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Beyond Xylella, Integrated Management Strategies for Mitigating Xylella fastidiosa Impact in Europe (BeXyl) (Grant Agreement 101060593). Partner/Coordinador principal: Blanca B. Landa del Castillo, Investigadora Científica del Instituto de Agricultura Sostenible (IAS-CSIC)., While there are documented host shifts in many bacterial plant pathogens, the genetic foundation of host shifts is largely unknown. Xylella fastidiosa is a bacterial pathogen found in over 600 host plant species. Two parallel host shifts occurred-in Brazil and Italy-in which X. fastidiosa adapted to infect olive trees, whereas related strains infected coffee. Using 10 novel whole-genome sequences from an olive-infecting population in Brazil, we investigated whether these olive-infecting strains diverged from closely related coffee-infecting strains. Several single-nucleotide polymorphisms, many derived from recombination events, and gene gain and loss events separated olive-infecting strains from coffee-infecting strains in this clade. The olive-specific variation suggests that this event was a host jump with genetic isolation between coffee- and olive-infecting X. fastidiosa populations. Next, we investigated the hypothesis of genetic convergence in the host shift from coffee to olive in both populations (Brazil and Italy). Each clade had multiple mutations and gene gain and loss events unique to olive, yet no overlap between clades. Using a genome-wide association study technique, we did not find any plausible candidates for convergence. Overall, this work suggests that the two populations adapted to infect olive trees through independent genetic solutions., This work was partially funded by the California Department of Food and Agriculture Pierce's Disease Control Program. Additional funding was from Horizon 2020 (XF Actors project number 727987), BeXyl Project (HORIZON, id: 101060593), and FAPESP (São Paulo Research Foundation 2016/02176-7). H.D.C.F. received CNPq research fellowships (proc. no. 313676/2017-8).

Since emerging in Brazil in 1985, wheat blast has spread throughout South America and recently appeared in Bangladesh and Zambia. Here we show that two wheat resistance genes, Rwt3 and Rwt4, acting as host-specificity barriers against non-Triticum blast pathotypes encode a nucleotide-binding leucine-rich repeat immune receptor and a tandem kinase, respectively. Molecular isolation of these genes will enable study of the molecular interaction between pathogen effector and host resistance genes.

Sweet potato yellow curling (YC), the most severe disease of sweet potato detected in Argentina, causes symptoms and damage to sweet potato crops in all cultivated regions. Since 2010/11, the presence of four viruses has been detected in symptomatic cv. Arapey INIA: two potyviruses non-persistently transmitted by Myzus persicae (sweet potato feathery mottle virus, SPFMV and sweet potato virus G, SPVG); a closterovirus, sweet potato chlorotic stunt virus (SPCSV) and a geminivirus, sweet potato leaf curl virus (SPLCV), both transmitted by Bemisia tabaci in a semi-persistent and persistent manner, respectively. All the plants were collected from fields in Colonia Caroya, Córdoba province, Argentina. The objectives of the present work are to isolate and identify the virus or viruses involved in YC disease of sweet potato, and to elucidate the viral combination that reproduces YC symptoms. The most severe YC symptoms for this genotype in the field were only reproduced by a combination of the four viruses. The symptoms include chlorosis, stunting, mosaic, blistering, leaf curling, chlorotic spots, chlorotic patterns, leaf area reduction and distortion, and upward curling of leaf edges. The presence of each virus was detected by serological (DAS, NCM and TAS-ELISA) and molecular (PCR) tests. It is concluded that the interaction of SPFMV, SPVG, SPCSV and SPLCV is needed for the development of YC symptoms. El encrespamiento amarillo (EA), la enfermedad más severa detectada en Argentina, causa síntomas y daños en cultivos de batata en toda la región productora. Desde 2010/11 se ha detectado la presencia de cuatro virus en plantas sintomáticas del cv. Arapey INIA recolectadas en lotes de Colonia Caroya, provincia de Córdoba. Los virus son sweet potato feathery mottle virus (SPFMV) y sweet potato virus G (SPVG), dos potyvirus transmitidos de forma no persistente por Myzus persicae; un closterovirus: sweet potato chlorotic stunt virus (SPCSV) y un geminivirus: sweet potato leaf curl virus (SPLCV), ambos transmitidos por Bemisia tacabi de manera semipersistente y persistente, respectivamente. Los objetivos de este trabajo fueron aislar e identificar el o los virus involucrados en la enfermedad EA de la batata y determinar la combinación de virus que reproduce la sintomatología de EA Solo la combinación de los cuatro virus permitió reproducir la sintomatología más severa del encrespamiento amarillo observada a campo en dicho genotipo. Los síntomas incluyen clorosis, achaparramiento, mosaico, ampollado, enrulado de la hoja, manchas cloróticas, diseños cloróticos, reducción y distorsión del área foliar, bordes de la hoja curvados hacia arriba. La presencia de cada uno de los virus se detectó mediante pruebas serológicas (DAS, NCM y TAS-ELISA) y moleculares (PCR). Se concluye que la interacción de SPFMV, SPVG, SPCSV y SPLCV es necesaria para el desarrollo de EA. Fil: Flamarique, Sofía Solange. Fondo para la Investigación Científica y Tecnológica; Argentina Fil: Flamarique, Sofía Solange. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Patología Vegetal; Argentina Fil: Vilanova Perez, Antonella. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Patología Vegetal; Argentina Fil: Luque, Andres Vicente. Consejo Nacional de Investigaciones Científicas y Técnicas. Unidad de Fitopatología y Modelización Agrícola (UFyMA); Argentina Fil: Luque, Andres Vicente. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Patología Vegetal; Argentina Fil: Rodriguez Pardina, Patricia. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Patología Vegetal; Argentina Fil: Rodriguez Pardina, Patricia. Consejo Nacional de Investigaciones Científicas y Técnicas. Unidad de Fitopatología y Modelización Agrícola (UFyMA); Argentina Fil: Di Feo, Liliana Del Valle. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Patología Vegetal; Argentina Fil: Di Feo, Liliana Del Valle. Consejo Nacional de Investigaciones Científicas y Técnicas. Unidad de Fitopatología y Modelización Agrícola (UFyMA); Argentina

Banded leaf and sheath blight (BLSB) is a devasting disease caused by the necrotrophic fungus Rhizoctonia solani that affects maize (Zea mays L.) fields worldwide, especially in China and Southeast Asia. Understanding how maize plants respond to R. solani infection is a key step towards controlling the spread of this fungal pathogen. In this study, we determined the transcriptome of maize plants infected by a low-virulence strain (LVS) and a high-virulence strain (HVS) of R. solani for 3 and 5 days by transcriptome deep-sequencing (RNA-seq). We identified 3,015 (for LVS infection) and 1,628 (for HVS infection) differentially expressed genes (DEGs). We confirmed the expression profiles of 10 randomly selected DEGs by quantitative reverse transcription PCR. We also performed a Gene Ontology (GO) enrichment analysis to establish which biological processes are associated with these DEGs, which revealed the enrichment of defense-related GO terms in LVS- and HVS-regulated genes. We selected 388 DEGs upregulated upon fungal infection as possible candidate genes. Among them, the overexpression of ZmNAC41 (encoding NAC transcription factor 41) or ZmBAK1 (encoding BRASSINOSTEROID INSENSITIVE 1-associated receptor kinase 1) in rice enhanced resistance to R. solani. In addition, overexpressing ZmBAK1 in rice also increased plant height, plant weight, thousand-grain weight, and grain length. The identification of 388 potential key maize genes related to resistance to R. solani provides significant insights into improving BLSB resistance.

It has been two decades since the first detection of the sudden oak death pathogen Phytophthora ramorum in Oregon forests. Although the epidemic was managed since its first discovery in 2001, at least three invasions of three separate variants (clonal lineages), NA1, EU1, and NA2, are documented to have occurred to date. Control of this epidemic has cost over US$32 million from 2001 to 2020. This is dwarfed by the predicted cost of the closure to the Coos Bay export terminal, estimated at $58 million per year, if the epidemic was allowed to spread unchecked. Management efforts in Oregon have reduced inoculum and limited the spread of the pathogen. An outreach and citizen scientist program has been piloted to help in early detection efforts and search for disease-resistant tanoak. This feature article documents the repeated emergence, impact, costs, and lessons learned from managing this devastating invasive pathogen.

Expression of the VaRPP13 in Arabidopsis and tobacco enhanced resistance to oomycete pathogens, and this enhancement is closely related to the activation of salicylic acid (SA) signaling pathway. Resistance (R) genes, which usually contain a nucleotide-binding site and a leucine-rich repeat (NBS-LRR) domain, play crucial roles in disease resistance. In this study, we cloned a CC-NBS-LRR gene VaRPP13 from Vitis amurensis 'Shuang Hong' grapevine, and investigated its function on disease resistance. VaRPP13 expression was induced by Plasmopara viticola, an oomycetes pathogen causing downy mildew disease in grapevine. Heterologous expression VaRPP13 could also enhance resistance to Hyaloperonospora arabidopsidis in Arabidopsis thaliana and Phytophthora capsici in Nicotiana benthamiana, both oomycete pathogens. Further study indicated that VaRPP13 could enhance the expression of genes in SA signal pathway, while exogenous SA could also induce the expression of VaRPP13. In conclusion, our studies demonstrated that VaRPP13 contributes to a broad-spectrum resistance to oomycetes via activating SA signaling pathway.

Next generation sequencing has been used to identify and characterize the full genome sequence of a cassava-infecting torradovirus, revealing the presence of a Maf/HAM1 domain downstream of the RNA-dependent RNA-polymerase (RdRp) domain in RNA1 in all isolates sequenced. A similar domain is also found in unrelated potyvirids infecting Euphorbiaceae hosts in the Americas and cassava in Africa. Even though cassava torrado-like virus (CsTLV) could not be mechanically transmitted to a series of herbaceous hosts, it can be efficiently transmitted by bud graft-inoculation to different cassava landraces. Our bioassays show that CsTLV has a narrow host range. Crystal-like structures of isometric virus-like particles were observed in cells of plants with single infection by CsTLV, and consistently induced chlorotic leaf spots and affected root yields significantly. Moreover, CsTLV infection induces changes in the accumulation of total sugars in storage roots. Field surveys indicated the presence of CsTLV in the main cassava growing regions of Colombia, and the occurrence of two different cassava-infecting torradovirus species. Profiles of small RNAs of 21 to 24 nucleotides in length, derived from CsTLV RNAs targeted by cassava RNA silencing defense mechanisms, are also reported.

We describe a standard method for characterizing the virulence profile of Plasmopara viticola, the causal agent of grapevine downy mildew. We used 33 European strains to inoculate six grapevine varieties carrying the principal factors for resistance to downy mildew (Rpv1, Rpv3.1, Rpv3.2, Rpv5, Rpv6, Rpv10, and Rpv12) and the susceptible Vitis vinifera ‘Chardonnay’. For each interaction, we characterized the level of sporulation by image analysis and the intensity of the grapevine hypersensitive response by visual score. We propose a definition for the breakdown of grapevine quantitative resistances combining these two traits. Among the 33 strains analyzed, 28 are virulent on at least one resistance factor. We identified five different pathotypes across the 33 strains analyzed: two pathotypes overcoming a single resistance factor (vir3.1 and vir3.2) and three complex pathotypes overcoming multiple resistance factors (vir3.1,3.2; vir3.2,12; vir3.1,3.2,10). Our findings confirm the widespread occurrence of P. viticola strains overcoming the Rpv3 haplotypes (28 strains). We also detected the first breakdown of resistance to the Rpv10 by a strain from Germany and the breakdown of Rpv12 factors by a strain from Hungary. The pathotyping method proposed here and the associated differential host range lay the groundwork for the early detection of resistance breakdown in grapevines. This approach will also facilitate the monitoring of the evolution of P. viticola populations at large spatial scales. This is an essential step forward to promoting durable management of the resistant grapevine varieties currently available.

Pyricularia oryzae and Pyricularia grisea are pathogens that cause blast disease in various monocots. It has been reported that P. oryzae infects the leaves and roots of rice via different mechanisms. However, it is unclear to what extent the tissue types affect the host specificities of P. oryzae and P. grisea. Here, we evaluated the tissue-specific infection strategies of P. oryzae and P. grisea in various gramineous plants. Generally, mycelial plug inoculation caused root browning but the degree of browning did not simply follow the disease index on leaves. Interestingly, the Triticum and Digitaria pathotypes caused strong root growth inhibition in rice, wheat, and barley. Moreover, the Digitaria pathotype inhibited root branching only in rice. Culture filtrate reproduced these inhibitory effects on root, suggesting that some secreted molecules are responsible for the inhibitions. Observation of root sections revealed that most of the infection hyphae penetrated intercellular spaces and further extended into root cells, regardless of pathotype and host plant. The infection hyphae of Digitaria and Triticum pathotypes tended to localize in the outer layer of rice roots, but not in those of wheat and barley roots. The infection hyphae of the Oryza pathotype were distributed in both the intercellular and intracellular spaces of rice root cells. Pathogenesis-related genes and reactive oxygen species accumulation were induced after root inoculation with all combinations. These results suggest that resistance reactions were induced in the roots of gramineous plants against the infection with Pyricularia isolates but failed to prevent fungal invasion.

Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici, is a destructive disease in many countries. In China, wheat stripe rust generally occurs in northwestern and southwestern China and sporadically in the Jiangsu and Zhejiang coastal epidemiological region (JZER), where an outbreak of the disease occurred in 2019. To understand the population structure and potential inoculum sources of the pathogen in this region, 171 isolates collected from 93 wheat fields of 53 counties in 10 provinces were phenotyped with two sets of wheat differentials and genotyped with 20 pairs of single-nucleotide polymorphism primers. Phenotype tests indicated that identical races (CYR34, CYR33, Su11-139, and Su11-14-1) detected in Jiangsu and Zhejiang were shared with the oversummering regions (Gansu), overwintering regions (Hubei, Henan, and Shaanxi), and Yun-Gui epidemiological regions (Yunnan and Guizhou). In JZER, races CYR32, G22-14, and G22-68 were detected in Jiangsu, but not in Zhejiang, and Su11-208 was identified in Zhejiang, but not in Jiangsu. Genotypic analysis revealed remarkable gene flows among the Jiangsu, Yunnan, Henan, and Anhui populations, as well as those of Zhejiang, Guizhou, and Sichuan, showing that wheat stripe rust in Zhejiang and Jiangsu was from spores that migrated from different routes. Major gene flows were detected between the Jiangsu and Zhejiang populations. P. striiformis f. sp. tritici from both overwintering regions (Yunnan, Sichuan, Guizhou, Henan, Hubei, and Shaanxi) and oversummering regions (Gansu) contributed to the wheat stripe rust epidemic in the JZER region in 2019.

The bacterial pathogens Xanthomonas oryzae pathovars oryzae (Xoo) and oryzicola (Xoc) cause leaf blight and leaf streak diseases on rice, respectively. Pathogenesis is largely defined by the virulence genes harboured in the pathogen genome. Recently, we demonstrated that the protein HpaP of the crucifer pathogen Xanthomonas campestris pv. campestris is an enzyme with both ATPase and phosphatase activities, and is involved in regulating the synthesis of virulence factors and the induction of the hypersensitive response (HR). In this study, we investigated the role of HpaP homologues in Xoo and Xoc. We showed that HpaP is required for full virulence of Xoo and Xoc. Deletion of hpaP in Xoo and Xoc led to a reduction in virulence and alteration in the production of virulence factors, including extracellular polysaccharide and cell motility. Comparative transcriptomics and reverse transcription-quantitative PCR assays revealed that in XVM2 medium, a mimic medium of the plant environment, the expression levels of hrp genes (for HR and pathogenicity) were enhanced in the Xoo hpaP deletion mutant compared to the wild type. By contrast, in the same growth conditions, hrp gene expression was decreased in the Xoc hpaP deletion mutant compared to the wild type. However, an opposite expression pattern was observed when the pathogens grew in planta, where the expression of hrp genes was reduced in the Xoo hpaP mutant but increased in the Xoc hpaP mutant. These findings indicate that HpaP plays a divergent role in Xoo and Xoc, which may lead to the different infection strategies employed by these two pathogens.

The oomycete pathogen Phytophthora sojae is a causal agent of soybean root rot. Upon colonization of soybeans, P. sojae secretes various RXLR effectors to suppress host immune responses, supporting successful infection. Previous research has demonstrated that the RXLR effector Avh94 functions as a virulence effector, but the molecular mechanism underlying its role in virulence remains unknown. Here, we demonstrate that Avh94 overexpression in plants and pathogens promotes Phytophthora infection. Avh94 interacts with soybean JAZ1/2, which is a repressor of jasmonic acid (JA) signaling. Avh94 stabilizes JAZ1/2 to inhibit JA signaling and silencing of JAZ1/2 enhances soybean resistance against P. sojae. Moreover, P. sojae lines overexpressing Avh94 inhibit JA signaling. Furthermore, exogenous application of methyl jasmonate improves plant resistance to Phytophthora. Taken together, these findings suggest that P. sojae employs an RXLR effector to hijack JA signaling and thereby promote infection.

According to their lifestyle, plant pathogens are divided into biotrophic and necrotrophic organisms. Biotrophic pathogens exclusively nourish living host cells, whereas necrotrophic pathogens rapidly kill host cells and nourish cell walls and cell contents. To this end, the necrotrophic fungus Botrytis cinerea secretes large amounts of phytotoxic proteins and cell wall-degrading enzymes. However, the precise role of these proteins during infection is unknown. Here, we report on the identification and characterization of the previously unknown toxic protein hypersensitive response-inducing protein 1 (Hip1), which induces plant cell death. We found the adoption of a structurally conserved folded Alternaria alternata Alt a 1 protein structure to be a prerequisite for Hip1 to exert its necrosis-inducing activity in a host-specific manner. Localization and the induction of typical plant defense responses by Hip1 indicate recognition as a pathogen-associated molecular pattern at the plant plasma membrane. In contrast to other secreted toxic Botrytis proteins, the activity of Hip1 does not depend on the presence of the receptor-associated kinases BRI1-associated kinase 1 and suppressor of BIR1-1. Our results demonstrate that recognition of Hip1, even in the absence of obvious enzymatic or pore-forming activity, induces strong plant defense reactions eventually leading to plant cell death. Botrytis hip1 overexpression strains generated by CRISPR/Cas9 displayed enhanced infection, indicating the virulence-promoting potential of Hip1. Taken together, Hip1 induces a noncanonical defense response which might be a common feature of structurally conserved fungal proteins from the Alt a 1 family.

The mannose-binding lectin gene MANNOSE-BINDING LECTIN 1 (MBL1) is a member of the G-type lectin family and is involved in defense in strawberry (Fragaria × ananassa). Genome-wide identification of the G-type lectin family was carried out in woodland strawberry, F. vesca, and 133 G-lectin genes were found. Their expression profiles were retrieved from available databases and indicated that many are actively expressed during plant development or interaction with pathogens. We selected MBL1 for further investigation and generated stable transgenic FaMBL1-overexpressing plants of F. ×ananassa to examine the role of this gene in defense. Plants were selected and evaluated for their contents of disease-related phytohormones and their reaction to biotic stresses, and this revealed that jasmonic acid decreased in the overexpressing lines compared with the wild-type (WT). Petioles of the overexpressing lines inoculated with Colletotrichum fioriniae had lower disease incidence than the WT, and leaves of these lines challenged by Botrytis cinerea showed significantly smaller lesion diameters than the WT and higher expression of CLASS II CHITINASE 2-1. Our results indicate that FaMBL1 plays important roles in strawberry response to fungal diseases caused by C. fioriniae and B. cinerea.

Humidity is a critical environmental factor affecting the epidemic of plant diseases. However, it is still unclear how ambient humidity affects the occurrence of diseases in plants. In this study, we show that high ambient humidity enhanced blast development in rice plants under laboratory conditions. Furthermore, we found that high ambient humidity enhanced the virulence of Magnaporthe oryzae by promoting conidial germination and appressorium formation. In addition, the results of RNA-sequencing analysis and the ethylene content assessment revealed that high ambient humidity suppressed the accumulation of ethylene and the activation of ethylene signaling pathway induced by M. oryzae in rice. Knock out of ethylene signaling genes OsEIL1 and OsEIN2 or exogenous application of 1-methylcyclopropene (ethylene inhibitor) and ethephon (ethylene analogues) eliminated the difference of blast resistance between the 70% and 90% relative humidity conditions, suggesting that the activation of ethylene signaling contributes to humidity-modulated basal resistance against M. oryzae in rice. In conclusion, our results demonstrated that high ambient humidity enhances the virulence of M. oryzae and compromises basal resistance by reducing the activation of ethylene biosynthesis and signaling in rice. Results from this study provide cues for novel strategies to control rice blast under global environmental changes.

Efficient pathogen diagnostics and genotyping methods enable effective disease management and breeding, improve crop productivity and ensure food security. However, current germplasm selection and pathogen detection techniques are laborious, time-consuming, expensive and not easy to mass-scale application in the field. Here, we optimized a field-deployable lateral flow assay, Bio-SCAN, as a highly sensitive tool to precisely identify elite germplasm and detect mutations, transgenes and phytopathogens in1 h, starting from sample isolation to result output using lateral flow strips. As a proof of concept, we genotyped various wheat germplasms for the Lr34 and Lr67 alleles conferring broad-spectrum resistance to stripe rust, confirmed the presence of synthetically produced herbicide-resistant alleles in the rice genome and screened for the presence of transgenic elements in the genome of transgenic tobacco and rice plants with 100% specificity. We also successfully applied this new assay to the detection of phytopathogens, including viruses and bacterial pathogens in Nicotiana benthamiana, and two destructive fungal pathogens (Puccinia striiformis f. sp. tritici and Magnaporthe oryzae Triticum) in wheat. Our results illustrate the power of Bio-SCAN in crop breeding, genetic engineering and pathogen diagnostics to enhance food security. The high sensitivity, simplicity, versatility and in-field deployability make the Bio-SCAN as an attractive molecular diagnostic tool for diverse applications in agriculture.

The Xanthomonas transcription activator-like effector (TALE) protein AvrBs3 transcriptionally activates the executor-type resistance (R) gene Bs3 from pepper (Capsicum annuum), thereby triggering a hypersensitive cell death reaction (HR). AvrBs3 also triggers an HR in tomato (Solanum lycopersicum) upon recognition by the nucleotide-binding leucine-rich repeat (NLR) R protein Bs4. Whether the executor-type R protein Bs3 and the NLR-type R protein Bs4 use common or distinct signalling components to trigger an HR remains unclear. CRISPR/Cas9-mutagenesis revealed, that the immune signalling node EDS1 is required for Bs4- but not for Bs3-dependent HR, suggesting that NLR- and executor-type R proteins trigger an HR via distinct signalling pathways. CRISPR/Cas9-mutagenesis also revealed that tomato Bs4 suppresses the virulence function of both TALEs, the HR-inducing AvrBs3 protein and of AvrHah1, a TALE that does not trigger an HR in tomato. Analysis of AvrBs3- and AvrHah1-induced host transcripts and disease phenotypes in CRISPR/Cas9-induced bs4 mutant plants indicates that both TALEs target orthologous transcription factor genes to promote disease in tomato and pepper host plants. Our studies display that tomato mutants lacking the TALE-sensing Bs4 protein provide a novel platform to either uncover TALE-induced disease phenotypes or genetically dissect components of executor-triggered HR.

The current study evaluated the effect of plant growth-promoting (PGP) strains of Streptomyces on yield, quality, and nitrate content of fruits, plant-microbe responses, and antifungal effect against blight disease caused by fungus pathogen Alternaria solani on tomato fruits in commercial greenhouse conditions. Greenhouse trials were done with four treatments including strains Y28, IC10, IT25, and commercial bio-fertilizer (Barvar NPK®) on tomato plants. In PGP treatments, the number of infected fruits significantly reduced (60%) compared to Barvar and control. Strain Y28 improved the quality of tomatoes more than other treatments. All three PGP treatments contained a higher level of total sugar concentration and antioxidant enzyme activities than Barvar and control. In contrast, PGP strains, especially Y28, significantly reduced nitrate accumulation (25%) compared to Barvar and control tomatoes. Streptomyces treatments induced more than a 20-fold increase in UDP and WRKY70 transcription factor gene expression relative to the control (P 0.01). Based on the results, microbe-dependent plant defense induced by these strains is positively correlated to WRKY70 expression and nitrate reduction in commercial greenhouse conditions. These findings suggest that the commercial application of specific strains not only can illustrate an eco-friendly solution to induce resistance against fungal pathogens but also improve the quality properties of food plants with lower nitrate content.

For many plant-pathogenic or endophytic fungi, production of mycotoxins, which are toxic to humans, may present a fitness gain. However, associations between mycotoxin production and plant pathogenicity or virulence is inconsistent and difficult due to the complexity of these host–pathogen interactions and the influences of environmental and insect factors. Aflatoxin receives a lot of attention due to its potent toxicity and carcinogenicity but the connection between aflatoxin production and pathogenicity is complicated by the pathogenic ability and prevalence of nonaflatoxigenic isolates in crops. Other toxins directly aid fungi in planta, trichothecenes are important virulence factors, and ergot alkaloids limit herbivory and fungal consumption due to insect toxicity. We review a panel discussion at the American Phytopathological Society's Plant Health 2021 conference, which gathered diverse experts representing different research sectors, career stages, ethnicities, and genders to discuss the diverse roles of mycotoxins in the lifestyles of filamentous fungi of the families Clavicipitaceae, Trichocomaceae (Eurotiales), and Nectriaceae (Hypocreales).

Dry flower disease caused by Pestalotiopsis/Neopestalotiopsis spp., green mold caused by Cladosporium spp., and gray mold caused by Botrytis spp., collectively known as flower blight cause significant yield losses in macadamia. Potential sources of inoculum of the various pathogens in macadamia tree canopy were examined using pathogenicity tests and a multiplex quantitative PCR (qPCR) assay developed in this study. The qPCR assay detected and quantified the relative abundance of the inoculum of flower blight pathogens. The assay revealed that remnant racemes contributed a high amount of inoculum of all the three groups of flower blight pathogens, while the yellow halo leaf spot contributed only Pestalotiopsis/Neopestalotiopsis species. The amount of conidia per gram of remnant racemes ranged from 7 × 103 to 2 × 104 for dry flower disease, 3 × 103 to 1 × 104 for green mold, and 5 to 8 × 103 for gray mold pathogens. Conidia of Pestalotiopsis/Neopestalotiopsis species quantified from leaf spots varied from 1 × 102 to 1 × 103 per cm2. Pathogenicity tests performed on developing racemes under field conditions, using conidial suspensions from both sources of inoculum (remnant racemes and yellow halo leaf spot), resulted in severe flower bight symptoms. Disease severity was not significantly different ( P > 0.05) when remnant racemes were incubated directly with the developing racemes compared with inoculation with conidial suspension from the material. This suggests that racemes from preceding seasons that remain in the tree canopy carryover inoculum between seasons and should be removed as a control option for flower blights in macadamia orchards.

Fusarium wilt, caused by Fusarium oxysporum f. sp. vasinfectum, is a severe disease of cotton (Gossypium spp.). Strains of the wilt pathogen in the United States, such as race 1, require the presence of nematodes such as southern root-knot nematode (Meloidogyne incognita) to cause appreciable disease. The exception is the race 4 strain of the wilt pathogen, which can attack cotton without concomitant infection by plant-parasitic nematodes and was first identified in California in 2001 and in Texas and New Mexico since 2017. The effects of the interaction between M. incognita and race 1 or race 4 on wilt severity and nematode reproduction on two Gossypium hirsutum cultivars, Acala 44 and FM 966, and a G. barbadense cultivar, Pima S-4, were directly compared in growth chamber assays. All three cultivars were susceptible to M. incognita. Suppression of nematode reproduction by the wilt pathogen was detected only for race 4 on all three cultivars on a per plant basis but not on a per gram root tissue basis. The control, M. incognita alone, and race 1 alone treatments caused no symptoms. Inoculation with race 1 and M. incognita caused moderate wilt symptoms in ‘Acala 44’ and ‘FM 966’ and mild symptoms in ‘Pima S-4’. However, race 4 treatment caused severe wilt in ‘Pima S-4’ and moderate wilt severity in ‘Acala 44’ and ‘FM 966’. The symptom severity of ‘Acala 44’ and ‘FM 966’ further increased in the presence of M. incognita. Thus, race 4 is not only capable of causing wilt in the absence of M. incognita but can also interact with the nematode to further increase disease severity. Though control of wilt caused by race 1 can be achieved mainly through breeding for nematode resistance, it will be imperative to incorporate both southern root-knot nematode and race 4 resistance to effectively control the disease should race 4 expand into southern root-knot nematode-infested fields.

Rice production is threatened by multiple pathogens. Breeding cultivars with broad-spectrum disease resistance is necessary to maintain and improve crop production. Previously we found that overexpression of miR160a enhanced rice blast disease resistance. However, it is unclear whether miR160a also regulates resistance against other pathogens, and what the downstream signaling pathways are. Here, we demonstrate that miR160a positively regulates broad-spectrum resistance against the causative agents of blast, leaf blight and sheath blight in rice. Mutations of miR160a-targeted Auxin Response Factors result in different alteration of resistance conferred by miR160a. miR160a enhances disease resistance partially by suppressing ARF8, as mutation of ARF8 in MIM160 background partially restores the compromised resistance resulting from MIM160. ARF8 protein binds directly to the promoter and suppresses the expression of WRKY45, which acts as a positive regulator of rice immunity. Mutation of WRKY45 compromises the enhanced blast resistance and bacterial leaf blight resistance conferred by arf8 mutant. Overall, our results reveal that a microRNA coordinates rice broad-spectrum disease resistance by suppressing multiple target genes that play different roles in disease resistance, and uncover a new regulatory pathway mediated by the miR160a-ARF8 module. These findings provide new resources to potentially improve disease resistance for breeding in rice.

ENHANCED DISEASE SUSCEPTIBILITY 1 (EDS1) mediates the induction of defense responses against pathogens in most angiosperms. However, it has recently been shown that a few species have lost EDS1. It is unknown how defense against disease unfolds and evolves in the absence of EDS1. We utilize duckweeds; a collection of aquatic species that lack EDS1, to investigate this question. We established duckweed-Pseudomonas pathosystems and used growth curves and microscopy to characterize pathogen-induced responses. Through comparative genomics and transcriptomics, we show that the copy number of infection-associated genes and the infection-induced transcriptional responses of duckweeds differ from other model species. Pathogen defense in duckweeds has evolved along different trajectories than in other plants, including genomic and transcriptional reprogramming. Specifically, the miAMP1 domain-containing proteins, which are absent in Arabidopsis, showed pathogen responsive upregulation in duckweeds. Despite such divergence between Arabidopsis and duckweed species, we found conservation of upregulation of certain genes and the role of hormones in response to disease. Our work highlights the importance of expanding the pool of model species to study defense responses that have evolved in the plant kingdom independent of EDS1.

Gram-negative bacterial plant pathogens inject effectors into their hosts to hijack and manipulate metabolism, eluding surveillance at the battle frontier on the cell surface. The effector AvrRpm1Pma from Pseudomonas syringae pv. maculicola functions as an ADP-ribosyl transferase that modifies RESISTANCE TO P. SYRINGAE PV MACULICOLA1 (RPM1)-INTERACTING PROTEIN4 (RIN4), leading to the activation of Arabidopsis thaliana (Arabidopsis) resistance protein RPM1. Here we confirmed the ADP-ribosyl transferase activity of another bacterial effector, AvrRpm2Psa from P. syringae pv. actinidiae, via sequential inoculation of Pseudomonas strain Pto DC3000 harboring avrRpm2Psa following Agrobacterium-mediated transient expression of RIN4 in Nicotiana benthamiana. We conducted mutational analysis in combination with mass spectrometry to locate the target site in RIN4. A conserved glutamate residue (Glu156) is the most likely target for AvrRpm2Psa, as only Glu156 could be ADP-ribosylated to activate RPM1 among candidate target residues identified from the MS/MS fragmentation spectra. Soybean (Glycine max) and snap bean (Phaseolus vulgaris) RIN4 homologs without glutamate at the positions corresponding to Glu156 of Arabidopsis RIN4 are not ADP-ribosylated by bacterial AvrRpm2Psa. In contrast to the effector AvrB, AvrRpm2Psa does not require the phosphorylation of Thr166 in RIN4 to activate RPM1. Therefore, separate biochemical reactions by different pathogen effectors may trigger the activation of the same resistance protein via distinct modifications of RIN4.

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.

Fusarium graminearum is a fungal pathogen that causes Fusarium head blight in cereal crops. The identification of proteins secreted from pathogens to overcome plant defenses and cause disease, collectively known as effectors, can reveal the etiology of a disease process. Proximity-dependent biotin identification (BioID) was used to identify potential effector proteins secreted in planta by F. graminearum during the infection of Arabidopsis. Mass spectrometry analysis of streptavidin affinity-purified proteins revealed over 300 proteins from F. graminearum, of which 62 were candidate effector proteins (CEPs). An independent analysis of secreted proteins from axenic cultures of F. graminearum showed a 42% overlap with CEPs, thereby assuring confidence in the BioID methodology. The analysis also revealed that 19 out of 62 CEPs (approx. 30%) had been previously characterized with virulence function in fungi. The functional characterization of additional CEPs was undertaken through deletion analysis by the CRISPR/Cas9 method, and by overexpression into Triticum aestivum (wheat) leaves by the Ustilago hordei delivery system. Deletion studies of 12 CEPs confirmed the effector function of three previously characterized CEPs and validated the function of another four CEPs on wheat inflorescence or vegetative tissues. Lastly, overexpression in wheat showed that all seven CEPs enhanced resistance against the bacterial pathogen Pseudomonas syringae DC3000.

Palm lethal decline is a disease that is always fatal to infected palm hosts and is caused by three species of phytoplasma in the Caribbean basin: ‘Candidatus Phytoplasma palmae’, ‘Ca. P. aculeata’, and ‘Ca. P. hispanola’. Movement of these pathogens throughout the Caribbean has been documented since their discovery in Jamaica. Over time, means of confirming infections in palms have improved. Current protocols utilize quantitative PCR (qPCR) for rapid amplification and distinction of these phytoplasmas using TaqMan probes and high-resolution melt-curve analysis (HRMA) of the 16S rRNA gene. These assays either do not detect all three phytoplasmas (HRMA) or do not distinguish between the three (TaqMan). In this study, a new qPCR-HRMA assay is developed that amplifies and distinguishes all three phytoplasmas currently known to kill palms in the Caribbean. Efficiency for the primer set secA614_F/secA759_R was shown to be consistent for all species at each concentration and yielded distinct melting temperature ranges for amplicons of ‘Ca. P. palmae’ (73.3 to 73.4°C), ‘Ca. P. aculeata’ (72.9 to 73.0°C), and ‘Ca. P. hispanola’ (73.5 to 73.6°C). This assay is a useful new tool not only for diagnostics that will contribute to monitoring and management programs, but it will also aid in basic research by allowing rapid screening of large samples in the context of vector surveys or identification of reservoir hosts.

Puccinia striiformis Westend. f. sp. tritici, commonly known as stripe rust, is an economically important pathogen of wheat (Triticum aestivum L.). The hexaploid club spring wheat cultivar JD contains both all-stage and adult plant resistance (APR) genes and exhibited consistent high resistance to stripe rust in the field. In this study, we aimed to identify the quantitative trait loci (QTL) for stripe rust resistance using a BC1F7 back-cross inbred-line population derived from the cross of JD and the recurrent parental line ‘Avocet’. The population was phenotyped in field plots in Washington State at the Spillman Agronomy Farm in Pullman and Mount Vernon Northwest Washington Research and Extension Center in between 2014 and 2016. A major QTL tentatively designated as QYrJD.wsu-1B, conferring all-stage resistance in JD background, was identified and mapped at the telomere region on the short arm of chromosome 1B using the genotyping-by-sequencing method. This QTL was further characterized with simple sequence repeat (SSR) markers and found to have the greatest logarithm-of-the-odds score and phenotypic effect, using SSR marker wmc798 on chromosome 1BS. Seven additional QTLs associated with APR were identified in the JD background on chromosomes 2D, 3A, 3B, 4A, 6B, and 7A with partial phenotypic effects.

Northern corn leaf blight (NCLB), caused by Exserohilum turcicum, is a devastating disease of corn in China. To enhance our understanding of NCLB epidemiology, the temporal progress and spatial patterns of NCLB were investigated. A susceptible corn cultivar, Xianyu 335, was planted in a field in Beijing in 2016 and 2017. Leaf lesions of NCLB on each plant were counted twice a week during the growing seasons. In addition, temporal disease progress was monitored for 8 weeks in three commercial corn fields in each of Yanqing, Miyun, Daxing, and Haidian Districts of Beijing in 2017, and the spatial patterns of diseased plants and NCLB lesion counts per plant were assessed in three commercial corn fields with moderate to high NCLB incidence in Yanqing District. The results demonstrated that a logistic model was the most appropriate to describe the temporal progress of NCLB incidence. The initial disease incidence was the key factor affecting disease epidemics under various conditions in the four districts of Beijing during the study. The higher the initial incidence of NCLB, the higher the final incidence. Thus, the earlier in the season NCLB incidence attained 1%, the higher was the final disease incidence. Greater than 1.0 variance-to-mean ratios suggested that the leaf lesions of NCLB tended to be aggregated on a plant. According to results from join-counts, variance of moving window averages, and semivariogram analysis, diseased corn plants and lesion numbers on each plant were aggregated in the field. The clustered pattern of NCLB lesions and infected plants suggested that conidia produced locally on diseased plants were important for disease spread within the field. The aggregated pattern of diseased plants suggested that plants should be sampled from more sites in a field to accurately estimate incidence of NCLB.

Pythium spp. is one of the major groups of pathogens that cause seedling diseases on soybean, leading to both preemergence and postemergence damping-off and root rot. More than 100 species have been identified within this genus, with Pythium irregulare, P. sylvaticum, P. ultimum var ultimum, and P. torulosum being particularly important for soybean production given their aggressiveness, prevalence, and abundance in production fields. This study investigated the antagonistic activity of potential biological control agents (BCAs) native to the U.S. Midwest against Pythium spp. First, in vitro screening identified BCAs that inhibit P. ultimum var. ultimum growth. Scanning electron microscopy demonstrated evidence of mycoparasitism of all potential biocontrol isolates against P. ultimum var. ultimum and P. torulosum, with the formation of appressorium-like structures, short hyphal branches around host hyphae, hook-shaped structures, coiling, and parallel growth of the mycoparasite along the host hyphae. Based on these promising results, selected BCAs were tested under field conditions against six different Pythium spp. Trichoderma afroharzianum 26 used alone and a mix of T. hamatum 16 + T. afroharzianum 19 used as seed treatments protected soybean seedlings from Pythium spp. infection, as BCA-treated plots had on average 15 to 20% greater plant stand and vigor than control plots. Our results also indicate that some of these potential BCAs could be added with a fungicide seed treatment with minimum inhibition occurring, depending on the fungicide active ingredient. This research highlights the need to develop tools incorporating biological control as a facet of soybean seedling disease management programs. The harnessing of native BCAs could be integrated with other management strategies to provide efficient control of seedling diseases.

Research on plant–virus–vector interactions has revealed that viruses can enhance their spread to new host plants by attracting nonviruliferous vectors to infected plants or driving viruliferous vectors to noninfected plants. However, whether viruses can also modulate the feeding preference of viruliferous vectors for different plant parts remains largely unknown. Here, by using rice stripe virus (RSV) and its vector, the small brown planthopper (SBPH), as a model, the effect of the virus on the feeding preference of its vector was studied by calculating the number of nonviruliferous and viruliferous SBPHs settling on different parts of rice plants. The results showed that the RSV-free SBPHs significantly preferred feeding on the stems of rice plants, whereas RSV-carrying SBPHs fed more on rice leaves. Moreover, the rice plants inoculated with RSV on the leaves showed more severe symptoms, with enhanced disease incidence and virus accumulation compared with rice plants inoculated at the top and bottom of stems, suggesting that the leaves are more susceptible to RSV than the stems of rice plants. These results demonstrate that RSV modulates the feeding preference of its transmitting vector SBPH from the stems to leaves of rice plants to promote virus infection. Interestingly, we also found that the leaves were more susceptible than the stems to rice black-streaked dwarf virus. This study proves that the feeding preference of insect vectors can be modulated by plant viruses to facilitate virus transmission.

Between 2000 and 2020, canker diseases of nut and stone fruit trees have become very widespread and severe in California. This study determined the effects of temperature on the development of canker-causing pathogens of almond and prune. Five pathogen taxa, Botryosphaeria dothidea, Cytospora leucostoma, Diaporthe (Phomopsis) neotheicola, Lasiodiplodia citricola, and Neofusicoccum mediterraneum, were used. Colony growth on medium and canker lesion development on detached shoots were measured at 10, 15, 20, 25, 30, and 35°C. The effects of temperature on colony growth differed among different pathogen taxa, although 25°C was the optimal temperature for most of the pathogens tested. The patterns of lesion growth as response to temperature were different among the different pathogens and tree crops. On almond, the highest growth rates appeared at 30°C for B. dothidea and L. citricola, but at 20°C for N. mediterraneum. The growth rates for C. leucostoma were lower than those of the other three pathogen taxa, with the highest rates recorded at 25°C. However, on prune, C. leucostoma showed greater lesion growth rates at different temperatures than the other pathogen taxa and maximum growth at 30 to 35°C. Similar trends were observed for L. citricola. The growth rates of B. dothidea and N. mediterraneum were comparatively lower than those of C. leucostoma and L. citricola.

Sudden death syndrome (SDS), caused by Fusarium virguliforme, causes substantial yield losses in soybean. However, relationships between soybean yield and components of disease progress, including time of disease onset, are poorly understood. Individual soybean plants (2018) and quadrats (2016 to 2018) were monitored in commercial fields and experimental plots in Iowa to quantify the impact of SDS foliar symptom onset on final SDS intensity, soybean yield components, and yield. The date when SDS foliar symptoms were first detected (onset time) and progress of SDS incidence and severity were recorded weekly. Individual soybean plants and quadrats were harvested at the end of each season. Beta-regression showed that date of SDS onset had a consistent and stable effect on final disease intensity both at individual plant and quadrat levels. The slope of the relationship between date of SDS onset and final SDS severity was common across all field sites and years. Weighted linear regression revealed that SDS onset explained 60 to 83% of the variation in number of pods, number of seeds, and total seed weight in individual plants, and 94 to 97% of the variation in seed yield in quadrats. Soybean yield damage functions (slopes) indicated that for each day SDS onset was delayed, soybean yield increased by 30.5 to 31.3 kg/ha. This new quantitative information improves understanding of the impact of SDS on final disease intensity and soybean yield. Further experiments are needed to determine how this relationship is affected by site-specific factors.

The plant journal 112(1), 84-103 (2022). doi:10.1111/tpj.15930, Published by Wiley-Blackwell, Oxford [u.a.]

The yield and quality of tomatoes (Solanum lycopersicum) is seriously affected by Phytophthora infestans. The long non-coding RNA (lncRNA) Sl-lncRNA39896 is induced after P. infestans infection and was previously predicted to act as an endogenous target mimic (eTM) for the microRNA Sl-miR166b, which function in stress responses. Here, we further examined the role of Sl-lncRNA39896 and Sl-miR166b in tomato resistance to P. infestans. Sl-miR166b levels were higher in Sl-lncRNA39896-knockout mutants than in wild-type plants, and the mutants displayed enhanced resistance to P. infestans. A six-point mutation in the region of Sl-lncRNA39896 that binds to Sl-miR166b disabled the interaction, suggesting that Sl-lncRNA39896 acts as an eTM for Sl-miR166b. Overexpressing Sl-miR166b yielded a similar phenotype to that produced by Sl-lncRNA39896-knockout, whereas silencing of Sl-miR166b impaired resistance. We verified that Sl-miR166b cleaved transcripts of its target class III homeodomain-leucine zipper genes SlHDZ34 and SlHDZ45. Silencing of SlHDZ34/45 decreased pathogen accumulation in plants infected with P. infestans. Additionally, jasmonic acid and ethylene contents were elevated following infection in the plants with enhanced resistance. Sl-lncRNA39896 is the first known lncRNA to negatively regulate resistance to P. infestans in tomato. We propose a novel mechanism in which the lncRNA39896-miR166b-HDZ module modulates resistance to P. infestans.

Root-knot nematodes (RKNs; Meloidogyne spp.) engage in complex parasitic interactions with many different host plants around the world, initiating elaborate feeding sites and disrupting host root architecture. Although RKNs have been the focus of research for many decades, new molecular tools have provided useful insights into the biological mechanisms these pests use to infect and manipulate their hosts. From identifying host defense mechanisms underlying resistance to RKNs to characterizing nematode effectors that alter host cellular functions, the past decade of research has significantly expanded our understanding of RKN–plant interactions, and the increasing number of quality parasite and host genomes promises to enhance future research efforts into RKNs. In this review, we have highlighted recent discoveries, summarized the current understanding within the field, and provided links to new and useful resources for researchers. Our goal is to offer insights and tools to support the study of molecular RKN–plant interactions.

Resistance to the soybean cyst nematode (SCN) is a topic incorporating multiple mechanisms and multiple types of science. It is also a topic of substantial agricultural importance, as SCN is estimated to cause more yield damage than any other pathogen of soybean, one of the world's main food crops. Both soybean and SCN have experienced jumps in experimental tractability in the past decade, and significant advances have been made. The rhg1-b locus, deployed on millions of farm acres, has been durable and will remain important, but local SCN populations are gradually evolving to overcome rhg1-b. Multiple other SCN resistance quantitative trait loci (QTL) of proven value are now in play with soybean breeders. QTL causal gene discovery and mechanistic insights into SCN resistance are contributing to both basic and applied disciplines. Additional understanding of SCN and other cyst nematodes will also grow in importance and lead to novel disease control strategies.

Although the phloem is a highly specialized tissue, certain pathogens, including phytoplasmas, spiroplasmas, and viruses, have evolved to access and live in this sequestered and protected environment, causing substantial economic harm. In particular, Candidatus Liberibacter spp. are devastating citrus in many parts of the world. Given that most phloem pathogens are vectored, they are not exposed to applied chemicals and are therefore difficult to control. Furthermore, pathogens use the phloem network to escape mounted defenses. Our review summarizes the current knowledge of phloem anatomy, physiology, and biochemistry relevant to phloem/pathogen interactions. We focus on aspects of anatomy specific to pathogen movement, including sieve plate structure and phloem-specific proteins. Phloem sampling techniques are discussed. Finally, pathogens that cause particular harm to the phloem of crop species are considered in detail.

The roles of mitogen-activated protein kinases (MAPKs) in plant–fungal pathogenic interactions are poorly understood in crops. Here, microscopic, phenotypic, proteomic, and biochemical analyses revealed that roots of independent transcription activator-like effector nuclease (TALEN)-based knockout lines of barley (Hordeum vulgare L.) MAPK 3 (HvMPK3 KO) were resistant against Fusarium graminearum infection. When co-cultured with roots of the HvMPK3 KO lines, F. graminearum hyphae were excluded to the extracellular space, the growth pattern of extracellular hyphae was considerably deregulated, mycelia development was less efficient, and number of appressoria-like structures and their penetration potential were substantially reduced. Intracellular penetration of hyphae was preceded by the massive production of reactive oxygen species (ROS) in attacked cells of the wild-type (WT), but ROS production was mitigated in the HvMPK3 KO lines. Suppression of ROS production in these lines coincided with elevated abundance of catalase (CAT) and ascorbate peroxidase (APX). Moreover, differential proteomic analysis revealed downregulation of several defense-related proteins in WT, and the upregulation of pathogenesis-related protein 1 (PR-1) and cysteine proteases in HvMPK3 KO lines. Proteins involved in suberin formation, such as peroxidases, lipid transfer proteins (LTPs), and the GDSL esterase/lipase (containing “GDSL” aminosequence motif) were differentially regulated in HvMPK3 KO lines after F. graminearum inoculation. Consistent with proteomic analysis, microscopic observations showed enhanced suberin accumulation in roots of HvMPK3 KO lines, most likely contributing to the arrested infection by F. graminearum. These results suggest that TALEN-based knockout of HvMPK3 leads to barley root resistance against Fusarium root rot.

Pattern recognition receptors (PRRs) sense ligands in pattern-triggered immunity (PTI). Plant PRRs include numerous receptor-like proteins (RLPs), but many RLPs remain functionally uncharacterized. Here, we examine an Arabidopsis thaliana RLP, RLP53, which positively regulates immune signaling. Our forward genetic screen for suppressors of enhanced disease resistance1 (edr1) identified a point mutation in RLP53 that fully suppresses disease resistance and mildew-induced cell death in edr1 mutants. The rlp53 mutants showed enhanced susceptibility to virulent pathogens, including fungi, oomycetes, and bacteria, indicating that RLP53 is important for plant immunity. The ectodomain of RLP53 contains leucine-rich repeat (LRR) motifs. RLP53 constitutively associates with the LRR receptor-like kinase SUPPRESSOR OF BRASSINOSTEROID INSENSITIVE1-ASSOCIATED KINASE (BAK1)-INTERACTING RECEPTOR KINASE1 (SOBIR1) and interacts with the co-receptor BAK1 in a pathogen-induced manner. The double mutation sobir1-12 bak1-5 suppresses edr1-mediated disease resistance, suggesting that EDR1 negatively regulates PTI modulated by the RLP53-SOBIR1-BAK1 complex. Moreover, the glycosylphosphatidylinositol (GPI)-anchored protein LORELEI-LIKE GPI-ANCHORED PROTEIN1 (LLG1) interacts with RLP53 and mediates RLP53 accumulation in the plasma membrane. We thus uncovered the role of a novel RLP and its associated immune complex in plant defense responses and revealed a potential new mechanism underlying regulation of RLP immune function by a GPI-anchored protein.

Plant biotrophic pathogens employ secreted molecules, called effectors, to suppress the host immune system and redirect the host's metabolism and development in their favour. Putative effectors of the gall-inducing maize pathogenic fungus Ustilago maydis were analysed for their ability to induce auxin signalling in plants. Using genetic, biochemical, cell-biological, and bioinformatic approaches we functionally elucidate a set of five, genetically linked effectors, called Topless (TPL) interacting protein (Tips) effectors that induce auxin signalling. We show that Tips induce auxin signalling by interfering with central corepressors of the TPL family. CRISPR-Cas9 mutants and deletion strain analysis indicate that the auxin signalling inducing subcluster effectors plays a redundant role in virulence. Although none of the Tips seem to have a conserved interaction motif, four of them bind solely to the N-terminal TPL domain and, for Tip1 and Tip4, we demonstrate direct competition with auxin/indole-3-acetic acid transcriptional repressors for their binding to TPL class of corepressors. Our findings reveal that TPL proteins, key regulators of growth-defence antagonism, are a major target of the U. maydis effectome.

Plants have many types of immune receptors that recognize diverse pathogen molecules and activate the innate immune system. The intracellular immune receptor family of nucleotide-binding domain leucine-rich repeat-containing proteins (NLRs) perceives translocated pathogen effector proteins and executes a robust immune response, including programmed cell death. Many plant NLRs have functionally specialized to sense pathogen effectors (sensor NLRs) or to execute immune signaling (helper NLRs). Sub-functionalized NLRs form a network-type receptor system known as the NLR network. In this review, we highlight the concept of NLR networks, discussing how they are formed, activated and regulated. Two main types of NLR networks have been described in plants: the ACTIVATED DISEASE RESISTANCE 1/N REQUIREMENT GENE 1 network and the NLR-REQUIRED FOR CELL DEATH network. In both networks, multiple helper NLRs function as signaling hubs for sensor NLRs and cell-surface-localized immune receptors. Additionally, the networks are regulated at the transcriptional and posttranscriptional levels, and are also modulated by other host proteins to ensure proper network activation and prevent autoimmunity. Plant pathogens in turn have converged on suppressing NLR networks, thereby facilitating infection and disease. Understanding the NLR immune system at the network level could inform future breeding programs by highlighting the appropriate genetic combinations of immunoreceptors to use while avoiding deleterious autoimmunity and suppression by pathogens.

Microbe‐associated molecular pattern (MAMP)‐triggered immunity (MTI) research has traditionally centred around signal transduction pathways originating from activated membrane‐localized pattern recognition receptors (PRRs), culminating in nuclear transcription and posttranslational modifications. More recently, chloroplasts have emerged as key immune signalling hubs, playing a central role in integrating environmental signals. Notably, MAMP recognition induces chloroplastic reactive oxygen species (cROS) that is suppressed by pathogen effectors, which also modify the balance of chloroplast‐synthesized precursors of the defence hormones, jasmonic acid, salicylic acid (SA) and abscisic acid. This study focuses on how well‐characterized PRRs and coreceptors modulate chloroplast physiology, examining whether diverse signalling pathways converge to similarly modulate chloroplast function. Pretreatment of receptor mutant plants with MAMP and D(Damage)AMP peptides usually protect against effector modulation of chlorophyll fluorescence and prevent Pseudomonas syringae effector‐mediated quenching of cROS and suppression of maximum dark‐adapted quantum efficiency (the ratio of variable/maximum fluorescence [Fv/Fm]). The MTI coreceptor double mutant, bak1‐5/bkk1‐1, exhibits a remarkable decrease in Fv/Fm compared to control plants during infection, underlining the importance of MTI‐mediated signalling in chloroplast immunity. Further probing the role of the chloroplast in immunity, we unexpectedly found that even moderate changes in light intensity can uncouple plant immune signalling.

Flavescence dorée (FD) is a quarantine disease threatening European vineyards. Its management is based on mandatory insecticide treatments and the uprooting of infected plants identified during annual surveys. Field surveys are currently not optimized because the drivers affecting FD spread in vineyard landscapes remain poorly understood. We collated a georeferenced dataset of FD detection, collected from 34,581 vineyard plots over 5 years in the South West France wine region. Spatial models fitted with integrated nested Laplace approximation were used to identify local and landscape factors affecting FD detection and infection. Our analysis highlights the importance of sampling period on FD detection and of local practices and landscape context on FD infection. At field scale, altitude and cultivar choice were the main factors affecting FD infection. In particular, the odds ratio of FD infection in fields planted with the susceptible Cabernet Sauvignon, Cabernet Franc, or Muscadelle varieties were approximately twice those in fields planted with the less susceptible Merlot. Field infection was also affected by the field’s immediate surroundings (within a circle with a radius of 150 to 200 m), corresponding to landscapes of 7 to 12 ha. In particular, the probability of FD infection increased with the proportions of forest and urban land and with the proportion of susceptible cultivars, demonstrating that the cultivar composition impacts FD epidemiology at landscape scale. The satisfactory predictive performance of the model for identifying districts with a prevalence of FD detection >10% of the fields suggests that it could be used to target areas in which future surveys would be most valuable.

Apoplastic ascorbate oxidases (AOs) play a critical role in reactive oxygen species (ROS)-mediated innate host immunity by regulating the apoplast redox state. To date, little is known about how apoplastic effectors of the rice blast fungus Magnaporthe oryzae modulate the apoplast redox state of rice to subvert plant immunity. In this study, we demonstrated that M. oryzae MoAo1 is an AO that plays a role in virulence by modulating the apoplast redox status of rice cells. We showed that MoAo1 inhibits the activity of rice OsAO3 and OsAO4, which also regulate the apoplast redox status and plant immunity. In addition, we found that MoAo1, OsAO3, and OsAO4 all exhibit polymorphic variations whose varied interactions orchestrate pathogen virulence and rice immunity. Taken together, our results reveal a critical role for extracellular redox enzymes during rice blast infection and shed light on the importance of the apoplast redox state and its regulation in plant-pathogen interactions.

Powdery mildew is one of the most destructive diseases in wheat production. Identifying novel resistance genes and deploying them in new cultivars is the most effective approach to minimize wheat losses caused by powdery mildew. In this study, wheat breeding line PBDH1607 showed high resistance to powdery mildew at both the seedling and adult plant stages. Genetic analysis of the seedling data demonstrated that the resistance was controlled by a single dominant gene, tentatively designated PmPBDH. The ΔSNP index based on bulked segregant RNA sequencing indicated that PmPBDH was associated with an interval of about 30.8 Mb (713.5 to 744.3 Mb) on chromosome arm 4AL. Using newly developed markers, we mapped PmPBDH to a 3.2-cM interval covering 7.1 Mb (719,055,516 to 726,215,121 bp). This interval differed from those of Pm61 (717,963,176 to 719,260,469 bp), MlIW30 (732,769,506 to 732,790,522 bp), and MlNSF10 (729,275,816 to 731,365,462 bp) reported on the same chromosome arm. PmPBDH also differed from Pm61, MlIW30, and MlNSF10 by its response spectrum, origin, or inheritance mode, suggesting that PmPBDH should be a new Pm gene. In the candidate interval, five genes were found to be associated with PmPBDH via time course gene expression analysis, and thus they are candidate genes of PmPBDH. Six closely linked markers, including two kompetitive allele-specific PCR markers, were confirmed to be applicable for tracking PmPBDH in marker-assisted breeding.

Underground microbial ecosystems have profound impacts on plant health

Trunk and scaffold canker diseases (TSCDs) of almond cause significant yield and tree losses and reduce the lifespan of orchards. In California, several pathogens cause TSCDs, including Botryosphaeriaceae, Ceratocystis destructans, Eutypa lata, Collophorina hispanica, Pallidophorina paarla, Cytospora, Diaporthe, and Phytophthora spp. Field diagnosis of TSCDs is challenging because symptom delineation among the diseases is not clear. Accurate diagnosis of the causal species requires detailed examination of symptoms and subsequent isolation on medium and identification using morphological criteria and subsequent confirmation using molecular tools. The process is time-consuming and difficult, particularly as morphological characteristics are variable and overlap among species. To facilitate diagnosis of TSCD, we developed PCR assays using 23 species-specific primers designed by exploiting sequence differences in the translation elongation factor, β-tubulin, or internal transcribed spacer gene. Using genomic DNA from pure cultures of each fungal and oomycete species, each primer pair successfully amplified a single DNA fragment from the target pathogen but not from selected nontarget pathogens or common endophytes. Although 10-fold serial dilution of fungal DNA extracted from either pure cultures or infected wood samples detected as little as 0.1 pg of DNA sample, consistent detection required 10 ng of pathogen DNA from mycelial samples or from wood chips or drill shavings from artificially or naturally infected almond wood samples with visible symptoms. The new PCR assay represents an improved tool for diagnostic laboratories and will be critical to implement effective disease surveillance and control measures.

Four common fungicidal products were evaluated for their effect on symptoms caused by two nectriaceous black root rot fungi, Calonectria ilicicola and Dactylonectria macrodidyma, when applied as pot drenches to avocado (Persea americana) seedlings in the greenhouse. Applications of fludioxonil, thiophanate-methyl + etridiazole, prochloraz, and prochloraz MnCl at 2 and 4 weeks after inoculation with C. ilicicola significantly reduced root necrosis and improved root and aboveground plant biomass compared with water-treated controls. Fludioxonil reduced necrosis by 60% and had a significantly lower frequency of reisolation of C. ilicicola than the other three fungicide treatments. D. macrodidyma inoculation caused less severe symptoms in seedlings than C. ilicicola despite the longer duration of the trial. Pot drenches with fludioxonil, thiophanate-methyl + etridiazole, and prochloraz MnCl, but not prochloraz alone, significantly reduced root necrosis caused by D. macrodidyma. Prochloraz MnCl was the only fungicide treatment to increase root and plant biomass compared with water-treated controls. Both fludioxonil and prochloraz MnCl reduced the frequency of reisolation of D. macrodidyma from necrotic roots by about 50% compared with the other fungicides or water controls. The results indicated that drenches with these fungicides may suppress existing low to moderate black root rot infection, allowing new root growth and improved establishment in the orchard. Fungicide drenching must not replace best-practice disease management strategies in nurseries but may be a useful tool in crisis situations.