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The LysM proteins have been reported to be important for the virulence and host immunity suppression in herbaceous plant pathogens, whereas far less information is documented in the woody plant pathogen Lasiodiplodia theobromae. To investigate the functional mechanism of LysM protein in L. theobromae, one gene LtScp1 was cloned and characterized detailedly in the current study. Transcription profiling revealed that LtScp1 was highly expressed at the infectious stages. Compared to wild type, overexpression and silencing of LtScp1 in L. theobromae led to significantly increased and decreased lesion areas, respectively. Moreover, LtScp1 was determined to be a secreted protein via a yeast signal peptide trapping system. Interestingly, LtScp1 was confirmed to be modified by the N-glycosylation, which is necessary for the homodimerization of LtScp1 molecules. Furthermore, it was found that LtScp1 interacted with the grapevine chitinase VvChi4 and interfered the ability of VvChi4 to bind chitin. Collectively, these results suggest that LtScp1 functions as a virulence factor to protect the fungus from degradation during the infection.

Rice blast, caused by Magnaporthe oryzae, is a destructive disease threatening the production of staple foods worldwide. Quinone outside inhibitors (QoIs) are a group of chemicals exhibiting excellent activity against a majority of plant pathogens, with the disadvantage that pathogens can easily develop resistance to QoIs.Here, we investigated the activity of picoxystrobin against M. oryzae, which showed a great inhibitory effect on 100 strains of M. oryzae with half-maximal effective concentrations (ECOur findings suggested that M. oryzae had a mid to high risk of resistance to picoxystrobin. Considering this, we should be vigilant to the resistance risk and apply picoxystrobin sensibly in the field. © 2022 Society of Chemical Industry.

The cyclic adenosine monophosphate (cAMP) signalling pathway plays an important role in the regulation of the development and pathogenicity of filamentous fungi. cAMP-dependent protein kinase A (PKA) is the conserved element downstream of cAMP, and its diverse mechanisms in multiple filamentous fungi are not well known yet. In the present study, gene knockout mutants of two catalytic subunits of PKA (PKA-C) in Setosphaeria turcica were created to illustrate the regulatory mechanisms of PKA-Cs on the development and pathogenicity of S. turcica. As a result, StPkaC2 was proved to be the main contributor of PKA activity in S. turcica. In addition, it was found that both StPkaC1 and StPkaC2 were necessary for conidiation and invasive growth, while only StPkaC2 played a negative role in the regulation of filamentous growth. We reveal that only StPkaC2 could interact with the transcription factor StEfg1, and it inhibited the transcription of StRAB1, a Rab GTPase homologue coding gene in S. turcica, whereas StPkaC1 could specifically interact with a transcriptional regulator StFlo8, which could rescue the transcriptional inhibition of StEfg1 on StRAB1. We also demonstrated that StRAB1 could positively influence the biosynthesis of chitin in hyphae, thus changing the filamentous growth. Our findings clarify that StPkaC2 participates in chitin biosynthesis to modulate mycelium development by targeting the Efg1-mediated transcription of StRAB1, while StFlo8, interacting with StPkaC1, acts as a negative regulator during this process.

The complex environments of plants force them to prioritize their immune responses to stimuli occurring simultaneously, including colonization by microbes or nutrient availability. Little is known about how the interplay between endophytes and nutrient status affects the immune responses of both plants and fungi. We primarily monitored immune responses in rice following inoculation with the endophytic fungus Phomopsis liquidambaris under different nitrogen (N) conditions. Ph. liquidambaris promoted plant growth under low N (LN) conditions, concomitant with higher root colonization. Plant production of oxidative signals, including hydrogen peroxide and nitric oxide, was activated by Ph. liquidambaris colonization under LN conditions, while salicylic acid (SA) was maintained at high levels and was involved in controlling rice-fungal interactions. High N (HN) conditions enhanced the ability of Ph. liquidambaris in suppressing plant cell death and the ability of roots to degrade Ph. liquidambaris cell walls. Furthermore, under both LN and HN conditions, the activity of plant defence-associated enzymes and fungal antioxidases was not affected in the interactive association. Our data reveal the alteration of plant immunity, including oxidative signalling and plant cell death, by fungal colonization in response to external N conditions and identify SA signalling as a potential controller for rice-Ph. liquidambaris interaction.

The K homology (KH) repeat is an RNA-binding motif that exists in various proteins, some of which participate in plant growth. However, the function of KH domain-containing proteins in plant defence is still unclear. In this study, we found that a KH domain-containing protein in apple (Malus domestica), HEN4-like (MdKRBP4), is involved in the plant immune response. Silencing of MdKRBP4 compromised reactive oxygen species (ROS) production and enhanced the susceptibility of apple to Valsa mali, whereas transient overexpression of MdKRBP4 stimulated ROS accumulation in apple leaves, indicating that MdKRBP4 is a positive immune regulator. Additionally, MdKRBP4 was proven to interact with the VmEP1 effector secreted by V. mali, which led to decreased accumulation of MdKRBP4. Coexpression of MdKRBP4 with VmEP1 inhibited cell death and ROS production induced by MdKRBP4 in Nicotiana benthamiana. These results indicate that MdKRBP4 functions as a novel positive regulatory factor in plant immunity in M. domestica and is a virulence target of the V. mali effector VmEP1.

This study aimed to develop a novel technique to retain and stabilize compounds contributing to truffle aroma by encapsulation using β-cyclodextrin. Two experiments were conducted. In the first experiment, the key volatile profile and microbial population of products resulting from three different encapsulation methods, namely direct mixing method (M1), direct mixing followed by ethanol addition method (M2), and paste method (M3), were compared with untreated truffles (positive control) over a 90-day period. The M2-derived product was the least optimal for retaining key volatile compounds despite showing the lowest microbial population. There was no significant difference in the volatile profile of products derived from M1 and M3 on day 0. However, it was observed that the M3-derived product could retain its volatile profile better than the M1-derived product by day 90. M3 was compared with freeze-drying in the second experiment. Freeze-dried truffles showed an overall higher relative percentage of volatiles than the M3-derived product on day 0. However, by day 90, some volatile changes occurred in the freeze-dried truffles but not in the M3-derived product. The findings indicate that while freeze-drying could adequately conserve truffle volatiles, the encapsulation of volatile compounds in β-cyclodextrin could improve the volatile stability of truffle products and allow for longer storage times. Microbes were found in all encapsulated truffle products and freeze-dried truffles on days 0 and 90, suggesting the need to explore the possibility of incorporating a decontamination step in the process prior to either encapsulation or freeze-drying. PRACTICAL APPLICATION: A technique to capture and stabilize compounds responsible for truffle aroma by encapsulation using β-cyclodextrin was developed and compared with freeze-drying in this study. The overall finding suggests that while freeze-drying of truffle could sufficiently preserve volatiles, encapsulating truffle volatiles with β-cyclodextrin may improve its stability, extending its shelf life, which can be applied in the development of a natural truffle ingredient that can be applied in food product development.

The necrotrophic fungus Ascochyta rabiei causes Ascochyta blight (AB) disease in chickpea. A. rabiei infects all aerial parts of the plant, which results in severe yield loss. At present, AB disease occurs in most chickpea-growing countries. Globally increased incidences of A. rabiei infection and the emergence of new aggressive isolates directed the interest of researchers toward understanding the evolution of pathogenic determinants in this fungus. In this review, we summarize the molecular and genetic studies of the pathogen along with approaches that are helping in combating the disease. Possible areas of future research are also suggested.kingdom Mycota, phylum Ascomycota, class Dothideomycetes, subclass Coelomycetes, order Pleosporales, family Didymellaceae, genus Ascochyta, species rabiei.A. rabiei survives primarily on Cicer species.A. rabiei infects aboveground parts of the plant including leaves, petioles, stems, pods, and seeds. The disease symptoms first appear as watersoaked lesions on the leaves and stems, which turn brown or dark brown. Early symptoms include small circular necrotic lesions visible on the leaves and oval brown lesions on the stem. At later stages of infection, the lesions may girdle the stem and the region above the girdle falls off. The disease severity increases at the reproductive stage and rounded lesions with concentric rings, due to asexual structures called pycnidia, appear on leaves, stems, and pods. The infected pod becomes blighted and often results in shrivelled and infected seeds.Crop failures may be avoided by judicious practices of integrated disease management based on the use of resistant or tolerant cultivars and growing chickpea in areas where conditions are least favourable for AB disease development. Use of healthy seeds free of A. rabiei, seed treatments with fungicides, and proper destruction of diseased stubbles can also reduce the fungal inoculum load. Crop rotation with nonhost crops is critical for controlling the disease. Planting moderately resistant cultivars and prudent application of fungicides is also a way to combat AB disease. However, the scarcity of AB-resistant accessions and the continuous evolution of the pathogen challenges the disease management process.https://www.ndsu.edu/pubweb/pulse-info/resourcespdf/Ascochyta%20blight%20of%20chickpea.pdf https://saskpulse.com/files/newsletters/180531_ascochyta_in_chickpeas-compressed.pdf http://www.pulseaus.com.au/growing-pulses/bmp/chickpea/ascochyta-blight http://agriculture.vic.gov.au/agriculture/pests-diseases-and-weeds/plant-diseases/grains-pulses-and-cereals/ascochyta-blight-of-chickpea http://www.croppro.com.au/crop_disease_manual/ch05s02.php https://www.northernpulse.com/uploads/resources/722/handout-chickpeaascochyta-nov13-2011.pdf http://oar.icrisat.org/184/1/24_2010_IB_no_82_Host_Plant https://www.crop.bayer.com.au/find-crop-solutions/by-pest/diseases/ascochyta-blight.

The effectiveness of metam potassium, 1,3-dichloropropene, chloropicrin, and different ratios of 1,3-dichloropropene and chloropicrin on the reduction of natural and artificial inoculum of Macrophomina phaseolina were investigated in laboratory and field experiments. Additionally, a multivariate meta-analysis with data from six field trials conducted in Florida from 2012 to 2018 was performed.In small-plot field experiments using drip stakes, the highest rate (468 L ha1,3-dichloropropene alone and in mixture with chloropicrin were more effective in reducing inoculum of M. phaseolina than chloropicrin alone, indicating the fungicidal efficacy of 1,3-dichloropropene. Formulation with higher 1,3-dichloropropene concentration performed better than the formulation with higher chloropicrin concentration in field trials. Metam potassium was effective when applied at the highest rate, but with limited lateral movement perpendicular to the drip irrigation line. © 2022 Society of Chemical Industry.

The cell wall provides a crucial barrier to stress imposed by the external environment. In the rice blast fungus Magnaporthe oryzae, this stress response is mediated by the cell wall integrity (CWI) pathway, consisting of a well-characterized protein phosphorylation cascade. However, other regulators that maintain CWI phosphorylation homeostasis, such as protein phosphatases (PPases), remain unclear. Here, we identified two PPases, MoPtc1 and MoPtc2, that function as negative regulators of the CWI pathway. MoPtc1 and MoPtc2 interact with MoMkk1, one of the key components of the CWI pathway, and are crucial for the vegetative growth, conidial formation, and virulence of M. oryzae. We also demonstrate that both MoPtc1 and MoPtc2 dephosphorylate MoMkk1 in vivo and in vitro, and that CWI stress leads to enhanced interaction between MoPtc1 and MoMkk1. CWI stress abolishes the interaction between MoPtc2 and MoMkk1, providing a means of deactivation for CWI signalling. Our studies reveal that CWI signalling in M. oryzae is a highly coordinated regulatory mechanism vital for stress response and pathogenicity.

Verticillium dahliae is a notorious soil-borne pathogen that enters hosts through the roots and proliferates in the plant water-conducting elements to cause Verticillium wilt. Historically, Verticillium wilt symptoms have been explained by vascular occlusion, due to the accumulation of mycelia and plant biomacromolecule aggregation, and also by phytotoxicity caused by pathogen-secreted toxins. Beyond the direct cytotoxicity of some members of the secretome, this review systematically discusses the roles of the V. dahliae secretome in vascular occlusion, including the deposition of polysaccharides as an outcome of plant cell wall destruction, the accumulation of fungal mycelia, and modulation of plant defence responses. By modulating plant defences and hormone levels, the secretome manipulates the vascular environment to induce Verticillium wilt. Thus, the secretome of V. dahliae colludes with plant defence responses to modulate Verticillium wilt symptoms, and thereby bridges the historical concepts of both toxin production by the pathogen and vascular occlusion as the cause of wilting symptoms.

Sclerotinia sclerotiorum is a devastating pathogen that infects a broad range of host plants. The mechanism underlying plant defence against fungal invasion is still not well characterized. Here, we report that ANGUSTIFOLIA (AN), a CtBP family member, plays a role in the defence against S. sclerotiorum attack. Arabidopsis an mutants exhibited stronger resistance to S. sclerotiorum at the early stage of infection than wild-type plants. Accordingly, an mutants exhibited stronger activation of pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) responses, including mitogen-activated protein kinase activation, reactive oxygen species accumulation, callose deposition, and the expression of PTI-responsive genes, upon treatment with PAMPs/microbe-associated molecular patterns. Moreover, Arabidopsis lines overexpressing AN were more susceptible to S. sclerotiorum and showed defective PTI responses. Our luminometry, bimolecular fluorescence complementation, coimmunoprecipitation, and in vitro pull-down assays indicate that AN interacts with allene oxide cyclases (AOC), essential enzymes involved in jasmonic acid (JA) biosynthesis, negatively regulating JA biosynthesis in response to S. sclerotiorum infection. This work reveals AN is a negative regulator of the AOC-mediated JA signalling pathway and PTI activation.

The soilborne ascomycete fungus Verticillium dahliae causes destructive vascular wilt disease in hundreds of dicotyledonous plant species. However, our understanding of the early invasion from the epidermis to the vasculature and the prompt proliferation and colonization in the xylem tissues remains poor. To elaborate the detailed infection strategy of V. dahliae in host plants, we traced the whole infection process of V. dahliae by live-cell imaging combined with high-resolution scanning electron microscopy. The 4D image series demonstrated that the apex of invading hyphae becomes tapered and directly invades the intercellular space of root epidermal cells at the initial infection. Following successful epidermal invasion, the invading hyphae extend in the intercellular space of the root cortex toward the vascular tissues. Importantly, the high-resolution microscopic and live-cell images demonstrated (a) that conidia are formed via budding at the apex of the hyphae in the xylem vessels to promote systemic propagation vertically, and (b) that the hyphae freely cross adjacent xylem vessels through the intertracheary pits to achieve horizontal colonization. Our findings provide a solid cellular basis for future studies on both intracellular invasion and vascular colonization/proliferation during V. dahliae infection and pathogenesis in host plants.

Mega-fires of unprecedented size, intensity and socio-economic impacts have surged globally due to climate change, fire suppression and development. Soil microbiomes are critical for post-fire plant regeneration and nutrient cycling, yet how mega-fires impact the soil microbiome remains unclear. We had a serendipitous opportunity to obtain pre- and post-fire soils from the same sampling locations after the 2016 Soberanes mega-fire burned with high severity throughout several of our established redwood-tanoak plots. This makes our study the first to examine microbial fire response in redwood-tanoak forests. We re-sampled soils immediately post-fire from two burned plots and one unburned plot to elucidate the effect of mega-fire on soil microbiomes. We used Illumina MiSeq sequencing of 16S and ITS1 sequences to determine that bacterial and fungal richness were reduced by 38%-70% in burned plots, with richness unchanged in the unburned plot. Fire altered composition by 27% for bacteria and 24% for fungi, whereas the unburned plots experienced no change in fungal and negligible change in bacterial composition. Pyrophilous taxa that responded positively to fire were phylogenetically conserved, suggesting shared evolutionary traits. For bacteria, fire selected for increased Firmicutes and Actinobacteria. For fungi, fire selected for the Ascomycota classes Pezizomycetes and Eurotiomycetes and for a Basidiomycota class of heat-resistant Geminibasidiomycete yeasts. We build from Grime's competitor-stress tolerator-ruderal (C-S-R) framework and its recent microbial applications to show how our results might fit into a trait-based conceptual model to help predict generalizable microbial responses to fire.

Brassica napus (oilseed rape, canola) seedling resistance to Leptosphaeria maculans, the causal agent of blackleg (stem canker) disease, follows a gene-for-gene relationship. The avirulence genes AvrLmS and AvrLep2 were described to be perceived by the resistance genes RlmS and LepR2, respectively, present in B. napus 'Surpass 400'. Here we report cloning of AvrLmS and AvrLep2 using two independent methods. AvrLmS was cloned using combined in vitro crossing between avirulent and virulent isolates with sequencing of DNA bulks from avirulent or virulent progeny (bulked segregant sequencing). AvrLep2 was cloned using a biparental cross of avirulent and virulent L. maculans isolates and a classical map-based cloning approach. Taking these two approaches independently, we found that AvrLmS and AvrLep2 are the same gene. Complementation of virulent isolates with this gene confirmed its role in inducing resistance on Surpass 400, Topas-LepR2, and an RlmS-line. The gene, renamed AvrLmS-Lep2, encodes a small cysteine-rich protein of unknown function with an N-terminal secretory signal peptide, which is a common feature of the majority of effectors from extracellular fungal plant pathogens. The AvrLmS-Lep2/LepR2 interaction phenotype was found to vary from a typical hypersensitive response through intermediate resistance sometimes towards susceptibility, depending on the inoculation conditions. AvrLmS-Lep2 was nevertheless sufficient to significantly slow the systemic growth of the pathogen and reduce the stem lesion size on plant genotypes with LepR2, indicating the potential efficiency of this resistance to control the disease in the field.

Crustose Verrucariaceae lichens form a distinctive black belt on seashores all over the world. This lifestyle is apparently enabled by a specific set of photobionts. However, their diversity is understudied. We sampled these lichens from the northern Patagonian Pacific coast of Chile. Using molecular markers, we identified both mycobionts and photobionts. The lichens, belonging to the genus Hydropunctaria and to the Wahlenbergiella group, hosted solely Ulvophycean photobionts. Pseudendoclonium submarinum (Kornmanniaceae, Ulvales) was the most common, but representatives of other closely related, yet undescribed, lineages were also found. Undulifilum symbioticum gen. et sp. nov. is described within Kornmanniaceae based on culture morphology and DNA sequence data. Furthermore, the free-living macroscopic genus Urospora (Acrosiphoniaceae, Ulotrichales) is reported as a lichen photobiont for the first time and is the first of its kind in the order. These results indicate that undescribed algal diversity is waiting to be uncovered in seashore lichens.

The diversity and ecological contribution of the fungus kingdom in the marine environment remain understudied. A recent survey in the Atlantic (Woods Hole, MA, USA) brought to light the diversity and unique biological features of marine fungi. The study revealed that black yeast species undergo an unconventional cell division cycle, which has not been documented in conventional model yeast species such as Saccharomyces cerevisiae (budding yeast) and Schizosaccharomyces pombe (fission yeast). The prevalence of this unusual property is unknown. Here, I collected and identified 65 marine fungi species across 40 genera from the surface ocean water, sediment, and the surface of macroalgae (seaweeds) in the Pacific (Sugashima, Toba, Japan). The Sugashima collection largely did not overlap with the Woods Hole collection and included several unidentifiable species, further illustrating the diversity of marine fungi. Three black yeast species were isolated, two of which were commonly found in Woods Hole (Aureobasidium pullulans and Hortaea werneckii). Surprisingly, their cell division mode was dependent on cell density, and the previously reported unconventional division mode was reproduced only at a certain cell density. For all three black yeast species, cells underwent filamentous growth with septations at low cell density and immediately formed buds at high cell density. At intermediate cell density, two black yeasts (H. werneckii and an unidentifiable species) showed rod cells undergoing septation at the cell equator. In contrast, all eight budding yeast species showed a consistent division pattern regardless of cell density. This study suggests the plastic nature of the growth/division mode of marine-derived black yeast.

Dragon fruit (Hylocereus polyrhizus) is an economically promising fruit in Bangladesh. The cultivation of dragon fruit has increased fourfold within a decade due to its popularity. Recently, a new disease known as stem canker was reported in some plantations of dragon fruit in Bangladesh, which forced some farmers to abandon their cultivation. This study aimed to explore the morphological, molecular, and cultural characteristics as well as host range of the causal agent associated with this destructive disease. Morphologically similar eight fungal isolates were recovered from eight canker symptomatic dragon fruit stems. Among them, two isolates (namely BU-DLa 01 and BU-DLa 02) were used for a detailed study. Morphological parameters and phylogeny of sequence data of internal transcribed spacer (ITS1, 5.8S rRNA, and ITS2), β-tubulin, and translation elongation factor 1-α identified the isolates as Lasiodiplodia theobromae. The cultural features were studied hinged on the growth of the two isolates on various media, temperature, and pH. Though the mycelial growth of the fungi was supported by all the media tested, potato dextrose agar was the most suitable one for both isolates. The fungi thrived well at a temperature of 25-35°C and 5.5-6.5 pH. Inoculation trials of dragon fruit stem ascertained Koch's postulate. In host range test, the isolates were found pathogenic toward mango, guava, banana, and the fruits of dragon fruit. These data will contribute not only to understanding the biology of L. theobromae as a newly recognized pathogen of H. polyrhizus but also will help in designing a proper management package against this pathogen.

Emerging infectious diseases have resulted in severe population declines across diverse taxa. In some instances, despite attributes associated with high extinction risk, disease emergence and host declines are followed by host stabilisation for unknown reasons. While host, pathogen, and the environment are recognised as important factors that interact to determine host–pathogen coexistence, they are often considered independently. Here, we use a translocation experiment to disentangle the role of host traits and environmental conditions in driving the persistence of remnant bat populations a decade after they declined 70–99% due to white-nose syndrome and subsequently stabilised. While survival was significantly higher than during the initial epidemic within all sites, protection from severe disease only existed within a narrow environmental space, suggesting host traits conducive to surviving disease are highly environmentally dependent. Ultimately, population persistence following pathogen invasion is the product of host–pathogen interactions that vary across a patchwork of environments. Published version

Genome-wide transcriptome analysis provides systems-level insights into plant biology. Due to the limited depth of quantitative proteomics our understanding of gene-protein-complex stoichiometry is largely unknown in plants. Recently, the complexity of the proteome and its cell-/tissue-specific distribution have boosted the research community to the integration of transcriptomics and proteomics landscapes in a proteogenomic approach. Herein, we generated a quantitative proteome and transcriptome abundance atlas of 15 major sweet cherry (Prunus avium L., cv 'Tragana Edessis') tissues represented by 29 247 genes and 7584 proteins. Additionally, 199 984 alternative splicing events, particularly exon skipping and alternative 3' splicing, were identified in 23 383 transcribed regions of the analyzed tissues. Common signatures as well as differences between mRNA and protein quantities, including genes encoding transcription factors and allergens, within and across the different tissues are reported. Using our integrated dataset, we identified key putative regulators of fruit development, notably genes involved in the biosynthesis of anthocyanins and flavonoids. We also provide proteogenomic-based evidence for the involvement of ethylene signaling and pectin degradation in cherry fruit ripening. Moreover, clusters of genes and proteins with similar and different expression and suppression trends across diverse tissues and developmental stages revealed a relatively low RNA abundance-to-protein correlation. The present proteogenomic analysis allows us to identify 17 novel sweet cherry proteins without prior protein-level annotation evidenced in the currently available databases. To facilitate use by the community, we also developed the Sweet Cherry Atlas Database (https://grcherrydb.com/) for viewing and data mining these resources. This work provides new insights into the proteogenomics workflow in plants and a rich knowledge resource for future investigation of gene and protein functions in Prunus species.

Background Phomopsis stem-end rot caused by Diaporthe citri, causes significant commercial postharvest losses of pummelo fruit during storage. Carvacrol (CVR) is a known generally recognized as safe (GRAS) and has the ability to prolong the preservation of harvested fruits. In the present study, the inhibitory effects of CVR treatment at the appropriate concentration on Phomopsis stem-end rot development of harvested pummelo fruit inoculated with D. citri were evaluated by the amounts of cell wall components, the activities and gene expressions of related enzymes involved in cell wall modification and lignin biosynthesis. Results Results indicated that CVR completely inhibited D. citri growth in vitro at 200 mg L-1 and significantly controlled Phomopsis stem-end rot development in harvested pummelo. The CVR treatment delayed peel softening and browning, and retarded electrolyte leakage, O2 •- production, and malondialdehyde content. The CVR-treated fruit maintained higher amounts of cell wall materials (CWM), protopectin, hemicelluloses, and cellulose, but exhibited lower water-soluble pectin (WSP) amount. Moreover, in D. citri-inoculated fruit, CVR treatment suppressed the activities and gene expressions of cell wall disassembling-enzymes, including pectin methylesterase, polygalacturonase, cellulase, and β-galactosidase, while the development of cell wall degradation was reduced. Meanwhile, the CVR treatment enhanced the lignin biosynthesis by increasing the activities and up-regulating the gene expressions of phenylalanine ammonialyase, cinnamic alcohol dehydrogenase, and peroxidase accompanied with elevated level of lignin in pummelo fruit. Conclusion The disease resistance to D. citri in pummelo fruit elicited by CVR treatment is related to delaying cell wall degradation and enhancing lignin biosynthesis. This article is protected by copyright. All rights reserved.

Solid organ transplant recipients are at increased risk of acquiring devastating infections with unusual pathogens. Nocardia are aerobic actinomycetes that affect the lungs, brain, skin and soft tissue. Cladophialophora species are dematiaceous fungi that overwhelmingly cause infections in the brain. Both organisms carry a high mortality rate. We present the first reported renal transplant case with Cladophialophora bantiana involving the renal allograft with concurrent invasive nocardiosis involving the lungs and brain.

PREMISE Populations of species with large spatial distributions are shaped by complex forces that differ throughout their ranges. To maintain the genetic diversity of species, genepool-based subsets of widespread species must be considered in conservation assessments. METHODS The population genetics of the lichenized fungus Lobaria pulmonaria and its algal partner, Symbiochloris reticulata, were investigated using microsatellite markers to determine population structure, genetic diversity, and degree of congruency in eastern and western North America. Data loggers measuring temperature and humidity were deployed at selected populations in eastern North America to test for climatic adaptation. To better understand the role Pleistocene glaciations played in shaping population patterns, a North American, range-wide species distribution model was constructed and hindcast to 22,000 years before present and at 500-year time slices from then to the present. KEY RESULTS The presence of two gene pools with minimal admixture was supported, one in the Pacific Northwest and one in eastern North America. Western populations were significantly more genetically diverse than eastern populations. There was no evidence for climatic adaptation among eastern populations, though there was evidence for range-wide adaptation to evapotranspiration rates. Hindcast distribution models suggest that observed genetic diversity may be due to a drastic Pleistocene range restriction in eastern North America, whereas a substantial coastal refugial area is inferred in the west. CONCLUSIONS Taken together the results show different, complex population histories of L. pulmonaria in eastern and western North America, and suggest that conservation planning for each gene pool should be considered separately. This article is protected by copyright. All rights reserved.

American chestnut (Castanea dentata) is a susceptible host of the invasive necrotrophic fungus Cryphonectria parasitica, which causes chestnut blight disease. The fungal pathogen attacks chestnut stems by invading wounded tissue and secreting oxalate. This process leads to the death of infected host cells and the formation of cankers, eventually girdling stems and killing the tree above the infections. To reduce damage caused by fungal oxalate, American chestnut has been genetically engineered to express a wheat oxalate oxidase (OxO). This enzyme degrades the oxalate produced by the pathogen and confers elevated tolerance to Cryphonectria parasitica infection. We report new lines of transgenic American chestnut that have been developed with the win3.12 inducible promoter from poplar (Populus deltoides) driving OxO expression. This promoter is responsive to both wounding and pathogen infection, with a low level of baseline expression. Targeted expression of OxO to wounded and infected tissue is sought as an alternative to constitutive expression for potential metabolic resource conservation and transgene stability over the long lifetime of a tree and over successive generations of breeding. Transgenic Castanea dentata lines harbouring the win3.12-OxO construct were evaluated for transgene expression patterns and tolerance to chestnut blight infection. OxO transcript levels were low in uninfected plants, but robust infection-induced expression levels were observed, with one transgenic line reaching levels comparable to those of previously characterized CaMV35S-OxO lines. In chestnut blight infection bioassays, win3.12-OxO lines showed elevated disease tolerance similar to blight-resistant Chinese chestnut (Castanea mollissima) controls.

Terpenes constitute a major part of secondary metabolites secreted by plants in the rhizosphere. However, their specific functions in fungal–plant interactions have not been investigated thoroughly. In this study we investigated the role of monoterpenes in interactions between oilseed rape (Brassica napus) and the soilborne pathogen Verticillium longisporum. We identified seven monoterpenes produced by B. napus, and production of α‐pinene, β‐pinene, 3‐carene, and camphene was significantly increased upon fungal infection. Among them, β‐pinene was chosen for further analysis. Transcriptome analysis of V. longisporum on exposure to β‐pinene resulted in identification of two highly expressed pleotropic drug transporters paralog genes named VlAbcG1a and VlAbcG1b. Overexpression of VlAbcG1a in Saccharomyces cerevisiae increased tolerance to β‐pinene, while deletion of the VlAbcG1a homologous gene in Verticillium dahliae resulted in mutants with increased sensitivity to certain monoterpenes. Furthermore, the VlAbcG1a overexpression strain displayed an increased tolerance to β‐pinene and increased virulence in tomato plants. Data from this study give new insights into the roles of terpenes in plant–fungal pathogen interactions and the mechanisms fungi deploy to cope with the toxicity of these secondary metabolites., A pleotropic drug transporter VIAbcG1 is required for tolerance to the plant host β‐pinene monoterpene and virulence in the soilborne plant pathogen Verticillium longisporum.

A rapid biodiversity assessment of insects and associated Laboulbeniales fungi was conducted over the course of five nights in August, 2018, at two central Florida lakes: Lake Eustis and the nearby protected and restored National Natural Landmark, Emeralda Marsh Conservation Area (EMCA), which encompasses a portion of Lake Griffin. Lake Eustis was surveyed for Laboulbeniales in 1897 by mycologist Dr. Roland Thaxter but has not since been investigated. Because Lake Eustis has been urbanized, with the lake perimeter almost entirely altered by human development, the site offers a look into Laboulbeniales diversity across a 121‐year timeline, before and after human development. By surveying Lake Eustis and EMCA, a modern case study comparison of Laboulbeniales and insect diversity between a developed and a protected and restored system is made. A total of 4022 insects were collected during the rapid assessment. Overall, insect abundance was greater at EMCA, with 3001 insects collected, than 1021 insects collected from Eustis. Although family‐level insect richness was comparable between sites, with 55 families present at EMCA and 56 at Eustis, 529 out of 3001 (17.6%) of the insects collected at EMCA were hosts to parasitic Laboulbeniales fungi, whereas only 2 out of 1021 (0.19%) collected from Eustis were infected. A total of 16 species of Laboulbeniales found at EMCA compared with only one at Eustis. The current number of Laboulbeniales species documented at Eustis was incredibly depauperate compared with the 26 species and two varieties recorded by Thaxter in 1897. These findings suggest the possibility of utilizing Laboulbeniales as indicators of ecosystem health, and future research should investigate this question further. A figure displaying host–parasite records and a species list of Laboulbeniales are presented. Finally, updated occurrence records for species of Ceratomyces and Hydrophilomyces are provided., Two lakes in central Florida were surveyed for insects and the associated fungal parasites, Laboulbeniales. One of the lakes had been the site of intensive restoration efforts while the other lake is highly developed and un‐restored. This study found stark differences in fungal diversity between the sites, and explores the possibility of the use of Laboulbeniales fungi as indicators of environmental health.

Emerging infectious diseases pose a major threat to human, animal, and plant health. The risk of species-extinctions increases when pathogens can survive in the absence of the host, for example in environmental reservoirs. However, identifying such reservoirs and modes of infection is often highly challenging. In this study, we investigated the presence and nature of an environmental reservoir for the ascomycete fungus Pseudogymnoascus destructans, the causative agent of white-nose disease. We also characterised the modes and timing of transmission of the pathogen; key elements to better understand the disease dynamics. Using 18 microsatellite markers, we determined the genotypic and genic (based on allele frequencies) differentiation between 1,497 P. destructans isolates collected from nine closely situated hibernacula in North-Eastern Germany. One hibernaculum was the focus of intensive sampling in which both the bats and walls of the site were sampled at regular intervals over five consecutive winter seasons (1,062 isolates). We found significant genic differentiation between sites and few multi-locus genotypes shared across hibernacula (genotypic differentiation). This demonstrates that each hibernaculum has an essentially unique population of the fungus. This would be expected if bats purge viable P. destructans over the summer, preventing the mixing and exchange of the pathogen in maternity colonies, where bats from all of the studied hibernacula meet. Results from the intensively sampled site show higher measures of genotypic richness on walls compared to bats, the absence of genic differentiation between bats and walls, and stable relative abundance of multi-locus genotypes over multiple winter seasons. This clearly implicates hibernacula walls as the main environmental reservoir of the pathogen, from which bats become re-infected annually.

The hemibiotrophic pathogen Bipolaris sorokiniana causes root rot, leaf blotching, and black embryos in wheat and barley worldwide, resulting in significant yield and quality reductions. However, the mechanism underlying the host–pathogen interactions between B. sorokiniana and wheat or barley remains unknown. The B. sorokiniana genome encodes a large number of uncharacterized putative effector proteins. In this study, we identified a putative secreted protein, CsSp1, with a classic N‐terminal signal peptide, that is induced during early infection. A split‐marker approach was used to knock out CsSP1 in the Lankao 9‐3 strain. Compared with the wild type, the deletion mutant ∆Cssp1 displayed less radial growth on potato dextrose agar plates and produced fewer spores, and complementary transformation completely restored the phenotype of the deletion mutant to that of the wild type. The pathogenicity of the deletion mutant in wheat was attenuated even though appressoria still penetrated the host. Additionally, the infectious hyphae in the deletion mutant became swollen and exhibited reduced growth in plant cells. The signal peptide of CsSp1 was functionally verified through a yeast YTK12 secretion system. Transient expression of CsSp1 in Nicotiana benthamiana inhibited lesion formation caused by Phytophthora capsici. Moreover, CsSp1 localized in the nucleus and cytoplasm of plant cells. In B. sorokiniana‐infected wheat leaves, the salicylic acid‐regulated genes TaPAL, TaPR1, and TaPR2 were down‐regulated in the ∆Cssp1 strain compared with the wild‐type strain under the same conditions. Therefore, CsSp1 is a virulence effector and is involved in triggering host immunity., CsSp1, a virulent effector in Bipolaris sorokiniana, also triggers immunity in plant cells.

Background Rice blast disease (Magnaporthe oryzae) is considered the most destructive rice diseases all over the world. Dimetindene maleate is used in medication against allergic reactions in humans. Dimetindene maleate used to induce systemic acquired resistance (SAR) in rice (Oryza sativa L.) in order to protect rice plants from blast disease. Results Dimetindene maleate was not effective against fungus linear growth in vitro. In greenhouse conditions, dimetindene maleate significantly improved resistance with 25, 50, 125, 250, 500, and 1000 mg/L concentrations. Leaf blast severity reached to 14.18% with a most suitable concentration of 125 mg/L compared with untreated check. In field conditions during the both seasons 2016 and 2017, dimetindene maleate at 125 mg/L decreased the disease severity to 1.1 and 2.7%, respectively after 30 days of treatment. Also, grain yield was increased to 13.27 and 12.90 tonnes/hectare, respectively. Moreover, dimetindene maleate induces some of the indicators for salicylic acid (SA) and jasmonic acid (JA) pathways via gene expression. These genes include OsWRKY45, OsNPR1, AOS2, JAMYB, and PBZ1 (OsPR10), recording 15.14, 16.47, 5.3, 5.37, and 5.1 fold changes, respectively, after 12-h inoculation. Conclusion The results overview investigated the effectiveness of dimetindene maleate to increase rice resistance to blast disease through induces SAR in rice plants under greenhouse and field conditions, which could be through SA defense pathway by expression of genes (OsWRKY45 and OsNPR1). This article is protected by copyright. All rights reserved.

Sclerotinia sclerotiorum, the notorious necrotrophic phytopathogenic fungus with wide distribution, is responsible for sclerotium disease in more than 600 plant species, including many economic crops such as soybean, oilseed rape, and sunflower. The compound appressorium is a crucial multicellular infection structure that is a prerequisite for infecting healthy tissues. Previously, the Forkhead‐box family transcription factors (FOX TFs) SsFoxE2 and SsFKH1 were shown to play a key regulatory role in the hyphae growth, sexual reproduction, and pathogenicity of S. sclerotiorum. However, little is known about the roles of SsFoxE3 regulating growth and development and pathogenicity. Here, we report SsFoxE3 contributes to sclerotium formation and deletion of SsFoxE3 leads to reduced formation of compound appressoria and developmental delays. Transcripts of SsFoxE3 were greatly increased during the initial stage of infection and SsFoxE3 deficiency reduced virulence on the host, while stabbing inoculation could partially restore pathogenicity. The SsFoxE3 mutant showed sensitivity to H2O2, and the expression of reactive oxygen species detoxification and autophagy‐related genes were reduced. Moreover, expression of SsAtg8 was also decreased during the infection process of the SsFoxE3 mutant. Yeast 1‐hybrid tests suggested that SsFoxE3 interacted with the promoter of SsAtg8. Disruption of SsAtg8 resulted in a phenotype similar to that of the SsFoxE3 mutant. Comparative analysis of the level of autophagy in the wild type and SsFoxE3 mutant showed that N starvation‐induced autophagy was reduced in the SsFoxE3 mutant. Taken together, our findings indicate that SsFoxE3 plays an important role in compound appressorium formation and is involved in transcriptional activation of SsAtg8 during infection by S. sclerotiorum., Transcription factor SsFoxE3 plays an important role in compound appressorium formation and is involved in transcriptional activation of SsAtg8 during infection by Sclerotinia sclerotiorum.

Viruses can play critical roles in symbioses by initiating horizontal gene transfer, affecting host phenotypes, or expanding their host's ecological niche. However, knowledge of viral diversity and distribution in symbiotic organisms remains elusive. Here we use deep-sequenced metagenomic DNA (PacBio Sequel II; two individuals), paired with a population genomics approach (Pool-seq; 11 populations, 550 individuals) to understand viral distributions in the lichen Umbilicaria phaea. We assess (i) viral diversity in lichen thalli, (ii) putative viral hosts (fungi, algae, bacteria) and (iii) viral distributions along two replicated elevation gradients. We identified five novel viruses, showing 28%-40% amino acid identity to known viruses. They tentatively belong to the families Caulimoviridae, Myoviridae, Podoviridae and Siphoviridae. Our analysis suggests that the Caulimovirus is associated with green algal photobionts (Trebouxia) of the lichen, and the remaining viruses with bacterial hosts. We did not detect viral sequences in the mycobiont. Caulimovirus abundance decreased with increasing elevation, a pattern reflected by a specific algal lineage hosting this virus. Bacteriophages showed population-specific patterns. Our work provides the first comprehensive insights into viruses associated with a lichen holobiont and suggests an interplay of viral hosts and environment in structuring viral distributions.

The fungal pathogen Setosphaeria turcica causes leaf blight on maize, which leads to considerable crop losses. However, how S. turcica establishes sustained systemic infection is largely unknown. Here, we report several novel factors contributing to S. turcica pathogenicity, identified using a genomic and transcriptional screen at different stages of S. turcica appressorium development. We identified two cytoskeleton regulators, SLM1 and SLM2, that are crucial for hypha and appressorium development. The SLM1 and SLM2 transcripts accumulated during germling stage but their levels were notably reduced at the appressorium stage. Deletion of SLM2 dramatically affected cell morphology, penetration ability, and pathogenicity. We also identified three different types of S. turcica glycosyl hydrolases that are critical for plant cell wall degradation. Their transcripts accumulated during the appressorium infection stage induced by cellophane and maize leaf. Most importantly, we characterized a novel and specific S. turcica effector, appressorium‐coupled effector 1 (StACE1), whose expression is coupled to appressorium formation in S. turcica. This protein is required for maize infection and induces cell death on expression in Nicotiana benthamiana. These observations suggest that the phytopathogen S. turcica is primed in advance with multiple strategies for maize infection, which are coupled to appressorium formation at the early infection stages., The phytopathogen Setosphaeria turcica is primed in advance with multiple strategies for maize infection, which are coupled to appressorium formation at the early infection stages.

Conidia of Trichoderma guizhouense (Hypocreales, Ascomycota) are frequently applied to the production of biofertilizers and biocontrol agents. Conidiation of some Trichoderma species depends on blue light and the action of different blue light receptors. However, the interplay between different blue-light receptors in light signalling remained elusive. Here, we studied the functions of the blue light receptors BLR1 and ENV1, and the MAP kinase HOG1 in blue light signalling in T. guizhouense. We found that the BLR1 dominates light responses and ENV1 is responsible for photoadaptation. Genome-wide gene expression analyses revealed that 1615 genes, accounting for ~13.4% of the genes annotated in the genome, are blue-light regulated in T. guizhouense, and remarkably, these differentially expressed genes (DEGs) including 61 transcription factors. BLR1 and HOG1 are the core components of the light signalling network, which control 79.9% and 73.9% of the DEGs respectively. In addition, the strict regulation of hydrophobin production by the blue light signalling network is impressive. Our study unravels the regulatory network based on the blue light receptors and the MAPK HOG pathway for conidiation, hydrophobin production and other processes in T. guizhouense.

Ectomycorrhizal (ECM) fungi associated with plants constitute one of the most successful symbiotic interactions in forest ecosystems. ECM support trophic exchanges with host plants and are important factors for the survival and stress resilience of trees. However, ECM clades often harbour morpho-species and cryptic lineages, with weak morphological differentiation. How this relates to intraspecific genome variability and ecological functioning is poorly known. Here, we analysed 16 European isolates of the ascomycete Cenococcum geophilum, an extremely ubiquitous forest symbiotic fungus with no known sexual or asexual spore-forming structures but with a massively enlarged genome. We carried out whole-genome sequencing to identify single-nucleotide polymorphisms. We found no geographic structure at the European scale but divergent lineages within sampling sites. Evidence for recombination was restricted to specific cryptic lineages. Lineage differentiation was supported by extensive copy-number variation. Finally, we confirmed heterothallism with a single MAT1 idiomorph per genome. Synteny analyses of the MAT1 locus revealed substantial rearrangements and a pseudogene of the opposite MAT1 idiomorph. Our study provides the first evidence for substantial genome-wide structural variation, lineage-specific recombination and low continent-wide genetic differentiation in C. geophilum. Our study provides a foundation for targeted analyses of intra-specific functional variation in this major symbiosis.

Highly reducing polyketide synthases (HR-PKSs) produce structurally diverse polyketides (PKs). The PK diversity is constructed by a variety of factors, including the β-keto processing, chain length, methylation pattern, and relative and absolute configurations of the substituents. We examined the stereochemical course of the PK processing for the synthesis of polyhydroxy PKs such as phialotides, phomenoic acid, and ACR-toxin. Heterologous expression of a HR-PKS gene, a trans-acting enoylreductase gene, and a truncated non-ribosomal peptide synthetase gene resulted in the formation of a linear PK with multiple stereogenic centers. The absolute configurations of the stereogenic centers were determined by chemical degradation followed by comparison of the degradation products with synthetic standards. A stereochemical rule was proposed to explain the absolute configurations of other reduced PKs and highlights an error in the absolute configurations of a reported structure. The present work demonstrates that focused functional analysis of functionally related HR-PKSs leads to a better understanding of the stereochemical course.

Magnaporthe oryzae is an important plant pathogen that causes rice blast. Hse1 and Vps27 are components of ESCRT-0 involved in the multivesicular body (MVB) sorting pathway and biogenesis. To date, the biological functions of ESCRT-0 in M. oryzae have not been determined. In this study, we identified and characterized Hse1 and Vps27 in M. oryzae. Disruption of MoHse1 and MoVps27 caused pleiotropic defects in growth, conidiation, sexual development, and pathogenicity, thereby resulting in loss of virulence in rice and barley leaves. Disruption of MoHse1 and MoVps27 triggered increased lipidation of MoAtg8 and degradation of GFP-MoAtg8, indicating that ESCRT-0 is involved in the regulation of autophagy. ESCRT-0 was determined to interact with coat protein complex II (COPII), a regulator functioning in homeostasis of the endoplasmic reticulum (ER homeostasis), and disruption of MoHse1 and MoVps27 also blocked activation of the unfolded protein response (UPR) and autophagy of the endoplasmic reticulum (ER-phagy). Overall, our results indicate that ESCRT-0 plays critical roles in regulating fungal development, virulence, autophagy, and ER-phagy in M. oryzae. This article is protected by copyright. All rights reserved.

Ectomycorrhiza-associated bacteria, especially endofungal bacterial microbiota (EBM) in the fruiting body, play important roles in driving the establishment and function of ectomycorrhizae. However, the influence of ectomycorrhizal fungus bolete identity on their EBM is still unclear. We analysed the EBM of three different bolete fruiting body species on Thousand Island Lake, including Tylopilus felleus, Tylopilus areolatus and Boletus queletii, and compared them with their corresponding mycosphere soil bacterial microbiota by high-throughput sequencing. The EBM was classified into Bacillus, Pseudomonas, Burkholderia and Stenotrophomonas genera. Proteobacteria, Bacteroidetes and Acidobacteria were predominant in the EBM of bolete fruiting bodies as well as their mycosphere soil, while Firmicutes was significantly higher in the EBM. Moreover, the core microbiome (342 operational taxonomic units) of the EBM was shared among the three bolete fungal species. The relative abundances of gene families related to cell cycle control and nucleotide, coenzyme and lipid metabolism were significantly higher in the EBM than in the corresponding mycosphere soil bacterial microbiota, but there was no difference among the three different boletes. The results suggested that the host identity of ectomycorrhizal fungus boletes could affect the EBM, which might be mainly due to the selection of host fungi for the different functional EBM needed.

Necrosis- and ethylene-inducing peptide 1 (Nep1)-like proteins (NLPs) constitute a superfamily of proteins toxic to dicot plants, but the molecular basis of this toxicity remains obscure. Using quantitative trait locus (QTL) analysis we investigated the genetic variation underlying ion leakage in Arabidopsis plants elicited with MoNLP1 derived from Magnaporthe oryzae. The QTL conditioning MoNLP1 toxicity was positionally cloned and further characterized to elucidate its mode of action. MoNLP1-triggered cell death varied significantly across > 250 Arabidopsis accessions and three QTLs were identified conferring the observed variation. The QTL on chromosome 4 was uncovered to encode a leucine-rich repeat (LRR)-only protein designated as NTCD4, which shares high sequence identity with a set of nucleotide-binding LRR proteins. NTCD4 was secreted into the apoplast and physically interacted with multiple NLPs. Apoplastic NTCD4 facilitated the oligomerization of NLP, which was closely associated with toxicity in planta. The natural genetic variation causing D3N change in NTCD4 reduced the secretion efficiency of NTCD4 and the infection of Botrytis cinerea on Arabidopsis plants. These observations demonstrate that the plant-derived NTCD4 is recruited by NLPs to promote toxicity via facilitating their oligomerization, which extends our understanding of a key step in the toxic mode of action of NLPs.

Summary Seventeen species of fungi belonging to thirteen genera were screened for the ability to carry out the transformation of 7‐oxo‐DHEA (7‐oxo‐dehydroepiandrosterone). Some strains expressed new patterns of catalytic activity towards the substrate, namely 16β‐hydroxylation (Laetiporus sulphureus AM498), Baeyer–Villiger oxidation of ketone in D‐ring to lactone (Fusicoccum amygdali AM258) and esterification of the 3β‐hydroxy group (Spicaria divaricata AM423). The majority of examined strains were able to reduce the 17‐oxo group of the substrate to form 3β,17β‐dihydroxy‐androst‐5‐en‐7‐one. The highest activity was reached with Armillaria mellea AM296 and Ascosphaera apis AM496 for which complete conversion of the starting material was achieved, and the resulting 17β‐alcohol was the sole reaction product. Two strains of tested fungi were also capable of stereospecific reduction of the conjugated 7‐keto group leading to 7β‐hydroxy‐DHEA (Inonotus radiatus AM70) or a mixture of 3β,7α,17β‐trihydroxy‐androst‐5‐ene and 3β,7β,17β‐trihydroxy‐androst‐5‐ene (Piptoporus betulinus AM39). The structures of new metabolites were confirmed by MS and NMR analysis. They were also examined for their cholinesterase inhibitory activity in an enzymatic‐based assay in vitro test.

White mold and stem rot is a common disease of Phaseolus vulgaris caused by Sclerotinia sclerotiorum. Biological control is a promising alternative for the control of this disease. In the present study, two Trichoderma spp., T. erinaceum and T. viride, and the consortium of both were evaluated as biocontrol agents against sclerotinia stem rot disease. The results revealed that T. erinaceum (NAIMCC-F-02171) and T. viride (NAIMCC-F-02500) when applied alone, significantly suppressed the infection rate of S. sclerotiorum and increased the rate of survival of plants by 74.5%. On the contrary, the combination of both the Trichoderma spp. was found to be more effective in reducing stem rot by 57.2% and increasing the survival of plants by 87.5% when compared to the individual Trichoderma applications. Further, the exogenous supplementation of Trichoderma activated antioxidative machineries, such as peroxidase, polyphenol oxidase, superoxide dismutase, catalase, and ascorbic acid in the plant. Besides, hydrogen peroxide and superoxide-free radical accumulation were also found to be reduced when T. erinaceum and T. viride were used either individually or in combination under the pathogen-challenged condition. Additionally, the photopigments in the bioprimed plants were markedly increased. Moreover, the combined inoculation of the two isolates yielded the highest records of growth parameters (root weight, shoot length, and leaf weight) compared with individual inoculation. Therefore, based on the above results, it was concluded that the combination of T. erinaceum and T. viride can be effectively used as an alternative to control white mold and stem rot caused by S. sclerotiorum.

Sclerotinia sclerotiorum infects host plant tissues by inducing necrosis to source nutrients needed for its establishment. Tissue necrosis results from an enhanced generation of reactive oxygen species (ROS) at the site of infection and apoptosis. Pathogens have evolved ROS scavenging mechanisms to withstand host‐induced oxidative damage. However, the genes associated with ROS scavenging pathways are yet to be fully investigated in S. sclerotiorum. We selected the S. sclerotiorum Thioredoxin1 gene (SsTrx1) for our investigations as its expression is significantly induced during S. sclerotiorum infection. RNA interference‐induced silencing of SsTrx1 in S. sclerotiorum affected the hyphal growth rate, mycelial morphology, and sclerotial development under in vitro conditions. These outcomes confirmed the involvement of SsTrx1 in promoting pathogenicity and oxidative stress tolerance of S. sclerotiorum. We next constructed an SsTrx1‐based host‐induced gene silencing (HIGS) vector and mobilized it into Arabidopsis thaliana (HIGS‐A) and Nicotiana benthamiana (HIGS‐N). The disease resistance analysis revealed significantly reduced pathogenicity and disease progression in the transformed genotypes as compared to the nontransformed and empty vector controls. The relative gene expression of SsTrx1 increased under oxidative stress. Taken together, our results show that normal expression of SsTrx1 is crucial for pathogenicity and oxidative stress tolerance of S. sclerotiorum., Normal expression of SsTrx1 is crucial for pathogenicity, sclerotial development, and oxidative stress tolerance of Sclerotinia sclerotiorum.

The functions of ubiquitin‐conjugating enzymes (E2) in plant immunity are not well understood. In this study, OsUBC26, a rice ubiquitin‐conjugating enzyme, was characterized in the defence against Magnaporthe oryzae. The expression of OsUBC26 was induced by M. oryzae inoculation and methyl jasmonate treatment. Both RNA interference lines and CRISPR/Cas9 null mutants of OsUBC26 reduced rice resistance to M. oryzae. WRKY45 was down‐regulated in OsUBC26 null mutants. In vitro E2 activity assay indicated that OsUBC26 is an active ubiquitin‐conjugating enzyme. Yeast two‐hybrid assays using OsUBC26 as bait identified the RING‐type E3 ligase UCIP2 as an interacting protein. Coimmunoprecipitation assays confirmed the interaction between OsUBC26 and UCIP2. The CRISPR/Cas9 mutants of UCIP2 also showed compromised resistance to M. oryzae. Yeast two‐hybrid screening using UCIP2 as bait revealed that APIP6 is a binding partner of UCIP2. Moreover, OsUBC26 working with APIP6 ubiquitinateds AvrPiz‐t, an avirulence effector of M. oryzae, and OsUBC26 null mutation impaired the proteasome degradation of AvrPiz‐t in rice cells. In summary, OsUBC26 plays important roles in rice disease resistance by regulating WRKY45 expression and working with E3 ligases such as APIP6 to counteract the effector protein AvrPiz‐t from M. oryzae., Rice ubiquitin‐conjugating enzyme OsUBC26 is essential for immunity to blast fungus by regulating WRKY45 expression and working with APIP6 to promote the ubiquitination and degradation of AvrPiz‐t.

The loss of function of exocyst subunit EXO70B1 leads to autoimmunity, which is dependent on TIR-NBS2 (TN2), a truncated intracellular nucleotide-binding and leucine-rich repeat receptor (NLR). However, how TN2 triggers plant immunity and whether typical NLRs are required in TN2-activated resistance remain unclear. Through the CRISPR/Cas9 gene editing system and knockout analysis, we found that the spontaneous cell death and enhanced resistance in exo70B1-3 were independent of the full-length NLR SOC3 and its closest homolog SOC3-LIKE 1 (SOC3-L1). Additionally, knocking out SOC3-L1 or TN2 did not suppress the chilling sensitivity conferred by chilling sensitive 1-2 (chs1-2). The ACTIVATED DISEASE RESISTANCE 1 (ADR1) family and the N REQUIREMENT GENE 1 (NRG1) family have evolved as helper NLRs for many typical NLRs. Through CRISPR/Cas9 gene editing methods, we discovered that the autoimmunity of exo70B1-3 fully relied on ADR1s, but not NRG1s, and ADR1s contributed to the upregulation of TN2 transcript levels in exo70B1-3. Furthermore, overexpression of TN2 also led to ADR1-dependent autoimmune responses. Taken together, our genetic analysis highlights that the truncated TNL protein TN2-triggered immune responses require ADR1s as helper NLRs to activate downstream signaling, revealing the importance and complexity of ADR1s in plant immunity regulation.