A conserved protein family in mirid bug Riptortus pedestris plays dual roles in regulating plant immunity.

The mirid bug (Riptortus pedestris), a major soybean pest, migrates into soybean fields during the pod filling stage and causes staygreen syndrome, which leads to substantial yield losses. The mechanism by which R. pedestris elicits soybean (Glycine max) defenses and counter-defenses remains largely unexplored. In this study, we characterized a protein family from R. pedestris, designated Riptortus pedestris HAMP 1 (RPH1) and its putative paralogs (RPH1L1, 2, 3, 4, and 5), whose members exhibit dual roles in triggering and inhibiting plant immunity. RPH1 and RPH1L1 function as herbivore-associated molecular patterns (HAMPs), activating pattern-triggered immunity (PTI) in tobacco (Nicotiana benthamiana) and G. max. Furthermore, RPH1 stimulates jasmonic acid and ethylene biosynthesis in G. max, thereby enhancing its resistance to R. pedestris feeding. Additionally, RPH1 homologs are universally conserved across various herbivorous species, with many homologs also acting as HAMPs that trigger plant immunity. Interestingly, the remaining RPH1 putative paralogs (RPH1L2-5) serve as effectors that counteract RPH1-induced PTI, likely by disrupting the extracellular perception of RPH1. This research uncovers a HAMP whose homologs are conserved in both chewing and piercing-sucking insects. Moreover, it unveils an extracellular evasion mechanism utilized by herbivores to circumvent plant immunity using functionally differentiated paralogs.

Profiling of phenolic composition in camellia oil and its correlative antioxidant properties analysis.

Less research has been conducted on the association between camellia oil's (CO) phenolic composition and antioxidant capability. In this study, the phenolic profile of CO and its connection to antioxidant capacity were examined utilizing a combination of widely-targeted phenolic metabolomics and multivariate statistical analysis. A total of 751 phenolics were discovered. The WGCNA was used to link phenols to antioxidants, yielding 161 antioxidant-related phenols from the blue module. In response to several antioxidant assays, 59 (FRAP), 59 (DPPH), and 53 (ABTS) phenolics were identified as differential phenolic markers (DPMs). Further stepwise multiple linear regression revealed six DPMs that substantially influenced the antioxidant capacities. Nine metabolic pathways and their associated network mechanisms for the most significant phenolics were developed. This study sheds light on the phenolic content of CO, elucidates their role in antioxidant activity, and lays the groundwork for improving extraction techniques and generating improved product.

Maturation-induced changes in phenolic forms and their antioxidant activities of walnuts: A dual view from kernel and pellicle.

The phenolic profiles and antioxidant activities during walnut maturation are not well understood. This study used UPLC-MS/MS to evaluate phenolic content in walnuts, including free, esterified, and bound forms, at different maturation stages. Findings showed that free phenolics were predominant, comprising 44.57 % in kernels and 56.54 % in pellicles. In vitro assays showed antioxidant capacity decreased with maturation, with IC50 values of 0.87-84.43 µg/mL in pellicles and 48.51-712.30 µg/mL in kernels. Most monomeric phenols decreased in concentration as the fruit ripened. OPLS-DA identified 5 and 8 maturity-sensitive phenolics (MSPs) in kernels and pellicles, respectively, with fold changes from 2.32 to 1664.72. Pearson correlation analysis showed a significant correlation between MSPs and antioxidant activity (r > 0.75, p < 0.05). Bioinformatics analysis elucidated three key metabolic pathways involved in these changes. This research provides insights into walnut phenolic composition, important for optimizing harvest practices and enhancing nutritional value.

The Pythium periplocum elicitin PpEli2 confers broad-spectrum disease resistance by triggering a novel receptor-dependent immune pathway in plants.

Elicitins are microbe-associated molecular patterns produced by oomycetes to elicit plant defense. It is still unclear whether elicitins derived from non-pathogenic oomycetes can be used as bioactive molecules for disease control. Here, for the first time we identify and characterize an elicitin named PpEli2 from the soil-borne oomycete Pythium periplocum, which is a non-pathogenic mycoparasite colonizing the root ecosystem of diverse plant species. Perceived by a novel cell surface receptor-like protein, REli, that is conserved in various plants (e.g. tomato, pepper, soybean), PpEli2 can induce hypersensitive response cell death and an immunity response in Nicotiana benthamiana. Meanwhile, PpEli2 enhances the interaction between REli and its co-receptor BAK1. The receptor-dependent immune response triggered by PpEli2 is able to protect various plant species against Phytophthora and fungal infections. Collectively, our work reveals the potential agricultural application of non-pathogenic elicitins and their receptors in conferring broad-spectrum resistance for plant protection.

A Phytophthora receptor-like kinase regulates oospore development and can activate pattern-triggered plant immunity.

Plant cell-surface leucine-rich repeat receptor-like kinases (LRR-RLKs) and receptor-like proteins (LRR-RLPs) form dynamic complexes to receive a variety of extracellular signals. LRR-RLKs are also widespread in oomycete pathogens, whereas it remains enigmatic whether plant and oomycete LRR-RLKs could mediate cell-to-cell communications between pathogen and host. Here, we report that an LRR-RLK from the soybean root and stem rot pathogen Phytophthora sojae, PsRLK6, can activate typical pattern-triggered immunity in host soybean and nonhost tomato and Nicotiana benthamiana plants. PsRLK6 homologs are conserved in oomycetes and also exhibit immunity-inducing activity. A small region (LRR5-6) in the extracellular domain of PsRLK6 is sufficient to activate BAK1- and SOBIR1-dependent immune responses, suggesting that PsRLK6 is likely recognized by a plant LRR-RLP. Moreover, PsRLK6 is shown to be up-regulated during oospore maturation and essential for the oospore development of P. sojae. Our data provide a novel type of microbe-associated molecular pattern that functions in the sexual reproduction of oomycete, and a scenario in which a pathogen LRR-RLK could be sensed by a plant LRR-RLP to mount plant immunity.

Quorum quenching by a type IVA secretion system effector.

Proteobacteria primarily utilize acyl-homoserine lactones (AHLs) as quorum-sensing signals for intra-/interspecies communication to control pathogen infections. Enzymatic degradation of AHL represents the major quorum-quenching mechanism that has been developed as a promising approach to prevent bacterial infections. Here we identified a novel quorum-quenching mechanism revealed by an effector of the type IVA secretion system (T4ASS) in bacterial interspecies competition. We found that the soil antifungal bacterium Lysobacter enzymogenes OH11 (OH11) could use T4ASS to deliver the effector protein Le1288 into the cytoplasm of another soil microbiome bacterium Pseudomonas fluorescens 2P24 (2P24). Le1288 did not degrade AHL, whereas its delivery to strain 2P24 significantly impaired AHL production through binding to the AHL synthase PcoI. Therefore, we defined Le1288 as LqqE1 (Lysobacter quorum-quenching effector 1). Formation of the LqqE1-PcoI complex enabled LqqE1 to block the ability of PcoI to recognize/bind S-adenosy-L-methionine, a substrate required for AHL synthesis. This LqqE1-triggered interspecies quorum-quenching in bacteria seemed to be of key ecological significance, as it conferred strain OH11 a better competitive advantage in killing strain 2P24 via cell-to-cell contact. This novel quorum-quenching also appeared to be adopted by other T4ASS-production bacteria. Our findings suggest a novel quorum-quenching that occurred naturally in bacterial interspecies interactions within the soil microbiome by effector translocation. Finally, we presented two case studies showing the application potential of LqqE1 to block AHL signaling in the human pathogen Pseudomonas aeruginosa and the plant pathogen Ralstonia solanacearum.

Plant secondary metabolite citral interferes with Phytophthora capsici virulence by manipulating the expression of effector genes.

Phytophthora capsici is a notorious pathogen that infects various economically important plants and causes serious threats to agriculture worldwide. Plants deploy a variety of plant secondary metabolites to fend off pathogen attacks, but the molecular mechanisms are largely unknown. In this study, we screened 11 plant secondary metabolites to evaluate their biofumigation effects against P. capsici, and found that citral, carvacrol, and trans-2-decenal exhibited strong antimicrobial effects. Intriguingly, a low concentration of citral was effective in restricting P. capsici infection in Nicotiana benthamiana, but it was unable to inhibit the mycelial growth. A high concentration of citral affected the mycelial growth and morphology, zoospore germination, and cell membrane permeability of P. capsici. Further investigations showed that citral did not induce expression of tested plant immunity-related genes and reactive oxygen species (ROS) production, suggesting that a low concentration of citral could not trigger plant immunity. Moreover, RNA-Seq analysis showed that citral treatment regulated the expression of some P. capsici effector genes such as RxLR genes and P. cactorum-fragaria (PCF)/small cysteine-rich (SCR)74-like genes during the infection process, which was also verified by reverse transcription-quantitative PCR assay. Five candidate effector genes suppressed by citral significantly facilitated P. capsici infection in N. benthamiana or inhibited ROS triggered by flg22, suggesting that they were virulence factors of P. capsici. Together, our results revealed that plant-derived citral exhibited excellent inhibitory efficacy against P. capsici by suppressing vegetative growth and manipulating expression of effector genes, which provides a promising application of citral for controlling Phytophthora blight.

Identification of Key Antioxidants of Free, Esterified, and Bound Phenolics in Walnut Kernel and Skin.

Walnut is a natural source of antioxidants. Its antioxidant capacity is determined by the distribution and composition of phenolics. The key phenolic antioxidants in various forms (free, esterified, and bound) in walnut kernel (particularly seed skin) are unknown. The phenolic compounds in twelve walnut cultivars were analyzed using ultra-performance liquid chromatography coupled with a triple quadrupole mass spectrometer in this study. A boosted regression tree analysis was used to identify the key antioxidants. Ellagic acid, gallic acid, catechin, ferulic acid, and epicatechin were abundant in the kernel and skin. The majority of phenolic acids were widely distributed in the free, esterified, and bound forms in the kernel but more concentrated in bound phenolics in the skin. The total phenolic levels of the three forms were positively correlated with antioxidant activities (R = 0.76-0.94, p < 0.05). Ellagic acid was the most important antioxidant in the kernel, accounting for more than 20%, 40%, and 15% of antioxidants, respectively. Caffeic acid was responsible for up to 25% of free phenolics and 40% of esterified phenolics in the skin. The differences in the antioxidant activity between the cultivars were explained by the total phenolics and key antioxidants. The identification of key antioxidants is critical for new walnut industrial applications and functional food design in food chemistry.

Lysobacter enzymogenes prevents Phytophthora infection by inhibiting pathogen growth and eliciting plant immune responses.

The Phytophthora pathogen causes enormous damage to important agricultural plants. This group of filamentous pathogens is phylogenetically distant from fungi, making them difficult to control by most chemical fungicides. Lysobacter enzymogenes OH11 (OH11) is a biocontrol bacterium that secretes HSAF (Heat-Stable Antifungal Factor) as a broad-spectrum antifungal weapon. Here, we showed that OH11 could also control a variety of plant Phytophthora diseases caused by three major oomycetes (P. sojae, P. capsici and P. infestans). We provided abundant evidence to prove that OH11 protected host plants from Phytophthora pathogen infection by inhibiting mycelial growth, digesting cysts, suppressing cyst germination, and eliciting plant immune responses. Interestingly, the former two processes required the presence of HSAF, while the latter two did not. This suggested that L. enzymogenes could prevent Phytophthora infection via multiple previously unknown mechanisms. Therefore, this study showed that L. enzymogenes could serve as a promising alternative resource for promoting plant resistance to multiple Phytophthora pathogens.

Level and risk assessment of selected polychlorinated biphenyls, polycyclic aromatic hydrocarbons, and organochlorine pesticides in walnut and soil.

It is unknown how hydrophobic organic contaminants (HOCs) are distributed and how they affect the environment in high-fat nuts and their planted soil. The profile of HOCs in walnut/soil system was investigated in this study. Polychlorinated biphenyls (PCBs), polycyclic aromatic hydrocarbons (PAHs), and organochlorine pesticides (OCPs) were found in walnuts at concentrations of 0.67, 127, and 116 µg/kg, respectively. The target hazard quotients (THQ) of 17 PCBs, 16 PAHs, and 21 OCPs from walnut consumption by human were 0.06, 0.01, and 0.11, respectively. The highest concentrations of HOC in the soil were found in Nap and toxaphene, with concentrations of 2580 and 902 µg/kg, respectively. Bioaccumulation factors (BAF) and biota-sediment accumulation factors (BSAF) in walnuts were ranged from <0.01 to 7.04 and <0.01 to 3.83, respectively. Concentrations of most individual HOCs in soil samples were significantly correlated with soil organic matter (SOM) (p < 0.01) and minerals (p < 0.01), with maximum correlation coefficients of 0.70 (OM-PCB81) and -0.84 (P-BaP). According to this study, high-fat walnuts do not have a high bioaccumulation of HOCs from soil, and the risk of consumption is within the safe range.

The Phytophthora sojae effector PsFYVE1 modulates immunity-related gene expression by targeting host RZ-1A protein.

Oomycete pathogens secrete numerous effectors to manipulate plant immunity and promote infection. However, relatively few effector types have been well characterized. In this study, members of an FYVE domain-containing protein family that are highly expanded in oomycetes were systematically identified, and one secreted protein, PsFYVE1, was selected for further study. PsFYVE1 enhanced Phytophthora capsici infection in Nicotiana benthamiana and was necessary for Phytophthora sojae virulence. The FYVE domain of PsFYVE1 had PI3P-binding activity that depended on four conserved amino acid residues. Furthermore, PsFYVE1 targeted RNA-binding proteins RZ-1A/1B/1C in N. benthamiana and soybean (Glycine max), and silencing of NbRZ-1A/1B/1C genes attenuated plant immunity. NbRZ-1A was associated with the spliceosome complex that included three important components, glycine-rich RNA-binding protein 7 (NbGRP7), glycine-rich RNA-binding protein 8 (NbGRP8), and a specific component of the U1 small nuclear ribonucleoprotein complex (NbU1-70K). Notably, PsFYVE1 disrupted NbRZ-1A-NbGRP7 interaction. RNA-seq and subsequent experimental analysis demonstrated that PsFYVE1 and NbRZ-1A not only modulated pre-mRNA alternative splicing (AS) of the necrotic spotted lesions 1 (NbNSL1) gene, but also co-regulated transcription of hydroxycinnamoyl-CoA shikimate/quinate hydroxycinnamoyl transferase (NbHCT), ethylene insensitive 2 (NbEIN2), and sucrose synthase 4 (NbSUS4) genes, which participate in plant immunity. Collectively, these findings indicate that the FYVE domain-containing protein family includes potential uncharacterized effector types and also highlight that plant pathogen effectors can regulate plant immunity-related genes at both AS and transcription levels to promote disease.

Cyclophilin effector Al106 of mirid bug Apolygus lucorum inhibits plant immunity and promotes insect feeding by targeting PUB33.

Apolygus lucorum (Meyer-Dur; Heteroptera: Miridae) is a major agricultural pest infesting crops, vegetables, and fruit trees. During feeding, A. lucorum secretes a plethora of effectors into its hosts to promote infestation. However, the molecular mechanisms of these effectors manipulating plant immunity are largely unknown. Here, we investigated the molecular mechanism underlying the effector Al106 manipulation of plant-insect interaction by RNA interference, electrical penetration graph, insect and pathogen bioassays, protein-protein interaction studies, and protein ubiquitination experiment. Expression of Al106 in Nicotiana benthamiana inhibits pathogen-associated molecular pattern-induced cell death and reactive oxygen species burst, and promotes insect feeding and plant pathogen infection. In addition, peptidyl-prolyl cis-trans isomerase (PPIase) activity of Al106 is required for its function to inhibit PTI.Al106 interacts with a plant U-box (PUB) protein, PUB33, from N. benthamiana and Arabidopsis thaliana. We also demonstrated that PUB33 is a positive regulator of plant immunity. Furthermore, an in vivo assay revealed that Al106 inhibits ubiquitination of NbPUB33 depending on PPIase activity. Our findings revealed that a novel cyclophilin effector may interact with plant PUB33 to suppress plant immunity and facilitate insect feeding in a PPIase activity-dependent manner.

Apolygus lucorum effector Al6 promotes insect feeding performance on soybean plants: RNAi analysis and feeding behaviour study with electrical penetration graph.

The mirid bug Apolygus lucorum, a dominant mirid species in northern China, is a notorious polyphagous pest with more than 200 hosts, including several major crops such as cotton and soybean, resulting in massive economic loss. Studies of insect salivary effectors may provide a novel control strategy for A. lucorum. An A. lucorum effector, that is, Al6, that inhibits plant immunity by using glutathione peroxidase to repress reactive oxidase accumulation was previously identified. In this study, we further explored the molecular functions of Al6 associated with feeding behaviour and insect survival on soybean, a major host of A. lucorum, using RNA interference and electrical penetration graph (EPG) techniques. We initially observed the injury symptom of this mirid bug and characterized feeding behaviour on soybean leaves using EPG. Our results revealed that A. lucorum preferred to feed on young plant organs such as tender leaves, shoots and buds. This mirid bug used cell rupture as a feeding strategy to ingest cell contents from plant tissues. Subsequently, we silenced the Al6 gene using RNAi and investigated the feeding behaviour, honeydew excretion, body weight, and survival rates of A. lucorum on soybean after Al6 knockdown. Our results demonstrated that silencing of Al6 significantly reduced feeding duration, amount of honeydew secretion, body weight, and survival rates of A. lucorum. Thus, our findings provide a novel molecular target of plant-mediated RNAi for the control of A. lucorum.

Novel EIicitin from Pythium oligandrum Confers Disease Resistance against Phytophthora capsici in Solanaceae Plants.

The mycoparasite Pythium oligandrum is a nonpathogenic oomycete that can boost plant immune responses. Elicitins are microbe-associated molecular patterns (MAMPs) specifically produced by oomycetes that activate plant defense. Here, we identified a novel elicitin, PoEli8, from P. oligandrum that exhibits immunity-inducing activity in plants. In vitro-purified PoEli8 induced strong innate immune responses and enhanced resistance to the oomycete pathogen Phytophthora capsici in Solanaceae plants, including Nicotiana benthamiana, tomato, and pepper. Cell death and reactive oxygen species (ROS) accumulation triggered by the PoEli8 protein were dependent on the plant coreceptors receptor-like kinases (RLKs) BAK1 and SOBIR1. Furthermore, REli from N. benthamiana, a cell surface receptor-like protein (RLP) was implicated in the perception of PoEli8 in N. benthamiana. These results indicate the potential value of PoEli8 as a bioactive formula to protect Solanaceae plants against Phytophthora.

A Phytophthora capsici RXLR effector manipulates plant immunity by targeting RAB proteins and disturbing the protein trafficking pathway.

The oomycete pathogen Phytophthora capsici encodes hundreds of RXLR effectors that enter the plant cells and suppress host immunity. Only a few of these genes are conserved across different strains and species. Such core effectors might target hub genes and immune pathways in hosts. Here, we describe the functional characterization of the core P. capsici RXLR effector RXLR242. The expression of RXLR242 was up-regulated during infection, and its ectopic expression in Nicotiana benthamiana, an experimental plant host, further promoted Phytophthora infection. RXLR242 physically interacted with a group of RAB proteins that belong to the small GTPase family and play a role in regulating transport pathways in the intracellular membrane trafficking system. In addition, RXLR242 impeded the secretion of PATHOGENESIS-RELATED 1 (PR1) protein to the apoplast. This phenomenon resulted from the competitive binding of RXLR242 to RABE1-7. We also found that RXLR242 interfered with the association between RABA4-3 and its binding protein, thereby disrupting the trafficking of the membrane receptor FLAGELLIN-SENSING 2. Thus, RXLR242 manipulates plant immunity by targeting RAB proteins and disrupting protein trafficking in the host plants.

Naringenin confers defence against Phytophthora nicotianae through antimicrobial activity and induction of pathogen resistance in tobacco.

Tobacco black shank caused by Phytophthora nicotianae is a serious disease in tobacco cultivation. We found that naringenin is a key factor that causes different sensitivity to P. nicotianae between resistant and susceptible tobacco. The level of basal flavonoids in resistant tobacco was distinct from that in susceptible tobacco. Of all flavonoids with different content, naringenin showed the best antimicrobial activity against mycelial growth and sporangia production of P. nicotianae in vitro. However, naringenin showed very low or no antimicrobial activity to other plant pathogens. We found that naringenin induced not only the accumulation of reactive oxygen species, but also the expression of salicylic acid biosynthesis-related genes. Naringenin induced the expression of the basal pathogen resistance gene PR1 and the SAR8.2 gene that contributes to plant resistance to P. nicotianae. We then interfered with the expression of the chalcone synthase (NtCHS) gene, the key gene of the naringenin synthesis pathway, to inhibit naringenin biosynthesis. NtCHS-RNAi rendered tobacco highly sensitive to P. nicotianae, but there was no change in susceptibility to another plant pathogen, Ralstonia solanacearum. Finally, exogenous application of naringenin on susceptible tobacco enhanced resistance to P. nicotianae and naringenin was very stable in this environment. Our findings revealed that naringenin plays a core role in the defence against P. nicotianae and expanded the possibilities for the application of plant secondary metabolites in the control of P. nicotianae.

Comprehensive lipidomics analysis reveals the changes in lipid profile of camellia oil affected by insect damage.

Information on changes in lipid composition of seed oils under biotic stresses is scare. The camellia weevil, Curculio chinensis (Coleoptera: Curculionidae) as a notorious seed predator of Camellia species, has caused huge economic losses in China. Lipidomics is used in this study to reveal the lipid composition of camellia oil and its changes after insect damage. 278 lipids including glycerolipids (GL) (221), glycerophospholipids (GP) (34), fatty acyls (FA) (13), sphingolipids (SP) (8), prenol lipids (PR) (1) and sterol lipids (ST) (1) were determined in camellia oils. Insect damage had a significant impact on lipids, particularly FA and GL. Ten significantly different lipids [FFA(18:2), FFA(24:6), TG(14:1/18:2/18:2), TG(16:0/23:0/18:2), TG(20:1/24:1/18:2), TG(18:2/24:0/18:2), TG(16:3/18:2/22:5), PI(16:1/18:1), PE(16:0/18:1), PE(18:1/18:2)] were identified as potential biomarkers for distinguishing oil extracted from non-infested oilseeds and oil from infested oilseeds. We also detected four most important metabolic pathways by bioinformatics analysis to explore the mechanisms underlying changes. Our findings may be useful for future camellia oil production and may provide new insight into improving of nutritional quality of camellia oil.

Genome of Pythium myriotylum Uncovers an Extensive Arsenal of Virulence-Related Genes among the Broad-Host-Range Necrotrophic Pythium Plant Pathogens.

The Pythium (Peronosporales, Oomycota) genus includes devastating plant pathogens that cause widespread diseases and severe crop losses. Here, we have uncovered a far greater arsenal of virulence factor-related genes in the necrotrophic Pythium myriotylum than in other Pythium plant pathogens. The genome of a plant-virulent P. myriotylum strain (~70 Mb and 19,878 genes) isolated from a diseased rhizome of ginger (Zingiber officinale) encodes the largest repertoire of putative effectors, proteases, and plant cell wall-degrading enzymes (PCWDEs) among the studied species. P. myriotylum has twice as many predicted secreted proteins than any other Pythium plant pathogen. Arrays of tandem duplications appear to be a key factor of the enrichment of the virulence factor-related genes in P. myriotylum. The transcriptomic analysis performed on two P. myriotylum isolates infecting ginger leaves showed that proteases were a major part of the upregulated genes along with PCWDEs, Nep1-like proteins (NLPs), and elicitin-like proteins. A subset of P. myriotylum NLPs were analyzed and found to have necrosis-inducing ability from agroinfiltration of tobacco (Nicotiana benthamiana) leaves. One of the heterologously produced infection-upregulated putative cutinases found in a tandem array showed esterase activity with preferences for longer-chain-length substrates and neutral to alkaline pH levels. Our results allow the development of science-based targets for the management of P. myriotylum-caused disease, as insights from the genome and transcriptome show that gene expansion of virulence factor-related genes play a bigger role in the plant parasitism of Pythium spp. than previously thought. IMPORTANCE Pythium species are oomycetes, an evolutionarily distinct group of filamentous fungus-like stramenopiles. The Pythium genus includes several pathogens of important crop species, e.g., the spice ginger. Analysis of our genome from the plant pathogen Pythium myriotylum uncovered a far larger arsenal of virulence factor-related genes than found in other Pythium plant pathogens, and these genes contribute to the infection of the plant host. The increase in the number of virulence factor-related genes appears to have occurred through the mechanism of tandem gene duplication events. Genes from particular virulence factor-related categories that were increased in number and switched on during infection of ginger leaves had their activities tested. These genes have toxic activities toward plant cells or activities to hydrolyze polymeric components of the plant. The research suggests targets to better manage diseases caused by P. myriotylum and prompts renewed attention to the genomics of Pythium plant pathogens.

Characterization of Salivary Secreted Proteins That Induce Cell Death From Riptortus pedestris (Fabricius) and Their Roles in Insect-Plant Interactions.

Riptortus pedestris (Fabricius) is a polyphagous hemipteran crop pest that mainly feeds on the leguminous plants, resulting in shriveled and dimpled seeds. With recent several outbreaks in the Huang-Huai-Hai region of China, as well as in South Korea and Japan, this species has caused enormous economic losses to soybean crops. In the present study, we found that R. pedestris feeding results in local lesions at the infestation sites. To identify the key effectors that induce plant damage during feeding, the salivary glands of R. pedestris were dissected for transcriptome sequencing, and 200 putative secreted proteins were transiently expressed in N. benthamiana. Among them, three intracellular effectors (RP191, RP246, and RP302) and one apoplastic effector (RP309) were identified as necrosis-inducing proteins (NIPs), which also triggered the reactive oxidative burst. Yeast signal sequence trap and qRT-PCR analysis suggested that these proteins might be secreted into plant tissue during R. pedestris infestation. Pathogenicity assays revealed that RP191, 246, and 302 promote Phytophthora capsici infection or induce Spodoptera litura feeding by inhibiting plant immunity. RP302 is localized to the cytoplasm and nuclei, while RP191 and 246 are endoplasmic reticulum (ER) resident proteins. RP309 stimulates the expression of PTI marker genes, and its induced cell death depends on co-receptors NbBAK1 and NbSOBIR1, indicating that it is a HAMP. Bioinformatics analysis demonstrated that four NIPs are recently evolved effectors and only conserved in the Pentatomidae. In this study, saliva-secreted proteins were used as the starting point to preliminarily analyze the harm mechanism of R. pedestris, which might provide a new idea and theoretical basis for this species control.

Phytophthora capsici CBM1-containing protein CBP3 is an apoplastic effector with plant immunity-inducing activity.

Carbohydrate-binding module family 1 (CBM1) is a cellulose-binding domain that is almost exclusively found in fungi and oomycetes. CBM1-containing proteins (CBPs) have diverse domain architectures and play pivotal roles in the plant-microbe interaction. However, only a few CBPs have been functionally investigated. In this study, we identified PcCBP3 in an oomycete pathogen, Phytophthora capsici. PcCBP3 contains two tandem CBM1 domains and its orthologs from other Phytophthora species exhibit diversity including gene loss, pseudogenization, variations in sequences, and domain structures. PcCBP3 is upregulated during infection and knockout of PcCBP3 results in significantly decreased virulence. Moreover, PcCBP3 requires signal peptide to induce BAK1-dependent cell death in Nicotiana benthamiana. Further studies indicate that PcCBP3-triggered cell death and plant immunity require its N-terminal region, which is conserved in CBM1-containing proteins and other small, secreted, cysteine-rich protein from oomycetes. These results suggest that PcCBP3 is an apoplastic effector and could be perceived by the plant immune system.