to P. infestans. Mutations in the N-terminus RNA recognition motif (RRM) of NbRBP3a abolished its interaction with Pi23014 and eliminated its capability to enhance plant resistance to leaf colonization by P. infestans. We further showed that silencing NbRBP3 reduced photosystem II activity, reduced host photosynthetic efficiency, attenuated Pi23014-mediated suppression of cell death triggered by P. infestans pathogen-associated molecular pattern elicitor INF1, and suppressed plant immunity.

G-U base-paired hpRNA confers potent inhibition of small RNA function in plants.

MicroRNA (miRNA) target mimicry technologies, utilizing naturally occurring miRNA decoy molecules, represent a potent tool for analyzing miRNA function. In this study, we present a highly efficient small RNA (sRNA) target mimicry design based on G-U base-paired hairpin RNA (hpG:U), which allows for the simultaneous targeting of multiple sRNAs. The hpG:U constructs consistently generate high amounts of intact, polyadenylated stem-loop (SL) RNA outside the nuclei, in contrast to traditional hairpin RNA designs with canonical base pairing (hpWT), which were predominantly processed resulting in a loop. By incorporating a 460-bp G-U base-paired double-stranded stem and a 312-576 nt loop carrying multiple miRNA target mimicry sites (GUMIC), the hpG:U construct displayed effective repression of three Arabidopsis miRNAs, namely miR165/166, miR157, and miR160, both individually and in combination. Additionally, a GUMIC construct targeting a prominent cluster of siRNAs derived from cucumber mosaic virus (CMV) Y-satellite RNA (Y-Sat) effectively inhibited Y-Sat siRNA-directed silencing of the chlorophyll biosynthetic gene CHLI, thereby reducing the yellowing symptoms in infected Nicotiana plants. Therefore, the G-U base-paired hpRNA, characterized by differential processing compared to traditional hpRNA, acts as an efficient decoy for both miRNAs and siRNAs. This technology holds great potential for sRNA functional analysis and the management of sRNA-mediated diseases.

Specific detection of Phytophthora parasitica by recombinase polymerase amplification (RPA) assays based on a unique multi-copy genomic sequence.

Phytophthora parasitica is a highly destructive oomycete plant pathogen that is capable of infecting a wide range of hosts including many agricultural cash crops, fruit trees, and ornamental garden plants. One of the most important diseases caused by P. parasitica worldwide is black shank of tobacco. Rapid, sensitive, and specific pathogen detection is crucial for early rapid diagnosis which can facilitate effective disease management. In this study, we used a genomics approach to identify repeated sequences in the genome of P. parasitica by genome sequence alignment, and identified a 203 bp P. parasitica-specific sequence, PpM34, that is present in 31-60 copies in the genome. The P. parasitica genome-specificity of PpM34 was supported by PCR amplification of 24 genetically diverse strains of P. parasitica, 32 strains representing twelve other Phytophthora species, one Pythium specie, six fungal species and three bacterial species, all of which are plant pathogens. Our PCR and real-time PCR assays showed that the PpM34 sequence was highly sensitive in specifically detecting P. parasitica. Finally, we developed a PpM34-based high-efficiency Recombinase Polymerase Amplification (RPA) assay, which allowed us to specifically detect as little as 1 pg of P. parasitica total DNA from both pure cultures and infected Nicotiana benthamiana at 39°C using a fluorometric thermal cycler. The sensitivity, specificity, convenience and rapidity of this assay represents a major improvement for early diagnosis of P. parasitica infection.

Symbiont-host interactome mapping reveals effector-targeted modulation of hormone networks and activation of growth promotion.

Plants have benefited from interactions with symbionts for coping with challenging environments since the colonisation of land. The mechanisms of symbiont-mediated beneficial effects and similarities and differences to pathogen strategies are mostly unknown. Here, we use 106 (effector-) proteins, secreted by the symbiont Serendipita indica (Si) to modulate host physiology, to map interactions with Arabidopsis thaliana host proteins. Using integrative network analysis, we show significant convergence on target-proteins shared with pathogens and exclusive targeting of Arabidopsis proteins in the phytohormone signalling network. Functional in planta screening and phenotyping of Si effectors and interacting proteins reveals previously unknown hormone functions of Arabidopsis proteins and direct beneficial activities mediated by effectors in Arabidopsis. Thus, symbionts and pathogens target a shared molecular microbe-host interface. At the same time Si effectors specifically target the plant hormone network and constitute a powerful resource for elucidating the signalling network function and boosting plant productivity.

A Phytophthora infestans RXLR effector targets a potato ubiquitin-like domain-containing protein to inhibit the proteasome activity and hamper plant immunity.

Ubiquitin-like domain-containing proteins (UDPs) are involved in the ubiquitin-proteasome system because of their ability to interact with the 26S proteasome. Here, we identified potato StUDP as a target of the Phytophthora infestans RXLR effector Pi06432 (PITG_06432), which supresses the salicylic acid (SA)-related immune pathway. By overexpressing and silencing of StUDP in potato, we show that StUDP negatively regulates plant immunity against P. infestans. StUDP interacts with, and destabilizes, the 26S proteasome subunit that is referred to as REGULATORY PARTICLE TRIPLE-A ATP-ASE (RPT) subunit StRPT3b. This destabilization represses the proteasome activity. Proteomic analysis and Western blotting show that StUDP decreases the stability of the master transcription factor SYSTEMIC ACQUIRED RESISTANCE DEFICIENT 1 (SARD1) in SA biosynthesis. StUDP negatively regulates the SA signalling pathway by repressing the proteasome activity and destabilizing StSARD1, leading to a decreased expression of the SARD1-targeted gene ISOCHORISMATE SYNTHASE 1 and thereby a decrease in SA content. Pi06432 stabilizes StUDP, and it depends on StUDP to destabilize StRPT3b and thereby supress the proteasome activity. Our study reveals that the P. infestans effector Pi06432 targets StUDP to hamper the homeostasis of the proteasome by the degradation of the proteasome subunit StRPT3b and thereby suppresses SA-related immunity.

Potato protein tyrosine phosphatase StPTP1a is activated by StMKK1 to negatively regulate plant immunity.

Phytophthora infestans causes severe losses in potato production. The MAPK kinase StMKK1 was previously found to negatively regulate potato immunity to P. infestans. Our results showed that StMKK1 interacts with a protein tyrosine phosphatase, referred to as StPTP1a, and StMKK1 directly phosphorylates StPTP1a at residues Ser-99, Tyr-223 and Thr-290. StPTP1a is a functional phosphatase and the phosphorylation of StPTP1a at these three residues enhances its stability and catalytic activity. StPTP1a negatively regulates potato immunity and represses SA-related gene expression. Furthermore, StPTP1a interacts with, and dephosphorylates, the StMKK1 downstream signalling targets StMPK4 and -7 at their Tyr-203 residue resulting in the repression of salicylic acid (SA)-related immunity. Silencing of NbPTP1a + NbMPK4 or NbPTP1a + NbMPK7 abolished the plant immunity to P. infestans caused by NbPTP1a silencing, indicating that PTP1a functions upstream of NbMPK4 and NbMPK7. StMKK1 requires StPTP1a to negatively regulate SA-related immunity and StPTP1a is phosphorylated and stabilized during immune activation to promote the de-phosphorylation of StMPK4 and -7. Our results reveal that potato StMKK1 activates and stabilizes the tyrosine phosphatase StPTP1a that in its turn de-phosphorylates StMPK4 and -7, thereby repressing plant SA-related immunity.

miR398b and AtC2GnT form a negative feedback loop to regulate Arabidopsis thaliana resistance against Phytophthora parasitica.

Oomycetes are diploid eukaryotic microorganisms that seriously threaten sustainable crop production. MicroRNAs (miRNAs) and corresponding natural antisense transcripts (NATs) are important regulators of multiple biological processes. However, little is known about their roles in plant immunity against oomycete pathogens. In this study, we report the identification and functional characterization of miR398b and its cis-NAT, the core-2/I-branching beta-1,6-N-acetylglucosaminyltransferase gene (AtC2GnT), in plant immunity. Gain- and loss-of-function assays revealed that miR398b mediates Arabidopsis thaliana susceptibility to Phytophthora parasitica by targeting Cu/Zn-Superoxidase Dismutase1 (CSD1) and CSD2, leading to suppressed expression of CSD1 and CSD2 and decreased plant disease resistance. We further showed that AtC2GnT transcripts could inhibit the miR398b-CSDs module via inhibition of pri-miR398b expression, leading to elevated plant resistance to P. parasitica. Furthermore, quantitative reverse transcription PCR, RNA ligase-mediated 5'-amplification of cDNA ends (RLM-5' RACE), and transient expression assays indicated that miR398b suppresses the expression of AtC2GnT. We generated AtC2GnT-silenced A. thaliana plants by CRISPR/Cas9 or RNA interference methods, and the Nicotiana benthamiana NbC2GnT-silenced plants by virus-induced gene silencing. Pathogenicity assays showed that the C2GnT-silenced plants were more susceptible, while AtC2GnT-overexpressing plants exhibited elevated resistance to P. parasitica. AtC2GnT encodes a Golgi-localized protein, and transient expression of AtC2GnT enhanced N. benthamiana resistance to Phytophthora pathogens. Taken together, our results revealed a positive role of AtC2GnT and a negative regulatory loop formed by miR398b and AtC2GnT in regulating plant resistance to P. parasitica.

Mutations in PpAGO3 Lead to Enhanced Virulence of Phytophthora parasitica by Activation of 25-26 nt sRNA-Associated Effector Genes.

Oomycetes represent a unique group of plant pathogens that are destructive to a wide range of crops and natural ecosystems. Phytophthora species possess active small RNA (sRNA) silencing pathways, but little is known about the biological roles of sRNAs and associated factors in pathogenicity. Here we show that an AGO gene, PpAGO3, plays a major role in the regulation of effector genes hence the pathogenicity of Phytophthora parasitica. PpAGO3 was unique among five predicted AGO genes in P. parasitica, showing strong mycelium stage-specific expression. Using the CRISPR-Cas9 technology, we generated PpAGO3ΔRGG1-3 mutants that carried a deletion of 1, 2, or 3 copies of the N-terminal RGG motif (QRGGYD) but failed to obtain complete knockout mutants, which suggests its vital role in P. parasitica. These mutants showed increased pathogenicity on both Nicotiana benthamiana and Arabidopsis thaliana plants. Transcriptome and sRNA sequencing of PpAGO3ΔRGG1 and PpAGO3ΔRGG3 showed that these mutants were differentially accumulated with 25-26 nt sRNAs associated with 70 predicted cytoplasmic effector genes compared to the wild-type, of which 13 exhibited inverse correlation between gene expression and 25-26 nt sRNA accumulation. Transient overexpression of the upregulated RXLR effector genes, PPTG_01869 and PPTG_15425 identified in the mutants PpAGO3ΔRGG1 and PpAGO3ΔRGG3 , strongly enhanced N. benthamiana susceptibility to P. parasitica. Our results suggest that PpAGO3 functions together with 25-26 nt sRNAs to confer dynamic expression regulation of effector genes in P. parasitica, thereby contributing to infection and pathogenicity of the pathogen.

Activation of the VQ Motif-Containing Protein Gene VQ28 Compromised Nonhost Resistance of Arabidopsis thaliana to Phytophthora Pathogens.

Nonhost resistance refers to resistance of a plant species to all genetic variants of a non-adapted pathogen. Such resistance has the potential to become broad-spectrum and durable crop disease resistance. We previously employed Arabidopsis thaliana and a forward genetics approach to identify plant mutants susceptible to the nonhost pathogen Phytophthora sojae, which resulted in identification of the T-DNA insertion mutant esp1 (enhanced susceptibility to Phytophthora). In this study, we report the identification of VQ motif-containing protein 28 (VQ28), whose expression was highly up-regulated in the mutant esp1. Stable transgenic A. thaliana plants constitutively overexpressing VQ28 compromised nonhost resistance (NHR) against P. sojae and P. infestans, and supported increased infection of P. parasitica. Transcriptomic analysis showed that overexpression of VQ28 resulted in six differentially expressed genes (DEGs) that are involved in the response to abscisic acid (ABA). High performance liquid chromatography-mass spectrometry (HPLC-MS) detection showed that the contents of endogenous ABA, salicylic acid (SA), and jasmonate (JA) were enriched in VQ28 overexpression lines. These findings suggest that overexpression of VQ28 may lead to an imbalance in plant hormone homeostasis. Furthermore, transient overexpression of VQ28 in Nicotiana benthamiana rendered plants more susceptible to Phytophthora pathogens. Deletion mutant analysis showed that the C-terminus and VQ-motif were essential for plant susceptibility. Taken together, our results suggest that VQ28 negatively regulates plant NHR to Phytophthora pathogens.

A mitochondrial RNA processing protein mediates plant immunity to a broad spectrum of pathogens by modulating the mitochondrial oxidative burst.

Mitochondrial function depends on the RNA processing of mitochondrial gene transcripts by nucleus-encoded proteins. This posttranscriptional processing involves the large group of nuclear-encoded pentatricopeptide repeat (PPR) proteins. Mitochondrial processes represent a crucial part in animal immunity, but whether mitochondria play similar roles in plants remains unclear. Here, we report the identification of RESISTANCE TO PHYTOPHTHORA PARASITICA 7 (AtRTP7), a P-type PPR protein, in Arabidopsis thaliana and its conserved function in immunity to diverse pathogens across distantly related plant species. RTP7 affects the levels of mitochondrial reactive oxygen species (mROS) by participating in RNA splicing of nad7, which encodes a critical subunit of the mitochondrial respiratory chain Complex I, the largest of the four major components of the mitochondrial oxidative phosphorylation system. The enhanced resistance of rtp7 plants to Phytophthora parasitica is dependent on an elevated mROS burst, but might be independent from the ROS burst associated with plasma membrane-localized NADPH oxidases. Our study reveals the immune function of RTP7 and the defective processing of Complex I subunits in rtp7 plants resulted in enhanced resistance to both biotrophic and necrotrophic pathogens without affecting overall plant development.

The Raf-like kinase Raf36 negatively regulates plant resistance against the oomycete pathogen Phytophthora parasitica by targeting MKK2.

Oomycetes represent a unique group of plant pathogens that are phylogenetically distant from true fungi and cause significant crop losses and environmental damage. Understanding of the genetic basis of host plant susceptibility facilitates the development of novel disease resistance strategies. In this study, we report the identification of an Arabidopsis thaliana T-DNA mutant with enhanced resistance to Phytophthora parasitica with an insertion in the Raf-like mitogen-activated protein kinase kinase kinase gene Raf36. We generated additional raf36 mutants by CRISPR/Cas9 technology as well as Raf36 complementation and overexpression transformants, with consistent results of infection assays showing that Raf36 mediates Arabidopsis susceptibility to P. parasitica. Using a virus-induced gene silencing assay, we silenced Raf36 homologous genes in Nicotiana benthamiana and demonstrated by infection assays the conserved immune function of Raf36. Mutagenesis analyses indicated that the kinase activity of Raf36 is important for its immune function and interaction with MKK2, a MAPK kinase. By generating and analysing mkk2 mutants and MKK2 complementation and overexpression transformants, we found that MKK2 is a positive immune regulator in the response to P. parasitica infection. Furthermore, infection assay on mkk2 raf36 double mutant plants indicated that MKK2 is required for the raf36-conferred resistance to P. parasitica. Taken together, we identified a Raf-like kinase Raf36 as a novel plant susceptibility factor that functions upstream of MKK2 and directly targets it to negatively regulate plant resistance to P. parasitica.

Quantitative Analysis of RNA Editing at Specific Sites in Plant Mitochondria or Chloroplasts Using DNA Sequencing.

Cytidine-to-uridine (C-to-U) RNA editing is one of the most important post-transcriptional RNA processing in plant mitochondria and chloroplasts. Several techniques have been developed to detect the RNA editing efficiency in plant mitochondria and chloroplasts, such as poisoned primer extension (PPE) assays, high-resolution melting (HRM) analysis, and DNA sequencing. Here, we describe a method for the quantitative detection of RNA editing at specific sites by sequencing cDNA from plant leaves to further evaluate the effect of different treatments or plant mutants on the C to U RNA editing in mitochondria and chloroplasts.

Perturbations in nitric oxide homeostasis promote Arabidopsis disease susceptibility towards Phytophthora parasitica.

Phytophthora species can infect hundreds of different plants, including many important crops, causing a number of agriculturally relevant diseases. A key feature of attempted pathogen infection is the rapid production of the redox active molecule nitric oxide (NO). However, the potential role(s) of NO in plant resistance against Phytophthora is relatively unexplored. Here we show that the level of NO accumulation is crucial for basal resistance in Arabidopsis against Phytophthora parasitica. Counterintuitively, both relatively low or relatively high NO accumulation leads to reduced resistance against P. parasitica. S-nitrosylation, the addition of a NO group to a protein cysteine thiol to form an S-nitrosothiol, is an important route for NO bioactivity and this process is regulated predominantly by S-nitrosoglutathione reductase 1 (GSNOR1). Loss-of-function mutations in GSNOR1 disable both salicylic acid accumulation and associated signalling, and also the production of reactive oxygen species, leading to susceptibility towards P. parasitica. Significantly, we also demonstrate that secreted proteins from P. parasitica can inhibit Arabidopsis GSNOR1 activity.

Genome Sequence Data of Three Formae Speciales of Phytophthora vignae Causing Phytophthora Stem Rot on Different Vigna Species.

Phytophthora vignae is an important oomycete pathogen causing Phytophthora stem rot on some Vigna spp. Three P. vignae isolates obtained from mung bean, adzuki bean, and cowpea exhibited high similarities in morphology and physiology but are specialized to infect different hosts. Here, we report the first de novo assembly of the draft genomes of three P. vignae isolates, which were performed using the PacBio SMRT Sequel platform. This study will extend the genomic resource available for the Phytophthora genus and provide a good foundation for further research on comparative genomics of Phytophthora spp. and interaction mechanism between hosts and pathogens.

Potato StMPK7 is a downstream component of StMKK1 and promotes resistance to the oomycete pathogen Phytophthora infestans.

A cascade formed by phosphorylation events of mitogen-activated protein kinases (MAPKs) takes part in plant stress responses. However, the roles of these MAPKs in resistance of potato (Solanum tuberosum) against Phytophthora pathogens is not well studied. Our previous work showed that a Phytophthora infestans RXLR effector targets and stabilizes the negative regulator of MAPK kinase 1 of potato (StMKK1). Because in Arabidopsis thaliana the AtMPK4 is the downstream phosphorylation target of AtMKK1, we performed a phylogenetic analysis and found that potato StMPK4/6/7 are closely related and are orthologs of AtMPK4/5/11/12. Overexpression of StMPK4/7 enhances plant resistance to P. infestans and P. parasitica. Yeast two-hybrid analysis revealed that StMPK7 interacts with StMKK1, and StMPK7 is phosphorylated on flg22 treatment and by expressing constitutively active StMKK1 (CA-StMKK1), indicating that StMPK7 is a direct downstream signalling partner of StMKK1. Overexpression of StMPK7 in potato enhances potato resistance to P. infestans. Constitutively active StMPK7 (CA-StMPK7; StMPK7D198G, E202A ) was found to promote immunity to Phytophthora pathogens and to trigger host cell death when overexpressed in Nicotiana benthamiana leaves. Cell death triggered by CA-StMPK7 is SGT1/RAR1-dependent. Furthermore, cell death triggered by CA-StMPK7 is suppressed on coexpression with the salicylate hydroxylase NahG, and StMPK7 activation promotes salicylic acid (SA)-responsive gene expression. We conclude that potato StMPK7 is a downstream signalling component of the phosphorelay cascade involving StMKK1 and StMPK7 plays a role in immunity to Phytophthora pathogens via an SA-dependent signalling pathway.

Susceptibility factor RTP1 negatively regulates Phytophthora parasitica resistance via modulating UPR regulators bZIP60 and bZIP28.

The unfolded protein response (UPR) is a conserved stress adaptive signaling pathway in eukaryotic organisms activated by the accumulation of misfolded proteins in the endoplasmic reticulum (ER). UPR can be elicited in the course of plant defense, playing important roles in plant-microbe interactions. The major signaling pathways of plant UPR rely on the transcriptional activity of activated forms of ER membrane-associated stress sensors bZIP60 and bZIP28, which are transcription factors that modulate expression of UPR genes. In this study, we report the plant susceptibility factor Resistance to Phytophthora parasitica 1 (RTP1) is involved in ER stress sensing and rtp1-mediated resistance against P. parasitica is synergistically regulated with UPR, as demonstrated by the simultaneous strong induction of UPR and ER stress-associated immune genes in Arabidopsis thaliana rtp1 mutant plants during the infection by P. parasitica. We further demonstrate RTP1 contributes to stabilization of the ER membrane-associated bZIP60 and bZIP28 through manipulating the bifunctional protein kinase/ribonuclease IRE1-mediated bZIP60 splicing activity and interacting with bZIP28. Consequently, we find rtp1bzip60 and rtp1bzip28 mutant plants exhibit compromised resistance accompanied with attenuated induction of ER stress-responsive immune genes and reduction of callose deposition in response to P. parasitica infection. Taken together, we demonstrate RTP1 may exert negative modulating roles in the activation of key UPR regulators bZIP60 and bZIP28, which are required for rtp1-mediated plant resistance to P. parasitica. This facilitates our understanding of the important roles of stress adaptive UPR and ER stress in plant immunity.

Phytophthora infestans RXLR effector PITG20303 targets a potato MKK1 protein to suppress plant immunity.

Pathogens secret a plethora of effectors into the host cell to modulate plant immunity. Analysing the role of effectors in altering the function of their host target proteins will reveal critical components of the plant immune system. Here we show that Phytophthora infestans RXLR effector PITG20303, a virulent variant of AVRblb2 (PITG20300) that escapes recognition by the resistance protein Rpi-blb2, suppresses PAMP-triggered immunity (PTI) and promotes pathogen colonization by targeting and stabilizing a potato MAPK cascade protein, StMKK1. Both PITG20300 and PITG20303 target StMKK1, as confirmed by multiple in vivo and in vitro assays, and StMKK1 was shown to be a negative regulator of plant immunity, as determined by overexpression and gene silencing. StMKK1 is a negative regulator of plant PTI, and the kinase activities of StMKK1 are required for its suppression of PTI and effector interaction. PITG20303 depends partially on MKK1, PITG20300 does not depend on MKK1 for suppression of PTI-induced reactive oxygen species burst, while the full virulence activities of nuclear targeted PITG20303 and PITG20300 are dependent on MKK1. Our results show that PITG20303 and PITG20300 target and stabilize the plant MAPK cascade signalling protein StMKK1 to negatively regulate plant PTI response.

The novel peptide NbPPI1 identified from Nicotiana benthamiana triggers immune responses and enhances resistance against Phytophthora pathogens.

In plants, recognition of small secreted peptides, such as damage/danger-associated molecular patterns (DAMPs), regulates diverse processes, including stress and immune responses. Here, we identified an SGPS (Ser-Gly-Pro-Ser) motif-containing peptide, Nicotiana tabacum NtPROPPI, and its two homologs in Nicotiana benthamiana, NbPROPPI1 and NbPROPPI2. Phytophthora parasitica infection and salicylic acid (SA) treatment induced NbPROPPI1/2 expression. Moreover, SignalP predicted that the 89-amino acid NtPROPPI includes a 24-amino acid N-terminal signal peptide and NbPROPPI1/2-GFP fusion proteins were mainly localized to the periplasm. Transient expression of NbPROPPI1/2 inhibited P. parasitica colonization, and NbPROPPI1/2 knockdown rendered plants more susceptible to P. parasitica. An eight-amino-acid segment in the NbPROPPI1 C-terminus was essential for its immune function and a synthetic 20-residue peptide, NbPPI1, derived from the C-terminus of NbPROPPI1 provoked significant immune responses in N. benthamiana. These responses led to enhanced accumulation of reactive oxygen species, activation of mitogen-activated protein kinases, and up-regulation of the defense genes Flg22-induced receptor-like kinase (FRK) and WRKY DNA-binding protein 33 (WRKY33). The NbPPI1-induced defense responses require Brassinosteroid insensitive 1-associated receptor kinase 1 (BAK1). These results suggest that NbPPI1 functions as a DAMP in N. benthamiana; this novel DAMP provides a potentially useful target for improving plant resistance to Pytophthora pathogens.

A Novel Disease of Mung Bean, Phytophthora Stem Rot Caused by a New Forma Specialis of Phytophthora vignae.

An emerging soilborne disease resembling Phytophthora stem rot was observed on mung bean plants grown in Anhui, China. To identify the causal agent, diseased plants and soil samples from 13 fields were collected to isolate the pathogen. Twenty-two Phytophthora isolates were recovered from the samples and detailed identification was conducted. Based on morphological and molecular characterizations, all of the isolates were consistently identified as P. vignae. Phylogenetic analysis using eight nuclear loci sequences of the internal transcribed spacer region, rRNA gene large subunit, a partial sequence of the ß-tubulin gene, translation elongation factor 1α, 60S ribosomal protein L10, the enolase gene, heat shock protein 90, and triose phosphate isomerase/glyceraldehyde-3-phosphate dehydrogenase and a mitochondrial locus cytochrome c oxidase subunit I revealed that the mung bean isolates grouped in the same clade as P. vignae and its two formae speciales, P. vignae f. sp. adzukicola and P. vignae f. sp. vignae. A host specificity test showed that the mung bean isolates of P. vignae were pathogenic toward mung bean with a much stronger virulence and toward adzuki bean with a relatively weak virulence, but they were nonpathogenic to the other tested legume crops, including soybean, cowpea, pea, common bean, faba bean, and chickpea. The host range of mung bean isolates significantly differs from those of P. vignae f. sp. adzukicola and P. vignae f. sp. vignae based on our results and on previous studies. Thus, the pathogen causing Phytophthora stem rot of mung bean is proposed as a new forma specialis of P. vignae, designated as P. vignae f. sp. mungcola.