Blumeria Research Articles

Genetic diversity of barley accessions from East Asian countries in terms of resistance to powdery mildew

Background. Barley is an ancient crop currently cultivated in more than 100 countries. High adaptability makes it possible to cultivate it in environments unfavorable for many other crops. At the same time, fungal diseases can have a negative impact on its grain yield and quality. One of the most harmful is the fungus Blumeria graminis (DC.) Golovin ex Speer f. sp. hordei Marchal. The pathogen can overcome a cultivar’s resistance, resulting in significant harvest losses. There is a need for a constant search for new effective sources of resistance to powdery mildew for barley breeding.Materials and methods. A set of 950 barley accessions from the East Asian center of the crop’s morphogenesis were studied at the adult stage of plant development. They were screened and crossed in the fields of Pushkin and Pavlovsk Laboratories of VIR (St. Petersburg, Russia). Genetic control of barley resistance to B. graminis was analyzed in a climate chamber under artificial infection pressure with the northwestern population of the fungus. Scoring scales were used for screening.Results and conclusions. Weak development of B. graminis during the heading phase was observed on plants of 38 barley accessions from the East Asian center. High resistance at all stages of development was manifested by 20 genotypes that can be used in breeding programs. Resistance genes in 18 selected forms differed in their effectiveness at the seedling stage of development and during the heading phase. Accessions k-3433, k-10931, k-10934, k-11608, k-17545, k-20272, k-20279, k-20354 and k-27867 had one dominant resistance gene each. Accessions k-11608, k-12278, and k-17545 are protected by the identical gene, while the resistance of k-3433 is induced by a gene that differs from the genes present in accessions k-10931 and k-20279.

Barley MLA3 recognizes the host-specificity effector Pwl2 from Magnaporthe oryzae.

Plant nucleotide-binding leucine-rich repeat (NLRs) immune receptors directly or indirectly recognize pathogen-secreted effector molecules to initiate plant defense. Recognition of multiple pathogens by a single NLR is rare and usually occurs via monitoring for changes to host proteins; few characterized NLRs have been shown to recognize multiple effectors. The barley (Hordeum vulgare) NLR gene Mildew locus a (Mla) has undergone functional diversification, and the proteins encoded by different Mla alleles recognize host-adapted isolates of barley powdery mildew (Blumeria graminis f. sp. hordei [Bgh]). Here, we show that Mla3 also confers resistance to the rice blast fungus Magnaporthe oryzae in a dosage-dependent manner. Using a forward genetic screen, we discovered that the recognized effector from M. oryzae is Pathogenicity toward Weeping Lovegrass 2 (Pwl2), a host range determinant factor that prevents M. oryzae from infecting weeping lovegrass (Eragrostis curvula). Mla3 has therefore convergently evolved the capacity to recognize effectors from diverse pathogens.

Resistance to Powdery Mildew Is Conferred by Different Genetic Loci at the Adult-Plant and Seedling Stages in Winter Wheat Line Tianmin 668.

Winter wheat line Tianmin 668 was crossed with susceptible cultivar Jingshuang 16 to develop 216 recombinant inbred lines (RILs) for dissecting its adult-plant resistance (APR) and all-stage resistance (ASR) against powdery mildew. The RIL population was genotyped on a 16K genotyping by target sequencing single-nucleotide polymorphism array and phenotyped in six field trials and in the greenhouse. Three loci-QPmtj.caas-2BL, QPmtj.caas-2AS, and QPmtj.caas-5AL-conferring APR to powdery mildew were detected on chromosomes 2BL, 2AS, and 5AL, respectively, of Tianmin 668. The effect of resistance to powdery mildew for QPmtj.caas-2BL was greater than that of the other two loci. A Kompetitive allele-specific PCR marker specific for QPmtj.caas-2BL was developed and verified on 402 wheat cultivars or breeding lines. Results of virulence and avirulence patterns to 17 Blumeria graminis f. sp. tritici isolates, bulked segregant analysis-RNA-sequencing, and a genetic linkage mapping identified a resistance allele at locus Pm4 in Tianmin 668 based on the seedling phenotypes of the RIL population. The PCR-based DNA sequence alignment and cosegregation of the functional marker with the phenotypes of the RIL population demonstrated that Pm4d was responsible for the ASR to isolate Bgt1 in Tianmin 668. The dissection of genetic loci for APR and ASR may facilitate the application of Tianmin 668 in developing powdery mildew-resistant wheat cultivars.

A chromosome-scale genome assembly of the grape powdery mildew pathogen Erysiphe necator reveals its genomic architecture and previously unknown features of its biology.

Erysiphe necator is an obligate fungal pathogen that causes grape powdery mildew, globally the most important disease on grapevines. Previous attempts to obtain a quality genome assembly for this pathogen were hindered by its high repetitive DNA content. Here, chromatin conformation capture (Hi-C) with long-read PacBio sequencing was combined to obtain a chromosome-scale assembly and a high-quality annotation for E. necator isolate EnFRAME01. The resulting 81.1 Mb genome assembly is 98% complete and consists of 34 scaffolds, 11 of which represent complete chromosomes. All chromosomes contain large centromeric-like regions and lack synteny to the 11 chromosomes of the cereal PM pathogen Blumeria graminis. Further analysis of their composition showed that repeats and transposable elements (TEs) occupy 62.7% of their content. TEs were almost evenly interspersed outside centromeric and telomeric regions and massively overlapped with regions of annotated genes, suggesting that they could have a significant functional impact. Abundant gene duplicates were observed as well, particularly in genes encoding candidate secreted effector proteins. Moreover, younger in age gene duplicates exhibited more relaxed selection pressure and were more likely to be located physically close in the genome than older duplicates. A total of 122 genes with copy number variations among six isolates of E. necator were also identified and were enriched in genes that were duplicated in EnFRAME01, indicating they may reflect an adaptive variation. Taken together, our study illuminates higher-order genomic architectural features of E. necator and provides a valuable resource for studying genomic structural variations in this pathogen. IMPORTANCE Grape powdery mildew caused by the ascomycete fungus Erysiphe necator is economically the most important and recurrent disease in vineyards across the world. The obligate biotrophic nature of E. necator hinders the use of typical genetic methods to elucidate its pathogenicity and adaptation to adverse conditions, and thus comparative genomics has been a major method to study its genome biology. However, the current reference genome of E. necator isolate C-strain is highly fragmented with many non-coding regions left unassembled. This incompleteness prohibits in-depth comparative genomic analyses and the study of genomic structural variations (SVs) that are known to affect several aspects of microbial life, including fitness, virulence, and host adaptation. By obtaining a chromosome-scale genome assembly and a high-quality gene annotation for E. necator, we reveal the organization of its chromosomal content, unearth previously unknown features of its biology, and provide a reference for studying genomic SVs in this pathogen.

Identification of Quantitative Trait Loci Associated with Powdery Mildew Resistance in Spring Barley under Conditions of Southeastern Kazakhstan.

Barley (Hordeum vulgare L.) is one of the most produced cereal crops in the world. It has traditionally been used for the production of animal feed and for malting, as well as for human consumption. However, its production is highly affected by biotic stress factors, particularly the fungal pathogen Blumeria graminis (DC.) f. sp. hordei (Bgh), which causes powdery mildew (PM). In this study, a collection of 406 barley accessions from the USA, Kazakhstan, Europe, and Africa were assessed for resistance to PM over a 3-year period in southeastern Kazakhstan. The collection was grown in the field in 2020, 2021, and 2022 and was genotyped using the 9K SNP Illumina chip. A genome-wide association study (GWAS) was conducted to identify the quantitative trait loci (QTLs) associated with PM resistance. As a result, seven QTLs for PM resistance were detected on chromosomes 4H, 5H, and 7H (FDR p-values < 0.05). Genetic positions of two QTLs were similar to those of PM resistance QTLs previously reported in the scientific literature, suggesting that the five remaining QTLs are novel putative genetic factors for the studied trait. Haplotype analysis for seven QTLs revealed three haplotypes which were associated with total PM resistance and one haplotype associated with the high PM severity in the barley collection. Identified QTLs and haplotypes associated with the PM resistance of barley may be used for further analysis, trait pyramiding, and marker-assisted selection.

Wheat Susceptibility Genes TaCAMTA2 and TaCAMTA3 Negatively Regulate Post-Penetration Resistance against Blumeria graminis forma specialis tritici.

Blumeria graminis forma specialis tritici (B.g. tritici) is the airborne fungal pathogen that causes powdery mildew disease on hexaploid bread wheat. Calmodulin-binding transcription activators (CAMTAs) regulate plant responses to environments, but their potential functions in the regulation of wheat-B.g. tritici interaction remain unknown. In this study, the wheat CAMTA transcription factors TaCAMTA2 and TaCAMTA3 were identified as suppressors of wheat post-penetration resistance against powdery mildew. Transient overexpression of TaCAMTA2 and TaCAMTA3 enhanced the post-penetration susceptibility of wheat to B.g. tritici, while knockdown of TaCAMTA2 and TaCAMTA3 expression using transient- or virus-induced gene silencing compromised wheat post-penetration susceptibility to B.g. tritici. In addition, TaSARD1 and TaEDS1 were characterized as positive regulators of wheat post-penetration resistance against powdery mildew. Overexpressing TaSARD1 and TaEDS1 confers wheat post-penetration resistance against B.g. tritici, while silencing TaSARD1 and TaEDS1 enhances wheat post-penetration susceptibility to B.g. tritici. Importantly, we showed that expressions of TaSARD1 and TaEDS1 were potentiated by silencing of TaCAMTA2 and TaCAMTA3. Collectively, these results implicated that the Susceptibility genes TaCAMTA2 and TaCAMTA3 contribute to the wheat-B.g. tritici compatibility might via negative regulation of TaSARD1 and TaEDS1 expression.

Wheat Class III Peroxidase TaPOD70 Is a Potential Susceptibility Factor Negatively Regulating Wheat Resistance to Blumeria graminis f. sp. tritici.

Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is one of the most important diseases on wheat worldwide and can lead to a large reduction in wheat production. Class III peroxidases (PODs), a kind of secretory enzyme and members of a multigene family in higher plants, have been linked to various plant physiological functions and defensive responses. However, the role of PODs in wheat resistance to Bgt remains unclear. TaPOD70, a class III POD gene, was identified from the proteomics sequencing of the incompatible interaction between wheat (Triticum aestivum) cultivar Xingmin 318 and Bgt isolate E09. After transient expression of the TaPOD70-GFP fusion protein in Nicotiana benthamiana leaves, TaPOD70 was located in the membrane region. Yeast secretion assay showed that TaPOD70 was a secretory protein. Furthermore, Bax-induced programmed cell death was inhibited by transient expression of TaPOD70 in N. benthamiana. The transcript expression level of TaPOD70 was significantly upregulated in the wheat-Bgt compatible interaction. More crucially, knocking down TaPOD70 using virus-induced gene silencing increased wheat resistance to Bgt compared with the control plants. In response to Bgt, histological analyses indicated that hyphal development of Bgt was significantly reduced, whereas H2O2 production was enhanced in TaPOD70-silenced leaves. These findings imply that TaPOD70 may act as a susceptibility factor, adversely regulating wheat resistance to Bgt.

Effective sources of powdery mildew resistance among spring bread wheat for the northwest of the russian federation

Background. Bread wheat (Triticum aestivum L.) is one of the world’s main food crops. In Russia, its gross harvest in recent years has reached 21.079 million tons. Blumeria graminis is one of the most harmful diseases of bread wheat. Annual harvest losses can reach 10–30%. In Leningrad Province, powdery mildew appears almost every year on cereals. Development of cultivars resistant to powdery mildew is the most important trend in spring bread wheat breeding. Its progress is determined by new sources of effective resistance genes and their incorporation into promising spring bread wheat cultivars. The purpose of this work was to retest the seedling and adult resistance to powdery mildew in spring bread wheat accessions selected over a period of more than twenty years and redefine the effectiveness of disease resistance sources at the present time.Materials and methods. Since 2000, VIR has conducted a search for powdery mildew resistance among 1283 spring bread wheat accessions. The identified 36 sources of adult and seedling powdery mildew resistance were reevaluated in 2022. Field and laboratory studies into the resistance to the pathogen were performed according to VIR’s guidelines.Results and discussion. In 2022, field and laboratory studies of spring bread wheat cultivars revealed accessions with resistance to the pathogen in all phases of plant development. Six sources remained resistant to powdery mildew for 13–22 years, another six showed resistance for 7–12 years, and 8 cultivars were immune for 6 years. Resistance of 9 accessions identified for this trait in 2020 was confirmed.Conclusion. Field and laboratory research into the resistance of spring bread wheat accessions to the pathogen identified sources retaining seedling and adult resistance to the Leningrad population of powdery mildew for 7 years or more. These accessions are recommended for use in wheat breeding programs.

Pipecolic acid synthesis is required for systemic acquired resistance and plant-to-plant-induced immunity in barley.

Defense responses in plants are based on complex biochemical processes. Systemic acquired resistance (SAR) helps to fight infections by (hemi-)biotrophic pathogens. One important signaling molecule in SAR is pipecolic acid (Pip), accumulation of which is dependent on the aminotransferase ALD1 in Arabidopsis. While exogenous Pip primes defense responses in the monocotyledonous cereal crop barley (Hordeum vulgare), it is currently unclear if endogenous Pip plays a role in disease resistance in monocots. Here, we generated barley ald1 mutants using CRISPR/Cas9, and assessed their capacity to mount SAR. Endogenous Pip levels were reduced after infection of the ald1 mutant, and this altered systemic defense against the fungus Blumeria graminis f. sp. hordei. Furthermore, Hvald1 plants did not emit nonanal, one of the key volatile compounds that are normally emitted by barley plants after the activation of SAR. This resulted in the inability of neighboring plants to perceive and/or respond to airborne cues and prepare for an upcoming infection, although HvALD1 was not required in the receiver plants to mediate the response. Our results highlight the crucial role of endogenous HvALD1 and Pip for SAR, and associate Pip, in particular together with nonanal, with plant-to-plant defense propagation in the monocot crop barley.

Silencing of the calcium-dependent protein kinase TaCDPK27 improves wheat resistance to powdery mildew

BackgroundCalcium ions (Ca2+), secondary messengers, are crucial for the signal transduction process of the interaction between plants and pathogens. Ca2+ signaling also regulates autophagy. As plant calcium signal-decoding proteins, calcium-dependent protein kinases (CDPKs) have been found to be involved in biotic and abiotic stress responses. However, information on their functions in response to powdery mildew attack in wheat crops is limited.ResultIn the present study, the expression levels of TaCDPK27, four essential autophagy-related genes (ATGs) (TaATG5, TaATG7, TaATG8, and TaATG10), and two major metacaspase genes, namely, TaMCA1 and TaMCA9, were increased by powdery mildew (Blumeria graminis f. sp. tritici, Bgt) infection in wheat seedling leaves. Silencing TaCDPK27 improves wheat seedling resistance to powdery mildew, with fewer Bgt hyphae occurring on TaCDPK27-silenced wheat seedling leaves than on normal seedlings. In wheat seedling leaves under powdery mildew infection, silencing TaCDPK27 induced excess contents of reactive oxygen species (ROS); decreased the activities of superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT); and led to an increase in programmed cell death (PCD). Silencing TaCDPK27 also inhibited autophagy in wheat seedling leaves, and silencing TaATG7 also enhanced wheat seedling resistance to powdery mildew infection. TaCDPK27-mCherry and GFP-TaATG8h colocalized in wheat protoplasts. Overexpressed TaCDPK27-mCherry fusions required enhanced autophagy activity in wheat protoplast under carbon starvation.ConclusionThese results suggested that TaCDPK27 negatively regulates wheat resistance to PW infection, and functionally links with autophagy in wheat.

Inoculum threshold for stripe rust infection in wheat

Stripe rust (Puccinia striiformis f. sp. tritici (Pst)) and powdery mildew (Blumeria graminis f. sp. tritici (Bgt)) are important diseases of wheat in Canada and worldwide. Molecular detection methods permit spore detection of few spores; therefore, there is a need to determine initial inoculum thresholds for pathogens to cause disease under both controlled environments and in the field. Susceptible wheat cultivars ‘Avocet’ and ‘AC Barrie’ were inoculated with different quantities of spores (0, 103, 104, 105, 106, and 107) of Pst and Bgt. Disease incidence, severity and infection type were evaluated. Results of controlled environment studies showed that the minimum number of spores necessary to cause appreciable incidence and severity for Pst was at higher spore concentrations of 105–106 spores. Conversely, low incidence and severity levels were observed at 103–104 spores for Bgt. Despite occurrence of natural Pst infection, results of field studies in 2016 and 2017 in Southern Alberta demonstrated that significant increases in severity levels were observed following application of 1.2 × 107 spores. Collectively, these results demonstrated that stripe rust severities increased with increasing spore concentration only at high spore levels. In contrast, Bgt severity increased with spore concentration from 103 to 107 spores mL−1. In vitro and in vivo spore germination tests demonstrated germination rates of Pst spores were reduced at lower spore concentrations compared to germination rates at the higher concentrations. Understanding of minimum spore numbers required for disease development will be a prerequisite for predicting epidemics and devising fungicide control measures for future sustainable agricultural systems.

TaSYP137 and TaVAMP723, the SNAREs Proteins from Wheat, Reduce Resistance to Blumeria graminis f. sp. tritici

SNARE protein is an essential factor driving vesicle fusion in eukaryotes. Several SNAREs have been shown to play a crucial role in protecting against powdery mildew and other pathogens. In our previous study, we identified SNARE family members and analyzed their expression pattern in response to powdery mildew infection. Based on quantitative expression and RNA-seq results, we focused on TaSYP137/TaVAMP723 and hypothesized that they play an important role in the interaction between wheat and Blumeria graminis f. sp. Tritici (Bgt). In this study, we measured the expression patterns of TaSYP132/TaVAMP723 genes in wheat post-infection with Bgt and found that the expression pattern of TaSYP137/TaVAMP723 was opposite in resistant and susceptible wheat samples infected by Bgt. The overexpression of TaSYP137/TaVAMP723 disrupted wheat's defense against Bgt infection, while silencing these genes enhanced its resistance to Bgt. Subcellular localization studies revealed that TaSYP137/TaVAMP723 are present in both the plasma membrane and nucleus. The interaction between TaSYP137 and TaVAMP723 was confirmed using the yeast two-hybrid (Y2H) system. This study offers novel insights into the involvement of SNARE proteins in the resistance of wheat against Bgt, thereby enhancing our comprehension of the role of the SNARE family in the pathways related to plant disease resistance.

Virulence variability and genetic diversity in Blumeria graminis f. sp. hordei in southeastern and southwestern China.

Powdery mildew is a key airborne foliar disease of barley in southeastern and southwestern China. Barley varieties usually partially or wholly lose resistance to the pathogen Blumeria graminis (DC.) f. sp. hordei (Bgh) three to five years after release due to the frequent acquirements of new virulences in the pathogen population. However, no Bgh virulence detection has been carried out in the recent decade, and thus no information is available on the present virulence components and major pathotypes in epidemic regions. Twenty-one near-isogenic lines (NILs) of Pallas were selected to detect Bgh virulence variation, with 97 pathotypes identified from the isolates collected from 2015 to 2019. The virulence complexities ranged from one to 12, with 1.5 isolates on average assigned per pathotype, suggesting a natural trait of high pathotype diversity and low virulence complexity (VC) in the Chinese Bgh populations. Eleven high virulence pathotypes were detected in the traditional barley-growing regions in Yunnan and Zhejiang. Six virulent pathotypes to resistance gene mlo-5 were detected only in the two traditional epidemic regions, with a virulence frequency (VF) of 4.8% (7/147). Compared to the results from a decade ago, VFs for resistance alleles Mla3, mlo-5, Mla6 + Mla14, Mla7 + Mlk, Mlg + MlCP, and Mla13 + MlRu3 + MlaRu4 increased from zero to 0.7 to 25.8%. Isolates from Yunnan and Zhejiang had similar virulence profiles, which differed from those identified in Tibet. In addition, genetic diversities differed in the isolate groups collected from Tibet, Yunnan, and Zhejiang.

Effectiveness of fungicidal protection of spring wheat against powdery mildew and tan spot

The results of the study of the effectiveness of plant protection products against powdery mildew (Blumeria graminis) and tan spot (Pyrenophora tritici-repentis) for spring wheat are presented. It has been noted that the probability of mass destruction of crops by phytopathogens is determined by the stock of the infectious beginning of pathogens, the susceptibility of the host plant and the favorable weather conditions for their development and spread. The experiment was carried out using the methods generally accepted in the Russian Federation. During the observation period, the degree of plant damage of more than 20% in the heading phase was noted in 46% of the years of observation (in 2009, 2013, 2014, 2016, 2017). Epiphytotic development of powdery mildew together with brown leaf rust (Puccinia triticina Eriks.) was registered in 2013–2014 and 2016–2017. For 2009–2019 the degree of plant damage of more than 20% in the heading phase was noted in 46% of the years of observation (in 2009, 2013, 2014, 2016, 2017). Effective protection of wheat against powdery mildew (75–87%) was provided by fungicides based on 2–3 active ingredients. The biological effectiveness of the biofungicide (Bacillus subtilis) was manifested only in the years with moderate wheat damage and did not exceed the average level of 58%. It is more expedient to carry out operational control of yellow spotting of wheat leaves with preparations based on such active substances as "azoxystrobin + epoxiconazole", "tebuconazole + propiconazole". The level of statistically significant yield preserved due to fungicidal protection during the years of depression was 5–6%, with a moderate development of leaf diseases due to chemical fungicides - 18%, and the biological product provided an increase in productivity of 9%. During the years of epiphytoties, the chemical protection of crops retained an average of 24% of the wheat yield, polycomponent preparations had the best performance and stability of action, and the biological protection of plants under these conditions was ineffective.

Discovery of HNPC-A9229: A Novel Pyridin-2-yloxy-Based Pyrimidin-4-amine Fungicide.

To search for novel pesticides, the synthesis around commercialized insecticide tebufenpyrad accidentally led us to the discovery of the fungicidal lead compound, 3-ethyl-1-methyl-N-((2-phenylthiazol-4-yl)methyl)-1H-pyrazole-5-carboxamide (1a) and its pyrimidin-4-amine-based optimization derivative 5-chloro-2,6-dimethyl-N-(1-(2-(p-tolyl)thiazol-4-yl)ethyl)pyrimidin-4-amine (2a). Compound 2a not only demonstrates fungicidal activity superior to commercial fungicides such as diflumetorim but also exhibits the good features that come with pyrimidin-4-amines, such as unique modes of action and no cross-resistance to other pesticide classes. However, 2a is highly toxic to rats. Further optimization of 2a by introducing pyridin-2-yloxy substructure finally led to the discovery of 5b5-6 (HNPC-A9229) (5-chloro-N-(1-((3-chloropyridin-2-yl)oxy)propan-2-yl)-6-(difluoromethyl)pyrimidin-4-amine). HNPC-A9229 exhibits excellent fungicidal activities with EC50 values of 0.16 mg/L against Puccinia sorghi and 1.14 mg/L against Erysiphe graminis, respectively. Not only that its fungicidal potency is significantly superior to or comparable to commercial fungicides including diflumetorim, tebuconazole, flusilazole, and isopyrazam, HNPC-A9229 possesses low toxicity to rats.

Haplotype Analysis Sheds Light on the Genetic Evolution of the Powdery Mildew Resistance Locus Pm60 in Triticum Species.

Wheat powdery mildew (Blumeria graminis f. sp. tritici, Bgt, recently clarified as B. graminis s. str.), is one of the most destructive diseases of wheat. Pm60 is a nucleotide-binding leucine-rich repeat (NLR) gene that confers race-specific resistance to Bgt. Allelic variants (Pm60, Pm60a, and Pm60b) were found in Triticum urartu and T. dicoccoides, the wild progenitors of wheat. In the present study, we studied the diversity of the Pm60 locus in a large set of wheat germplasm and found 20 tetraploid wheats harboring the Pm60 alleles, which correspond to three novel haplotypes (HapI-HapIII). HapI (Pm60 allele) and HapII (Pm60a allele) were present in domesticated tetraploid wheats, whereas HapIII (Pm60a allele) was identified in wild tetraploid T. araraticum. A sequence comparison of HapII and HapIII revealed that they differed by three SNPs and a GCC deletion. Results of the phylogenetic analysis revealed that HapII was more closely related to the functional haplotype MlIW172. Infection tests showed that HapII-carrying lines display a partial resistance response to Bgt#GH, while HapI was susceptible. Our results provide insights into the genetic evolution of the Pm60 locus and potential valuable alleles for powdery mildew resistance breeding.

Novel quinolin-2(1H)-one analogues as potential fungicides targeting succinate dehydrogenase: design, synthesis, inhibitory evaluation and molecular modeling.

Succinate dehydrogenase is an important target of fungicides. Succinate dehydrogenase inhibitors (SDHIs) have widely been used to combat destructive plant pathogenic fungi because they possess efficient and broad-spectrum antifungal activities and as well as unique mode of action. The research and development of novel SDHIs have been ongoing. Thirty-six novel quinolin-2(1H)-one derivatives were designed, synthesized and characterized. The single crystal structure of compound 3c was determined through the X-ray diffraction of single crystals. The bioassay results displayed that most compounds had good antifungal activities at 16 μg mL-1 against Rhizoctonia cerealis, Erysiphe graminis, Botrytis cinerea, Penicillium italicum and Phytophthora infestans. Compounds 6o, 6p and 6r had better antifungal activities than the commercialized fungicide pyraziflumid against Botrytis cinerea. Their half maximal effective concentration (EC50 ) values were 0.398, 0.513, 0.205 and 0.706 μg mL-1 , respectively. Moreover, the inhibiting activities of the bioactive compounds were tested against succinate dehydrogenase. The results indicated that they possessed outstanding activities. Compounds 6o, 6p and 6r also exhibited better inhibiting activities than pyraziflumid against succinate dehydrogenase. Their half maximal inhibitory concentration (IC50 ) values were 0.450, 0.672, 0.232 and 0.858 μg mL-1 , respectively. The results of molecular dynamic (MD) simulations indicated that compound 6r displayed stronger affinity to succinate dehydrogenase than pyraziflumid. The results of the present study displayed that quinolin-2(1H)-one derivative could be one scaffold of potential SDHIs and will provide some valuable information for the research and development of new SDHIs. © 2022 Society of Chemical Industry.

Spore-Trapping Device: An Efficient Tool to Manage Fungal Diseases in Winter Wheat Crops.

Leaf airborne diseases cause major shortfalls in agricultural crops. The introduction of technical means can significantly improve early-warning systems for plant diseases as well as provide timely and accurate forecasts. In this paper, we aim to evaluate the possibilities of detecting a phytopathogenic infection using a spore-catching device developed at the Federal Research Center of Biological Plant Protection (FRCBPP) on winter wheat varieties of different levels of susceptibility to major economically important leaf diseases, taking into account climatic conditions. The device captures spores in the surface layer of air among crop plants. We conducted research in the experimental fields of FRCBPP in 2019-2021. The objects of the study were four cultivars of winter wheat. They were selected according to the degree of resistance to various leaf diseases. We studied the progress of wheat diseases according to generally accepted international scales the onset of the first manifestations to their maximum development. We studied the aerogenic infection in wheat crops using the FRCBPP developed portable device for determining the infestation of plants. Sampling was carried out in the same period as the visual assessment. The samples were taken in the crops of each variety at five points. The sampling time was one minute. As a result of research on experimental crops of four varieties of winter wheat, we observed the development of such diseases as powdery mildew (Erysiphe graminis), yellow spot (Pyrenophora tritici-repentis), septoria leaf spot (Septoria tritici), yellow (Puccinia striiformis) and brown rust (Puccinia triticina). In a laboratory study of samples under a light microscope, all of the listed pathogens were found except for septoria leaf spot. Two-way analysis of variance confirmed the statistically significant separate and cumulative influence of the cultivar and year factor on winter wheat diseases. A generalized correlation analysis for three growing seasons (2019-2021) showed that an average statistically significant correlation coefficient (0.5-0.6) remains for the total groups for powdery mildew, yellow and brown rust. This indicator for the causative agent of yellow spot was equal to 0.4 with a high level of statistical significance. Thus, we conclude that by using a spore-catching device, it is possible to identify spores of economically significant pathogens in winter wheat crops and predict the further development of pathogens, taking into account the cultivar and annual climate factors.

Identification of Key Modules and Candidate Genes for Powdery Mildew Resistance of Wheat-Agropyron cristatum Translocation Line WAT-2020-17-6 by WGCNA

As one of the serious diseases of wheat, powdery mildew (Blumeria graminis f. sp. tritici) is a long-term threat to wheat production. Therefore, it is of great significance to explore new powdery mildew-resistant genes for breeding. The wild relative species of wheat provide gene resources for resistance to powdery mildew breeding. Agropyron cristatum (2n = 4x = 28, genomes PPPP) is an important wild relative of wheat, carrying excellent genes for high yield, disease resistance, and stress resistance, which can be used for wheat improvement. To understand the molecular mechanism of powdery mildew resistance in the wheat-A. cristatum translocation line WAT2020-17-6, transcriptome sequencing was performed, and the resistance genes were analyzed by weighted gene co-expression network analysis (WGCNA). In the results, 42,845 differentially expressed genes were identified and divided into 18 modules, of which six modules were highly correlated with powdery mildew resistance. Gene ontology (GO) enrichment analysis showed that the six interested modules related to powdery mildew resistance were significantly enriched in N-methyltransferase activity, autophagy, mRNA splicing via spliceosome, chloroplast envelope, and AMP binding. The candidate hub genes of the interested modules were further identified, and their regulatory relationships were analyzed based on co-expression data. The temporal expression pattern of the 12 hub genes was verified within 96 h after powdery mildew inoculation by RT-PCR assay. In this study, we preliminarily explained the resistance mechanism of the wheat-A. cristatum translocation lines and obtained the hub candidate genes, which laid a foundation in the exploration of resistance genes in A. cristatum for powdery mildew-resistant breeding in wheat.

Prediction of effector protein structures from fungal phytopathogens enables evolutionary analyses

Elucidating the similarity and diversity of pathogen effectors is critical to understand their evolution across fungal phytopathogens. However, rapid divergence that diminishes sequence similarities between putatively homologous effectors has largely concealed the roots of effector evolution. Here we modelled the structures of 26,653 secreted proteins from 14 agriculturally important fungal phytopathogens, six non-pathogenic fungi and one oomycete with AlphaFold 2. With 18,000 successfully predicted folds, we performed structure-guided comparative analyses on two aspects of effector evolution: uniquely expanded sequence-unrelated structurally similar (SUSS) effector families and common folds present across the fungal species. Extreme expansion of lineage-specific SUSS effector families was found only in several obligate biotrophs, Blumeria graminis and Puccinia graminis. The highly expanded effector families were the source of conserved sequence motifs, such as the Y/F/WxC motif. We identified new classes of SUSS effector families that include known virulence factors, such as AvrSr35, AvrSr50 and Tin2. Structural comparisons revealed that the expanded structural folds further diversify through domain duplications and fusion with disordered stretches. Putatively sub- and neo-functionalized SUSS effectors could reconverge on regulation, expanding the functional pools of effectors in the pathogen infection cycle. We also found evidence that many effector families could have originated from ancestral folds conserved across fungi. Collectively, our study highlights diverse effector evolution mechanisms and supports divergent evolution as a major force in driving SUSS effector evolution from ancestral proteins.