Incompatible Interactions Research Articles

Impact of Rhg1 copy number variation on a soybean cyst nematode resistance transcriptional network.

Soybean yield loss due to soybean cyst nematode (SCN) infestation has a negative impact on the U.S. economy. Most SCN-resistant soybeans carry a common resistance locus (Rhg1), conferred by copy number variation of a 31.2-kb segment at the Rhg1 locus. To identify the effects of Rhg1 copy number on the plant prior to SCN infection, we investigated genome-wide expression profiles in isogenic Fayette plants carrying different copy numbers at the Rhg1 locus (9-11 copies), that confer different levels of resistance to SCN. We found that even small differences in copy number lead to large changes in expression of downstream defense genes. The co-expression network constructed from differentially expressed genes (DEGs) outside the Rhg1 locus revealed complex effects of Rhg1 copy number on transcriptional regulation involving signal transduction and ethylene-mediated signaling pathways. Moreover, we report a variation in expression levels of phytoalexin biosynthesis-related genes that is correlated with copy number, and the activation of different NBS-LRR gene sets, indicating a broad effect of copy number on defense responses. Using qRT-PCR time series during SCN infection, we validated the SCN responses of DEGs detected in the copy number comparison and showed a stable upregulation of genes related to phytoalexin biosynthesis in resistant Fayette lines during the early stages of the incompatible interaction between soybeans and SCN, before syncytium formation. These results suggest additional genes that could enhance Rhg1-mediated SCN resistance.

Puccinia triticina avirulence protein AvrLr21 directly interacts with wheat resistance protein Lr21 to activate wheat immune response

Wheat leaf rust, caused by Puccinia triticina (Pt), remains a constant threat to wheat production worldwide. Deployment of race-specific leaf rust (Lr) resistance genes in wheat provides effective protection against leaf rust, but often leads to selective pressures that drive the rapid emergence of new virulent Pt isolates in nature. However, the molecular mechanisms underlying the evasion of Lr-delivered resistance by leaf rust remain largely unknown. Here, we identify an avirulence gene AvrLr21 in Pt that triggers Lr21-dependent immune responses. BSMV (Barley stripe mosaic virus)-mediated host-induced gene silencing assay shows that silencing AvrLr21 compromises Lr21-mediated immunity. AvrLr21 interacts directly with Lr21 protein to induce a hypersensitive response in tobacco leaves. The evolved Lr21-breaking Pt isolates can suppress Lr21-mediated immunity. Our data provide a basis for studying the molecular determinants in Pt-wheat incompatible interaction and monitoring natural Pt populations to prioritize the deployment of Lr resistance genes in the field.

The effect of polyvinyl chloride and polyethylene terephthalate on the mechanical properties of butyl rubber (IIR)-based thermoplastic elastomer using semi-efficient vulcanisation system and a nano zinc oxide as activator

Thermoplastic elastomers (TPEs), a class of materials bridging the gap between rubber and thermoplastics, have revolutionised diverse fields with their unique properties and applications. This study highlights the critical importance of compatibility between plastics, polyvinyl chloride (PVC), polyethylene terephthalate (PET), and butyl rubber (isobutylene-isoprene rubber, IIR) components in TPEs for optimal performance. Several TPEs were prepared with different amounts of plastics. Subsequently, the TPEs were cured, and curing and mechanical properties, and correspondent thermal stability and solvent resistance were measured. It revealed that the thermal stability of TPEs, containing PVC, PET and IIR, was influenced by the type and quantity of each plastic, affecting degradation temperatures. PVC and PET exhibited distinct effects on the curing behaviour of IIR-based TPEs, impacting scorch time and optimum cure time differently. The incorporation of PVC and PET enhanced the organic solvent resistance of the TPEs, attributed to physical barrier effects, enhanced crosslinking, increased crystallinity, and specific interfacial interactions. Tensile strength and elongation at break generally decreased with the addition of plastics, reflecting incompatible interactions and increased crystallinity. Meanwhile, hardness increased due to the rigid structure and filler effects of plastics. Achieving an optimal TPE formulation involves balancing these competing effects.

Insights into bs5 resistance mechanisms in pepper against Xanthomonas euvesicatoria through transcriptome profiling

BackgroundBacterial spot of pepper (BSP), caused by four different Xanthomonas species, primarily X. euvesicatoria (Xe), poses a significant challenge in pepper cultivation. Host resistance is considered the most important approach for BSP control, offering long-term protection and sustainability. While breeding for resistance to BSP for many years focused on dominant R genes, introgression of recessive resistance has been a more recent focus of breeding programs. The molecular interactions underlying recessive resistance remain poorly understood.ResultsIn this study, transcriptomic analyses were performed to elucidate defense responses triggered by Xe race P6 infection by two distinct pepper lines: the Xe-resistant line ECW50R containing bs5, a recessive resistance gene that confers resistance to all pepper Xe races, and the Xe-susceptible line ECW. The results revealed a total of 3357 upregulated and 4091 downregulated genes at 0, 1, 2, and 4 days post-inoculation (dpi), with the highest number of differentially expressed genes (DEGs) observed at 2 dpi. Pathway analysis highlighted DEGs in key pathways such as plant-pathogen interaction, MAPK signaling pathway, plant hormone signal transduction, and photosynthesis – antenna proteins, along with cysteine and methionine metabolism. Notably, upregulation of genes associated with PAMP-Triggered Immunity (PTI) was observed, including components like FLS2, Ca-dependent pathways, Rboh, and reactive oxygen species (ROS) generation. In support of these results, infiltration of ECW50R leaves with bacterial suspension of Xe led to observable hydrogen peroxide accumulation without a rapid increase in electrolyte leakage, suggestive of the absence of Effector-Triggered Immunity (ETI). Furthermore, the study confirmed that bs5 does not disrupt the effector delivery system, as evidenced by incompatible interactions between avirulence genes and their corresponding dominant resistant genes in the bs5 background.ConclusionOverall, these findings provide insights into the molecular mechanisms underlying bs5-mediated resistance in pepper against Xe and suggest a robust defense mechanism in ECW50R, primarily mediated through PTI. Given that bs5 provides early strong response for resistance, combining this resistance with other dominant resistance genes will enhance the durability of resistance to BSP.

Strain-tuned incompatible magnetic exchange-interaction in La2NiO4

Magnetic frustration is a route for novel ground states, including spin liquids and spin ices. Such frustration can be introduced through either lattice geometry or incompatible exchange interactions. Here, we find that epitaxial strain is an effective tool for tuning antiferromagnetic exchange interactions in a square-lattice system. By studying the magnon excitations in La2NiO4 films using resonant inelastic x-ray scattering, we show that the magnon displays substantial dispersion along the antiferromagnetic zone boundary, at energies that depend on the lattice of the film’s substrate. Using first principles simulations and an effective spin model, we demonstrate that the antiferromagnetic next-nearest neighbour coupling is a consequence of the two-orbital nature of La2NiO4. Altogether, we illustrate that compressive epitaxial strain enhances this coupling and, as a result, increases the level of incompatibility between exchange interactions within a model square-lattice system.

Quantitative phosphoproteomics reveals molecular pathway network in wheat resistance to stripe rust

Protein phosphorylation plays an important role in immune signaling transduction in plant resistance to pathogens. Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), severely devastates wheat production. Nonetheless, the molecular mechanism of wheat resistance to stripe rust remains limited. In this study, quantitative phosphoproteomics was employed to investigate the protein phosphorylation changes in wheat challenged by Pst. A total of 1537 and 2470 differentially accumulated phosphoproteins (DAPs) were identified from four early infection stage (6, 12, 18 and 24 h post-inoculation) in incompatible and compatible wheat-Pst interactions respectively. KEGG analysis revealed that Oxidative Phosphorylation, Phosphatidylinositol Signaling, and MAPK signaling processes are distinctively enriched in incompatible interaction, while Biosynthesis of secondary metabolites and RNA degradation process were significantly enriched in compatible interactions. In particular, abundant changes in phosphorylation levels of chloroplast proteins were identified, suggesting the regulatory role of photosynthesis in wheat-Pst interaction, which is further emphasized by protein-protein interaction (PPI) network analysis. Motif-x analysis identified [xxxxSPxxxx] motif, likely phosphorylation sites for defensive response-related kinases, and a new [xxxxSSxxxx] motif significantly enriched in incompatible interaction. The results shed light on the early phosphorylation events contributing to wheat resistance against Pst. Moreover, our study demonstrated that the phosphorylation levels of Nucleoside diphosphate kinase TaNAPK1 are upregulated at 12 hpi with CYR23 and at 24 hpi with CYR31. Transient silencing of TaNAPK1 was able to attenuate wheat resistance to CYR23 and CYR31. Our study provides new insights into the mechanisms underlying Pst-wheat interactions and may provide database to find potential targets for the development of new resistant varieties.

Changes in RNase activities and in expression of two RNase genes in powdery mildew infected barley, wheat and Brachypodium distachyon leaves

AbstractChanges in RNase activities were investigated in extracts from barley near isogenic lines without or with various powdery mildew resistance genes and were compared to changes in wheat and Brachypodium distachyon leaves after powdery mildew infections. In barley, the compatible interaction with powdery mildew induced the highest increase in RNase activity as measured spectrophotometrically. The incompatible interaction that accompanied with hypersensitive reaction in Mla leaves gave less increase, whereas incompatible interactions in Mlg and mlo barley leaves without visible symptoms gave the least increase of RNase activity. In wheat, the largest RNase activity was found in leaves infected with the compatible wheat powdery mildew or wheat stem and leaf rusts. RNase activity in B. distachyon was higher than that in healthy wheat and especially barley leaves. The electrophoretic RNase enzyme activity patterns were different in barley, wheat and B. distachyon plants, but showed similar activities as determined spectrophotometrically. Barley genes encoding endonuclease 2 and ribonuclease 3-like protein X3 showed the highest expression in the compatible barley - barley powdery mildew interaction as measured by RT-qPCR. This correlated with RNase activities in leaf extracts suggesting that RNases in barley and wheat may act as susceptibility factors of powdery mildew and rust diseases.

Infection of Alfalfa Cotyledons by an Incompatible but Not a Compatible Species of Colletotrichum Induces Formation of Paramural Bodies and Secretion of EVs.

Hemibiotrophic fungi in the genus Colletotrichum employ a biotrophic phase to invade host epidermal cells followed by a necrotrophic phase to spread through neighboring mesophyll and epidermal cells. We used serial block face-scanning electron microscopy (SBF-SEM) to compare subcellular changes that occur in Medicago sativa (alfalfa) cotyledons during infection by Colletotrichum destructivum (compatible on M. sativa) and C. higginsianum (incompatible on M. sativa). Three-dimensional reconstruction of serial images revealed that alfalfa epidermal cells infected with C. destructivum undergo massive cytological changes during the first 60 h following inoculation to accommodate extensive intracellular hyphal growth. Conversely, inoculation with the incompatible species C. higginsianum resulted in no successful penetration events and frequent formation of papilla-like structures and cytoplasmic aggregates beneath attempted fungal penetration sites. Further analysis of the incompatible interaction using focused ion beam-scanning electron microscopy (FIB-SEM) revealed the formation of large multivesicular body-like structures that appeared spherical and were not visible in compatible interactions. These structures often fused with the host plasma membrane, giving rise to paramural bodies that appeared to be releasing extracellular vesicles (EVs). Isolation of EVs from the apoplastic space of alfalfa leaves at 60 h postinoculation showed significantly more vesicles secreted from alfalfa infected with incompatible fungus compared with compatible fungus, which in turn was more than produced by noninfected plants. Thus, the increased frequency of paramural bodies during incompatible interactions correlated with an increase in EV quantity in apoplastic wash fluids. Together, these results suggest that EVs and paramural bodies contribute to immunity during pathogen attack in alfalfa. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Exploring Potential Surrogate Systems for Studying the Early Steps of the Sporisorium scitamineum Pathogenesis.

Despite its global importance as a primary source of table sugar and bioethanol, sugarcane faces a significant threat to its production due to diseases. One of these diseases, sugarcane smut, involves the emergence of a whip-like structure from the host apical shoot. The slow onset of this pathogenesis is the most substantial challenge for researchers to investigate the molecular events leading to resistance or susceptibility. In this study, we explored the early interaction between the smut fungus Sporisorium scitamineum and foliar tissues of the model plants Arabidopsis thaliana and Nicotiana benthamiana. Upon inoculation with the fungus, A. thaliana showed a compatible reaction, producing lesions during fungus colonization, whereas N. benthamiana showed signs of nonhost resistance. In addition, we propose a sugarcane detached leaf assay using plants cultivated in vitro to reveal sugarcane smut response outcomes. We used two sugarcane genotypes with known contrasting reactions to smut in the field. Although there is no evidence of sugarcane smut fungus infecting host leaves naturally, the sugarcane detached leaf assay enabled a rapid assessment of disease outcomes. Different symptoms in the detached leaves after inoculation distinguished smut-susceptible and smut-resistant sugarcane genotypes. Microscopic observations and gene expression analysis of S. scitamineum candidate effectors confirmed the fungal growth and its restriction on the compatible and incompatible interactions, respectively. These findings offer new prospects into the disease phenotyping of S. scitamineum, which could greatly expedite the comprehension of the initial stages of the pathogenesis and predict smut resistance in sugarcane genotypes.

Alterations in Gene Expression during Incompatible Interaction between Amendoim Cavalo Common Bean and Colletotrichum lindemuthianum.

Anthracnose, caused by the fungus Colletotrichum lindemuthianum, poses a significant and widespread threat to the common bean crop. The use of plant genetic resistance has proven to be the most effective strategy for managing anthracnose disease. The Amendoim Cavalo (AC) Andean cultivar has resistance against multiple races of C. lindemuthianum, which is conferred by the Co-AC gene. Fine mapping of this resistance gene to common bean chromosome Pv01 enabled the identification of Phvul.001G244300, Phvul.001G244400, and Phvul.001G244500 candidate genes for further validation. In this study, the relative expression of Co-AC candidate genes was assessed, as well as other putative genes in the vicinity of this locus and known resistance genes, in the AC cultivar following inoculation with the race 73 of C. lindemuthianum. Gene expression analysis revealed significantly higher expression levels of Phvul.001G244500. Notably, Phvul.001G244500 encodes a putative Basic Helix-Loop-Helix transcription factor, suggesting its involvement in the regulation of defense responses. Furthermore, a significant modulation of the expression of defense-related genes PR1a, PR1b, and PR2 was observed in a time-course experiment. These findings contribute to the development of improved strategies for breeding anthracnose-resistant common bean cultivars, thereby mitigating the impact of this pathogen on crop yields and ensuring sustainable bean production.

Unravelling the role of key genes involved coffee leaf rust resistance

The biotrophic fungus Hemileia vastatrix is the pathogen responsible for coffee leaf rust, a devastating disease in several coffee-producing countries. Despite the importance of studying the interaction between Coffea and H. vastatrix, a more comprehensive understanding of the mechanisms involved in this pathosystem is necessary. The role of eight candidate genes was analyzed aiming at identifying and validating new important coffee resistance genes and understanding their interaction with H. vastatrix. These genes were identified in the most important sources of coffee resistance, the Híbrido de Timor CIFC 832/2 and CIFC 832/1. Previous works found six resistance genes and, in our research, other two new genes were identified in BAC clones and validated by RT-qPCR during compatible and incompatible interactions between Coffea and H. vastatrix. An interactome approach was performed using Coffea-H. vastatrix and Coffea-Coffea proteins to better understand the biological process and the interaction of the host-pathogen. Two networks of interactions from the compiled data were built focused on candidate genes associated with pre-haustorial resistance (12 and 24 h.a.i) in coffee plants against the pathogen. The results showed, for the first time, differentially expressed proteins (DEPs) positively regulated in the incompatible interaction Coffea-H. vastatrix. These coffee proteins interact with each other and with secreted and/or transmembrane pathogen proteins. The obtained results also show that DEPs found are involved in important plant defense pathways such as pathways associated with the response to wounds, signaling, regulation of the innate immune response and the transmembrane receptor protein serine/threonine kinase pathway. The present work shows the involvement of genes in both pathogen recognition and signaling cascades, which act in pre-haustorial defense mechanisms of HdT coffee. Therefore, the candidate genes analyzed, together with the biological processes elucidated, have the potential to contribute to the development of new control strategies against the fungus H. vastatrix within coffee breeding programs aiming to develop cultivars with durable resistance.

De novo assembly-based transcriptome analysis of resistant and susceptible potato varieties to Phytophthora infestans

AbstractAn effective tool for discovering differentially expressed genes (DEGs) related to late blight (LB) resistance is the transcriptome sequencing of potatoes. The aim of this study was to compare transcriptome expression analysis in incompatible and compatible interactions via high-throughput sequencing. Furthermore, we performed a bioinformatics analysis to screen a large number of specific transcription factors (TFs) and DEGs linked to Phytophthora infestans infection. Two locally cultivated potato varieties were chosen from evaluation assays conducted in two consecutive seasons and based on the disease severity (DS) values. These varieties were the highly resistant Jelly (HR) to P. infestans and the moderately susceptible Annabelle (MS). Ribonucleic acid-sequencing (RNA-seq) was achieved for the two varieties with their controls through the BGISEQ-500 sequencing platform. The RNA-seq analysis identified P. infestans-responsive genes and their expression in potatoes. The mechanism of the response of these cultivars to the P. infestans pathogen by TFs and DEG genes, which play an important role in defense response, was investigated. The Gene Ontology (GO) analysis classified 46,248 unigenes in the HR and 26,921 unigenes in MS into the following three categories: biological process, cellular component, and molecular functions. More genes were responsible for the cellular component category, biological process, and molecular functions in HR compared to MS. Moreover, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the significantly enriched DEGs were included in the plant–pathogen interaction, biosynthesis of secondary metabolites, and ribosome. In addition, 1874 transcription factor genes belonging to 85 families were indicated in the DEGs, of which MYB and AP2-EREBP genes were the most abundant. Besides, multiple genes related to LB resistance showed differential expression during infection. It also sheds light on the molecular mechanisms behind potato resistance to P. infestans infection.

OsATL32 ubiquitinates the reactive oxygen species-producing OsRac5-OsRbohB module to suppress rice immunity.

Ubiquitination-mediated protein degradation is integral to plant immunity, with E3 ubiquitin ligases acting as key factors in this process. Here, we report the functions of OsATL32, a plasma membrane-localized Arabidopsis Tóxicos En Levadura (ATL)-type E3 ubiquitin ligase, in rice (Oryza sativa) immunity and its associated regulatory network. We found that the expression of OsATL32 is downregulated in both compatible and incompatible interactions between rice and the rice blast fungus Magnaporthe oryzae. The OsATL32 protein level declines in response to infection by a compatible M. oryzae strain or to chitin treatment. OsATL32 negatively regulates rice resistance to blast and bacterial leaf blight diseases, as well as chitin-triggered immunity. Biochemical and genetic studies revealed that OsATL32 suppresses pathogen-induced reactive oxygen species (ROS) accumulation by mediating ubiquitination and degradation of the ROS-producing OsRac5-OsRbohB module, which enhances rice immunity against M. oryzae. The protein phosphatase PHOSPHATASE AND TENSIN HOMOLOG enhances rice blast resistance by dephosphorylating OsATL32 and promoting its degradation, preventing its negative effect on rice immunity. This study provides insights into the molecular mechanism by which the E3 ligase OsATL32 targets a ROS-producing module to undermine rice immunity.

Global changes in gene expression during compatible and incompatible interactions of faba bean (Vicia faba L.) during Orobanche foetida parasitism.

Orobanche foetida Poiret is the main constraint facing faba bean crop in Tunisia. Indeed, in heavily infested fields with this parasitic plant, yield losses may reach 90%, and the recent estimation of the infested area is around 80,000 ha. Identifying genes involved in the Vicia faba/O. foetida interaction is crucial for the development of effective faba bean breeding programs. However, there is currently no available information on the transcriptome of faba bean responding to O. foetida parasitism. In this study, we employed RNA sequencing to explore the global gene expression changes associated with compatible and incompatible V. faba/O. foetida interactions. In this perspective, two faba bean varieties (susceptible and resistant) were examined at the root level across three stages of O. foetida development (Before Germination (BG), After Germination (AG) and Tubercule Stage (TS)). Our analyses presented an exploration of the transcriptomic profile, including comprehensive assessments of differential gene expression and Gene Ontology (GO) enrichment analyses. Specifically, we investigated key pathways revealing the complexity of molecular responses to O. foetida attack. In this study, we detected differential gene expression of pathways associated with secondary metabolites: flavonoids, auxin, thiamine, and jasmonic acid. To enhance our understanding of the global changes in V. faba response to O. foetida, we specifically examined WRKY genes known to play a role in plant host-parasitic plant interactions. Furthermore, considering the pivotal role of parasitic plant seed germination in this interaction, we investigated genes involved in the orobanchol biosynthesis pathway. Interestingly, we detected the gene expression of VuCYP722C homolog, coding for a key enzyme involved in orobanchol biosynthesis, exclusively in the susceptible host. Clearly, this study enriches our understanding of the V. faba/O. foetida interaction, shedding light on the main differences between susceptible and resistant faba bean varieties during O. foetida infestation at the gene expression level.

Photosynthetic Costs and Impact on Epidemiological Parameters Associated with Ht Resistance Genes in Maize Lines Infected with Exserohilum turcicum.

Northern corn leaf blight, caused by Exserohilum turcicum, is mainly controlled by the use of resistant cultivars. Maize lines carrying individual resistance genes B37Ht1, B37Ht2, B37Ht3, and B37Htn1 express different defense symptoms having an impact on the photosynthetic activity, the accumulation of reactive oxygen species, and epidemiological parameters. Plants were inoculated with a race 0 isolate of E.turcicum conferring a compatible interaction with B37 and incompatible interactions with plants carrying resistance genes. Five days postinoculation (dpi), the resistant lines displayed a reduction in leaf CO2 assimilation of 30 to 80% compared with healthy plants. At 14 dpi, inoculated plants of B37Ht1 showed a significant decrease in leaf CO2 assimilation, similar to B37 (up to 94%). The instantaneous carboxylation efficiency was significantly reduced on inoculated plants of the lines B37Ht2, B37Ht3, and B37Htn1 (54 to 81%) at 5 dpi. Curiously, the reduction in carboxylation efficiency for B37 and B37Ht1 (up to 95%) was higher at 14 dpi than at 5 dpi (up to 81%). At 6 dpi, low levels of H2O2 were detected in B37Ht1, in contrast to B37Htn1, where a high H2O2 level and peroxidase activity were observed. The sporulation rate on B37Ht1, B37Ht3, and B37Htn1 decreased by 92% compared with the susceptible control, whereas strong sporulation occurred in lesions on line B37Ht2. The resistance in maize to E. turcicum conferred by Ht resistance genes is associated with photosynthetic costs and may have quite contrasting effects on host physiology and major epidemiological parameters, such as sporulation, which contributes inoculum for secondary infections.

Wide transcriptional outlook to uncover Penicillium expansum genes underlying fungal incompatible infection

Pathogenesis of P. expansum involved different processes and one of them is the recognition between pathogen-host, which in the case of P. expansum is preferably pome fruit. In this work, the possible mechanisms connected to host recognition are addressed through the generation of a subtractive library carried out during the incompatible P. expansum-orange interaction in the initial stages of infection. The generated library was analyzed by massive sequencing and bioinformatic analysis. Of the identified genes, a total of 24 were selected for subsequent expression analysis by RT-qPCR in two incompatible interaction situations. The characterization of the overexpressed genes revealed the presence of CWDEs, ATPases, aldolases, detoxifying enzymes and virulent determinants that could act as effectors related to fungal virulence independently of the host. However, several identified genes, which could not be associated with the virulence of P. expansum under compatible conditions, were related to enzymes to obtain the nutrients necessary for the growth and development of the pathogen under stress conditions through basal metabolism that contributes to expand the range of adaptation of the pathogen to the environment and different hosts.

TaCRT3 Is a Positive Regulator of Resistance to Blumeria graminis f. sp. tritici in Wheat.

Wheat powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is one of the most prevalent diseases of wheat worldwide and can lead to severe yield reductions. Identifying genes involved in powdery mildew resistance will be useful for disease resistance breeding and control. Calreticulin (CRT) is a member of multigene family widely found in higher plants and is associated with a variety of plant physiological functions and defense responses. However, the role of CRT in wheat resistance to powdery mildew remains unclear. TaCRT3 was identified from the proteomic sequence of an incompatible interaction between the wheat (Triticum aestivum) cultivar Xingmin 318 and the Bgt isolate E09. Following analysis of transient expression of the GFP-TaCRT3 fusion protein in Nicotiana benthamiana leaves, TaCRT3 was localized in the nucleus, cytoplasm, and cell membrane. Transcript expression levels of TaCRT3 were significantly upregulated in the wheat-Bgt incompatible interaction. More critically, knockdown of TaCRT3 using virus-induced gene silencing resulted in attenuated resistance to Bgt in wheat. Histological analysis showed a significant increase in Bgt development in TaCRT3-silenced plants, whereas the pathogen-related gene was significantly downregulated in TaCRT3-silenced leaves. In addition, overexpression of TaCRT3 in wheat enhanced the resistance to powdery mildew, the growth of Bgt was significantly inhibited, and the area of H2O2 near the infection site and the expression of defense-related genes of the salicylic acid pathway significantly increased. These findings imply that TaCRT3 may act as a disease resistance factor that positively regulates resistance to powdery mildew, during which SA signaling is probably activated.

Heat Pre-Treatment Modified Host and Non-Host Interactions of Powdery Mildew with Barley Brassinosteroid Mutants and Wild Types.

High temperatures associated with climate change may increase the severity of plant diseases. This study investigated the effect of heat shock treatment on host and non-host barley powdery mildew interactions using brassinosteroid (BR) mutants of barley. Brassinosteroids are plant steroid hormones, but so far little is known about their role in plant-fungal interactions. Wild type barley cultivar Bowman and its near-isogenic lines with disturbances in BR biosynthesis or signalling showed high compatibility to barley powdery mildew race A6, while cultivar Delisa and its BR-deficient mutants 522DK and 527DK were fully incompatible with this pathogen (host plant-pathogen interactions). On the other hand, Bowman and its mutants were highly resistant to wheat powdery mildew, representing non-host plant-pathogen interactions. Heat pre-treatment induced shifts in these plant-pathogen interactions towards higher susceptibility. In agreement with the more severe disease symptoms, light microscopy showed a decrease in papillae formation and hypersensitive response, characteristic of incompatible interactions, when heat pre-treatment was applied. Mutant 527DK, but not 522DK, maintained high resistance to barley powdery mildew race A6 despite heat pre-treatment. By 10 days after heat treatment and infection, a noticeable shift became apparent in the chlorophyll a fluorescence and in various leaf reflectance parameters at all genotypes.

Pathogenically altered Colletotrichum lindemuthianum transformants help in understanding the biochemical defense and colonization dynamics in Phaseolus vulgaris

Colletotrichum lindemuthianum poses a serious threat to common bean production and impacts food security. It is essential to comprehend the intricate nature of the pathogen infection process and the underlying biochemical systems regulating plant defence to create tactics that effectively increase plant resistance. This study leveraged a stable C. lindemuthianum transformant library to examine these intrinsic mechanisms in a susceptible Phaseolus vulgaris cultivar, Jawala. Among the 57 mitotically stable transformants employed to screen for altered virulence, 51 were pathogenically similar to wild type and the remaining six transformants were either reduced or impaired. Nine transformants (three pathogenically similar and all impaired and reduced mutants) were selected for subsequent studies. These mutants exhibited an array of morpho-cultural variations. The biochemical studies of these mutants revealed free radicals as double-edged swords, exerting a pivotal influence on the infection process. The free radical (H2O2) elevated at an early stage (24 hpi) in the cases of reduced (CT-51) and impaired (CT-225) mutants. However, at the later stage (72 hpi), H2O2 was promoted in the case of R-2047 and equal to wild type (CT-101) transformant. Contrary, superoxide (O2−) increased considerably at 48 hpi in the incompatible interaction of reduced (CT-51) and impaired (CT-225) mutants. The microscopic images revealed how easily the wild type (CT-101) could germinate, penetrate, and infect the leaves. The pathogenically reduced transformant (CT-51), however, had delayed germination as well as a decreased growth rate and spore production. Additionally, the pathogenicity of the impaired transformant (CT-225) was completely lost, and its conidia failed to properly germinate. This is the first account of C. lindemuthianum in planta colonization with a GFP tag. This study increased our understanding of the mechanics of the reactions triggered by C. lindemuthianum and may be used to develop novel strategies for the effective control of bean anthracnose. Additionally, identifying the genes disrupted by the insertion of Agrobacterium T-DNA will help in identifying the genes necessary for bean plant colonization.

A rapid molecular diagnostic tool to discriminate alleles of avirulence genes and haplotypes of Phytophthora sojae using high-resolution melting analysis.

Effectors encoded by avirulence genes (Avr) interact with the Phytophthora sojae resistance gene (Rps) products to generate incompatible interactions. The virulence profile of P. sojae is rapidly evolving as a result of the large-scale deployment of Rps genes in soybean. For a successful exploitation of Rps genes, it is recommended that soybean growers use cultivars containing the Rps genes corresponding to Avr genes present in P. sojae populations present in their fields. Determination of the virulence profile of P. sojae isolates is critical for the selection of soybean cultivars. High-resolution melting curve (HRM) analysis is a powerful tool, first applied in medicine, for detecting mutations with potential applications in different biological fields. Here, we report the development of an HRM protocol, as an original approach to discriminate effectors, to differentiate P. sojae haplotypes for six Avr genes. An HRM assay was performed on 24 P. sojae isolates with different haplotypes collected from soybean fields across Canada. The results clearly confirmed that the HRM assay discriminated different virulence genotypes. Moreover, the HRM assay was able to differentiate multiple haplotypes representing small allelic variations. HRM-based prediction was validated by phenotyping assays. This HRM assay provides a unique, cost-effective and efficient tool to predict virulence pathotypes associated with six different Avr (1b, 1c, 1d, 1k, 3a and 6) genes from P. sojae, which can be applied in the deployment of appropriate Rps genes in soybean fields.