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MPMI: Identification and Characterization of In-planta Expressed Secreted Effectors from Magnaporthe oryzae that Induce Cell Death in Rice

MPMI: Identification and Characterization of In-planta Expressed Secreted Effectors from Magnaporthe oryzae that Induce Cell Death in Rice | Rice Blast | Scoop.it

A total of 6,413 in planta–expressed fungal genes, including 851 genes encoding predicted effector proteins, were identified. We used a protoplast transient expression system to assess 42 of the predicted effector proteins for the ability to induce plant cell death. This study demonstrates that our integrative genomic approach is effective for the identification of in planta–expressed cell death–inducing effectors from M. oryzae that may play an important role facilitating colonization and fungal growth during infection.


Via Kamoun Lab @ TSL, Elsa Ballini
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Rice Blast
Scientific articles on rice blast and wheat blast 20 new articles each month !
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INCREASE OF FUNGAL PATHOGENICITY AND ROLE OF PLANT GLUTAMINE IN NITROGEN-INDUCED SUSCEPTIBILITY (NIS) TO RICE BLAST

INCREASE OF FUNGAL PATHOGENICITY AND ROLE OF PLANT GLUTAMINE IN NITROGEN-INDUCED SUSCEPTIBILITY (NIS) TO RICE BLAST | Rice Blast | Scoop.it
Understanding why nitrogen fertilization increase the impact of many plant diseases is of major importance. The interaction between Magnaporthe oryzae and rice was used as a model for analyzing the molecular mechanisms underlying Nitrogen-Induced Susceptibility (NIS). We show that our experimental system in which nitrogen supply strongly affects rice blast susceptibility only slightly affects plant growth. In order to get insights into the mechanisms of NIS, we conducted a dual RNA-seq experiment on rice infected tissues under two nitrogen fertilization regimes. On the one hand, we show that enhanced susceptibility was visible despite an over-induction of defense gene expression by infection under high nitrogen regime. On the other hand, the fungus expressed to high levels effectors and pathogenicity-related genes in plants under high nitrogen regime. We propose that in plants supplied with elevated nitrogen fertilization, the observed enhanced induction of plant defense is over-passed by an increase in the expression of the fungal pathogenicity program, thus leading to enhanced susceptibility. Moreover, some rice genes implicated in nitrogen recycling were highly induced during NIS. We further demonstrate that the OsGS1-2 glutamine synthetase gene enhances plant resistance to M. oryzae and abolishes NIS and pinpoint glutamine as a potential key nutrient during NIS.
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Genetic diversity of disease resistance genes in foxtail millet (Setaria italica L.)

Genetic diversity of disease resistance genes in foxtail millet (Setaria italica L.) | Rice Blast | Scoop.it
Foxtail millet (Setaria italica L.) is a potential biofuel plant species, which is also one of the most commonly cultivated millet species for food and fodder. In this study we aimed to conduct a genome-wide identification of Coiled-coil, Nucleotide-binding site, Leucine-rice repeat (CNL) disease resistance genes (R-genes) in foxtail millet and study their evolutionary relationships. We identified 242 CNL genes with domains for NBS-LRR receptor function, with the addition of a few genes that contained transmembrane or zinc finger domains. Of the identified CNL genes, more than half formed gene clusters within the foxtail millet genome, with the majority showing evidence of tandem duplications. Syntenic analysis displayed chromosomal similarities among foxtail millet, rice, and barley, identifying strong syntenic relationships between foxtail millet and rice. Approximately 30% of the foxtail millet CNL gene clusters were found on chromosome Si08, exhibiting strong synteny with chromosome Os11 of rice. Selection pressure analysis showed a prevalence of purifying selection among all phylogenetic clades. We also identified the foxtail millet homologs of several well-studied R-genes in other species. The results from this study will have implications in research on CNL gene signaling pathways and development of durable resistance in foxtail millet and other crop species in general.
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Daily chart: Asian countries are eating more wheat

Daily chart: Asian countries are eating more wheat | Rice Blast | Scoop.it
“SO CENTRAL is rice to life in Asia that in many countries, rather than asking “how are you?” people ask, “have you eaten rice yet?” Around 90% of the world’s rice is consumed in Asia—60% of it in China, India and Indonesia alone. In every large country except Pakistan, Asians eat more rice than the global average.”
Via CIMMYT, Int., Rey Thomas
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Calcium-triggered accumulation of defense-related transcripts enhances wheat resistance to leaf blast

Calcium (Ca) is an essential macronutrient for plant growth, but its role in plant disease resistance remains largely elusive. Here, we investigated whether Ca increases the abundance of defense-related transcripts and enhances wheat resistance to leaf blast, caused by Pyricularia oryzae. To address these issues, wheat plants were grown in nutrient solution containing three Ca concentrations (0.26, 2.6 and 5 mM) either non-inoculated or inoculated with P. oryzae. Ca content in the leaves increased from 1.3 to 4.1% and blast severity decreased from 36.8 to 8.2% as the Ca concentration in the nutrient solution increased from 0.26 to 5 mM. The expression level of salicylic (SA) and jasmonic acid (JA) pathways gene markers (pathogenesis-related 1 and allene oxide synthase, respectively) as well as chitinase, β-1,3-glucanase, phenylalanine ammonia lyase, peroxidase and polyphenol oxidase was higher in inoculated plants grown with 5 mM Ca than in those grown with 0.26 mM Ca. Collectively, results suggest that Ca, in addition to its well-known effect in cell wall strengthening and inhibition of fungal-secreted pectolytic enzymes, plays an active role in wheat resistance to leaf blast through upregulation of defense-related gene expression upon P. oryzae infection. Both of SA and JA pathways seems to make up a central node in such Ca-mediated resistance.
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Profiling the extended phenotype of plant pathogens

Profiling the extended phenotype of plant pathogens | Rice Blast | Scoop.it
One of the most fundamental questions in plant pathology is what determines whether a pathogen grows within a plant? This question is frequently studied in terms of the role of elicitors and pathogenicity factors in the triggering or overcoming of host defences. However, this focus fails to address the basic question of how the environment in host tissues acts to support or restrict pathogen growth. Efforts to understand this aspect of host–pathogen interactions are commonly confounded by several issues, including the complexity of the plant environment, the artificial nature of many experimental infection systems and the fact that the physiological properties of a pathogen growing in association with a plant can be very different from the properties of the pathogen in culture. It is also important to recognize that the phenotype and evolution of pathogen and host are inextricably linked through their interactions, such that the environment experienced by a pathogen within a host, and its phenotype within the host, is a product of both its interaction with its host and its evolutionary history, including its co-evolution with host plants. As the phenotypic properties of a pathogen within a host cannot be defined in isolation from the host, it may be appropriate to think of pathogens as having an ‘extended phenotype’ that is the product of their genotype, host interactions and population structure within the host environment. This article reflects on the challenge of defining and studying this extended phenotype, in relation to the questions posed below, and considers how knowledge of the phenotype of pathogens in the host environment could be used to improve disease control.

What determines whether a pathogen grows within a plant?

What aspects of pathogen biology should be considered in describing the extended phenotype of a pathogen within a host?

How can we study the extended phenotype in ways that provide insights into the phenotypic properties of pathogens during natural infections?

Via Christophe Jacquet, Freddy Monteiro, Tatsuya Nobori
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7th European Conference on Prokaryotic and Fungal Genomics

7th European Conference on Prokaryotic and Fungal Genomics | Rice Blast | Scoop.it

It is our great pleasure to cordially invite you to the 7th European Conference on Prokaryotic and Fungal Genomics – ProkaGENOMICS 2017. This meeting will take place from 19–22 September 2017 in Göttingen/Germany, and focus on the presentation of the latest results in the rapidly growing field of prokaryotic and fungal functional genomics. The ProkaGENOMICS conference series has been launched already in 2003 when the genomics revolution was still in its infancy. At present, the increasing power of high-throughput technologies drives research in microbiology. The OMICS-technologies have become the new mantra in functional microbial genomics, allowing the simultaneous examination of thousands of genes, transcripts, proteins, or metabolites. These powerful tools yielded completely new scientific objectives, e.g. monitoring microbial community dynamics as well as comprehensive cellular processes under a wide range of different conditions. ProkaGENOMICS 2017 will address this challenging field of microbiology. The main conference topics will be: • Towards Biotechnology – Metabolic Modeling & Genome/ Enzyme Engineering • Systems and Synthetic Biology • Omics at its best – Global Gene Regulation & Proteome Dynamics • MetaOmics & Microbial Ecosystems • Infectiongenomics & Host Microbe Interplay • Microbial Evolution, Population Genomics & Epidemiology • Microbiome Host Interaction • Functional Phage Genomics

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SP-LL-37, human antimicrobial peptide, enhances disease resistance in transgenic rice

SP-LL-37, human antimicrobial peptide, enhances disease resistance in transgenic rice | Rice Blast | Scoop.it
Human LL-37 is a multifunctional antimicrobial peptide of cathelicidin family. It has been shown in recent studies that it can serve as a host’s defense against influenza A virus. We now demonstrate in this study how signal peptide LL-37 (SP-LL-37) can be used in rice resistance against bacterial leaf blight and blast. We synthesized LL-37 peptide and subcloned in a recombinant pPZP vector with pGD1 as promoter. SP-LL-37 was introduced into rice plants by Agrobacterium mediated transformation. Stable expression of SP-LL-37 in transgenic rice plants was confirmed by RT-PCR and ELISA analyses. Subcellular localization of SP-LL-37-GFP fusion protein showed evidently in intercellular space. Our data on testing for resistance to bacterial leaf blight and blast revealed that the transgenic lines are highly resistant compared to its wildtype. Our results suggest that LL-37 can be further explored to improve wide-spectrum resistance to biotic stress in rice.
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Towards understanding the mechanism of cytoplasmic effector translocation during biotrophic development of Magnaporthe oryzae.

Rice blast caused by Magnaporthe oryzae (synonym of Pyricularia oryzae), a hemibiotroph and facultative pathogen, is the most destructive disease of rice worldwide. During the infection process, M. oryzae secretes various effectors, which are hypothesized to be involved in effective host infection. Effectors are classified by their destinations in the interaction court, with apoplastic effectors residing in the extracellular plant compartment and cytoplasmic effectors translocating into the cytoplasm of living plant cells. Notably, cytoplasmic effectors of M. oryzae are associated with a specialized interfacial structure, the biotrophic interfacial complex (BIC). To date, little is known about the mechanisms of effector uptake into plant cells during fungal infection. Here we show evidence for translocation of the cytoplasmic effectors Bas1, Pwl1 and Pwl2 in vesicles from BICs to rice cytoplasm during biotrophic development. Using fluorescent protein tagging, we found that cytoplasmic effectors Bas1, Pwl1 and Pwl2 are sorted into different vesicles in BICs formed on primary hyphae (PH), revealing new levels of functional complexity for this biotrophic structure. In contrast, most of the vesicles from BICs on mature bulbous hyphae showed colocalization of the cytoplasmic effectors. Whereas BICs on primary hyphae deliver effectors in micro-vesicles, BICs on mature bulbous hypha deliver effectors in macro-vesicles, at times reaching diameter sizes over 3 µm. Furthermore, we demonstrate that endocytosis inhibitors Cantharidin, Triclosan and Wortmannin induce abnormally-shaped and swollen BICs as well as the accumulation of cytoplasmic effectors in the BICs. Moreover, Cantharidin, Triclosan and Wortamannin treatment induced the accumulation of the cytoplasmic effectors under penetration pores, suggesting that effector uptake begins even before host penetration. Based on these results, it appears that cytoplasmic effector translocation is mediated by vesicle formation and may be characteristic of appressoria as well as biotrophic invasive hyphae. Our results also suggest a potential role of M. oryzae effectors for manipulation of the host cell endocytosis process.
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Characterization of the interaction between bacteria and Magnaporthe oryzae.

Like many other foliar pathogens, the rice blast fungus Magnaporthe oryzae form appressoria for plant penetration. Interestingly, bacteria attached to appressoria are often observed in in vitro assays for appressorium formation. In this study, Pseudomonas fluorescens strain TR3 was isolated from rice leaves and used for studying for its interaction with M. oryzae. TR3 could attached to conidia, germ tubes, and appressoria and caused fungal cell death. In co-cultivation assays, TR3 was inhibitory to colony growth in a contact-based manner, excluding the possibility of small antifungal metabolites. Attachment of TR3 to fungal spores was inhibitory to germination and plant infection. Antifungal activities were specifically detected in culture filtrates of TR3 grown in M9 medium together with M. oryzae, suggesting the involvement of the type 3 secretion system (T3SS). Mutants detected of the hrcC gene encoding the outer membrane pore forming protein of T3SS failed to form flagella and had reduced inhibitory activities against M. oryzae. In contrast, deletion of the pilC gene that is essential for T4SS had no impact on its antifungal potential although affected attachment to fungal hyphae or conidia. The phosphorylation level of Pmk1 MAP kinase was significantly increased after incubated with TR3 for 2 h, which was not observed in the sho1 deletion mutant. Taken together, these results indicate that the interaction of P. fluorescens with M. oryzae is inhibitory to fungal growth, development, and pathogenesis and likely involves T3SS effectors affecting intracellular signaling pathways.
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Magnaporthe lunch meeting 12:15 PM - 2:30 PM on Friday, March 17, 2017

Magnaporthe lunch meeting 12:15 PM - 2:30 PM on Friday, March 17, 2017 | Rice Blast | Scoop.it

March 17-22, 2015, Pacifc Grove, California #FUNGAL15


On behalf of the International Rice Blast Policy Committee, we would like to invite you to a Magnaporthe lunch meeting that will be held during the Fungal Genetics Conference in Asilomar this week. When: 12:15 PM - 2:30 PM on Friday, March 17, 2017 Where: To be announced (a conference room in Asilomar) What: (1) Short research talks and (2) discussion on wheat blast and future Magnaporthe meetings To help us organize the meeting, please send the following information to us (Chang-Hyun Khang and Marc-Henri Lebrun ). Please let us know if you plan to attend. We will have lunch boxes delivered to the meeting room. Students, postdocs, or new PIs: If you wish to give a talk, please let us know asap. We have one or two slots for presentations (each for 10 minutes). Send us a talk title (no abstract needed). Despite short notice, we hope to see you in Asilomar! Chang-Hyun Khang and Marc-Henri Lebrun

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Durable resistance to rice blast

Durable resistance to rice blast | Rice Blast | Scoop.it
The blast fungus Magnaporthe oryzae (Pyricularia oryzae) destroys rice crops worldwide (see the photo) (1) and now threatens global wheat production as well. Wheat blast disease has been spreading in South America since 1985, and last year it devastated wheat crops in Bangladesh (2, 3). Incorporation of resistance (R) genes into rice presents an effective, economical, and environmentally sound way to control blast (4). However, it takes years to move an R gene into a rice variety, whereas the highly variable fungus can overcome the R gene effects within 2 or 3 years after planting. Also, many R genes are associated with yield losses, so farmers often prefer rice varieties without R genes. On page 962 of this issue, Deng et al. (5) report a new R gene that comes with a homologous partner to counteract the yield penalty.
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The Sequence of 1504 Mutants in the Model Rice Variety Kitaake Facilitates Rapid Functional Genomic Studies

The Sequence of 1504 Mutants in the Model Rice Variety Kitaake Facilitates Rapid Functional Genomic Studies | Rice Blast | Scoop.it

ABSTRACT The availability of a whole-genome sequenced mutant population and the cataloging of mutations of each line at a single-nucleotide resolution facilitates functional genomic analysis. To this end, we generated and sequenced a fast-neutron-induced mutant population in the model rice cultivar Kitaake (Oryza sativa L. ssp. japonica), which completes its life cycle in 9 weeks. We sequenced 1,504 mutant lines at 45-fold coverage and identified 91,513 mutations affecting 32,307 genes, 58% of all rice genes. We detected an average of 61 mutations per line. Mutation types include single base substitutions, deletions, insertions, inversions, translocations, and tandem duplications. We observed a high proportion of loss-of-function mutations. Using this mutant population, we identified an inversion affecting a single gene as the causative mutation for the short-grain phenotype in one mutant line with a small segregating population. This result reveals the usefulness of the resource for efficient identification of genes conferring specific phenotypes. To facilitate public access to this genetic resource, we established an open access database called KitBase that provides access to sequence data and seed stocks, enabling rapid functional genomic studies of rice.


Via Loïc Lepiniec, Jennifer Mach
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‘Wheat blast’ disease enters India from Bangladesh, ICAR official says damage contained

‘Wheat blast’ disease enters India from Bangladesh, ICAR official says damage contained | Rice Blast | Scoop.it
Wheat blast’, a fungal disease that attacks the standing crop, has been reported in a few pockets in Nadia and Murshidabad districts of Bengal. According to an official with the Indian Council for Agricultural Research (ICAR), the wheat blast disease was reported in a couple of places a few weeks back in Nadia and Murshidabad districts. However, it is unlikely to cause any major damage to the standing crop harvesting of which will commence within a next few weeks.

“The disease has entered our territory from Bangladesh, especially in the bordering districts of Bengal. However, in most of the parts of Bihar and Bengal, farmers sow the HD 2967 variety, which has in-built capacity to deal with the fungus,” Trilochan Mohapatra, director general of the ICAR, told FE.
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A web-based microsatellite database for the Magnaporthe oryzae genome

A web-based microsatellite database for the Magnaporthe oryzae genome | Rice Blast | Scoop.it
Rice blast disease caused by a fungal pathogen Magnaporthe oryzae, which results up to 90% crop yield loss during severe epidemics [1]. Rice fulfils food energy requirement of 2/3rd of world’s population. Understanding about the pathogen and its diversity among different strains of a particular geographic region can help breeders for proper deployments of blast resistance genes in the region. DNA markers are the unique regions within a genome which may be associated with the genes responsible for specific trait in an organism. Microsatellite or simple sequence repeat markers are stretches of DNA in which the same short nucleotide sequence is tandemly repeated within the genome [2]. These sequences are also known as simple sequence repeats (SSRs) or simple tandem repeats (STRs). SSR is generally a 1-6 nucleotide sequence variations present across the eukaryotic genome [3,4, 5]. Polymorphism, or variation, among SSR markers is determined by the number of times of the base sequence repeats. Its hyper-variability in among the related organisms makes them excellent markers for phenotype mapping, marker assisted selection, genotype identification and analysis of genetic diversity. The nature of SSRs gives them a number of advantages over other molecular markers by their abundances in genome, high-level of polymorphisms, codominance nature, open accessibility, simple assay method, and feasibility to use at high-throughput level. They are one of the most advanced marker technologies, after single nucleotide polymorphism (SNP) markers, available in genetic research. Considering these above benefits, a SSR marker database was developed from genomes of two M. oryzae isolates, RML-29 (avirulent, designated name Mo-nwi-55) and RP-2421 (virulent, designated name Mo-nwi-31). These two isolates were selected for the database development on the basis of their virulence spectrum towards different rice blast resistance genes.
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Anti-blast mutant screened from tissue cultured with crude toxin from Pyricularia oryzae.

The crude toxin was extracted from liquid of different Jiangsu province's physiological races of rice blast fungus, the local rice variety which was sensitive to rice blast was chosen as tested material. The effects of crude toxin on its seed germination, mature embryo callus's growth and differentiation were studied. Under appropriate concentrations of crude toxin, the blast-resistant mutant during the whole time of embryo callus's growth was screen out. The results showed that the seeds' germination was inhibited strongly with the increasing of the toxin concentration, and the germination was totally inhibited when the concentration was 1 mL/10 mL. The growth of embryo callus was inhibited gradually with the increasing of toxin's concentration, and the callus's growth was completely stopped when the concentration was 25%. The differentiation of embryo callus was sensitive to toxin, and the differentiation rate of seedling was dropped significantly when the concentration was 5%. The rice seedlings from the mature embryo cultured with crude toxin were injected by rice blast fungus in the field, their disease resistance was improved compared with CK. This study suggested that the method of using the media including toxin to induct mutant was a effective protocol to improve the rice resistance of rice-blast.
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Comparison and Validation of Putative Pathogenicity-Related Genes Identified by T-DNA Insertional Mutagenesis and Microarray Expression Profiling in Magnaporthe oryzae

Comparison and Validation of Putative Pathogenicity-Related Genes Identified by T-DNA Insertional Mutagenesis and Microarray Expression Profiling in Magnaporthe oryzae | Rice Blast | Scoop.it

High-throughput technologies of functional genomics such as T-DNA insertional mutagenesis and microarray expression profiling have been employed to identify genes related to pathogenicity in Magnaporthe oryzae. However, validation of the functions of individual genes identified by these high-throughput approaches is laborious. In this study, we compared two published lists of genes putatively related to pathogenicity in M. oryzae identified by T-DNA insertional mutagenesis (comprising 1024 genes) and microarray expression profiling (comprising 236 genes), respectively, and then validated the functions of some overlapped genes between the two lists by knocking them out using the method of target gene replacement. Surprisingly, only 13 genes were overlapped between the two lists, and none of the four genes selected from the overlapped genes exhibited visible phenotypic changes on vegetative growth, asexual reproduction, and infection ability in their knockout mutants. Our results suggest that both of the lists might contain large proportions of unrelated genes to pathogenicity and therefore comparing the two gene lists is hardly helpful for the identification of genes that are more likely to be involved in pathogenicity as we initially expected.

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Functional Characterization of Endophytic Fungal Community Associated with Oryza sativa L. and Zea mays L.

Functional Characterization of Endophytic Fungal Community Associated with Oryza sativa L. and Zea mays L. | Rice Blast | Scoop.it
In a natural ecosystem, the plant is in a symbiotic relationship with beneficial endophytes contributing huge impact on its host plant. Therefore, exploring beneficial endophytes and understanding its interaction is a prospective area of research. The present work aims to characterize the fungal endophytic communities associated with healthy maize and rice plants and to study the deterministic factors influencing plant growth and biocontrol properties against phytopathogens, viz, Pythium ultimum, Sclerotium oryzae, Rhizoctonia solani, and Pyricularia oryzae. A total of 123 endophytic fungi was isolated using the culture-dependent approach from different tissue parts of the plant. Most dominating fungal endophyte associated with both the crops belong to genus Fusarium, Sarocladium, Aspergillus, and Penicillium and their occurrence was not tissue specific. The isolates were screened for in vitro plant growth promotion, stress tolerance, disease suppressive mechanisms and based on the results, each culture from both the cereal crops was selected for further study. Acremonium sp. (ENF 31) and Penicillium simplicisssum (ENF22), isolated from maize and rice respectively could potentially inhibit the growth of all the tested pathogens with 46.47 ± 0.16 mm to 60.09 ± 0.04 mm range zone of inhibition for ENF31 and 35.48 ± 0.14 to 62.29 ± 0.15 mm for ENF22. Both significantly produce the defensive enzymes, ENF31 could tolerate a wide range of pH from 2 to 12, very important criteria, for studying plant growth in different soil types, especially acidic as it is widely prevalent here, making more land unsuitable for cultivation. ENF22 grows in pH range 3–12, with 10% salt tolerating ability, another factor of consideration. Study of root colonization during 7th to 30th days of growth phase reveals that ENF31 could colonize pleasantly in rice, though a maize origin, ranging from 1.02 to 1.21 log10 CFU/g root and in maize, it steadily colonizes ranging from 0.95 to 1.18 log10 CFU, while ENF22 could colonize from 0.98 to 1.24 Log10CFU/g root in rice and 1.01 to 1.24Log10CFU/g root in maize, just the reverse observed in Acremonium sp. Therefore, both the organism has the potency of a promising Bio-resource agent, that we must definitely explore to fill the gap in the agriculture industry.
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Transposon-mediated epigenetic regulation contributes to phenotypic diversity and environmental adaptation in rice

Transposon-mediated epigenetic regulation contributes to phenotypic diversity and environmental adaptation in rice | Rice Blast | Scoop.it

Most rice genes are associated with transposable elements (TEs).
TE-based epigenetic regulation of expression of adjacent genes is prevalent in rice.
Environmental conditions can activate TEs to generate genetic or epigenetic variations.
TE-driven variations contributed to rice evolution and northward expansion.

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Epidemiology, virulence and molecular diversity in blast [Magnaporthe grisea (Hebert) Barr.]

Epidemiology, virulence and molecular diversity in blast [Magnaporthe grisea (Hebert) Barr.] of pearl millet [Pennisetum glaucum (L.) R. Br.] and resistanc
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Novel interkingdom sensing and signaling in rice blast.

Rice Blast represents a model pathosystem, wherein the causal fungus Magnaporthe oryzae perceives and responds to host cues to initiate pathogenic development. Recently, we gained insight into the role of the CFEM motif in surface sensing/response function of the Pth11 receptor, which upon activation utilizes the late endosomal compartment as a scaffold for sustained G-protein/cyclic AMP signaling to induce appressorium formation. Highly regulated redox homeostasis was found to be important for Pth11-mediated appressorium formation in M. oryzae. Swapping of the CFEM motifs and in vitro binding assays provided further insight into the molecular signatures that are likely recognized by the individual CFEM domains.

Our data suggest that a key step in the appressorium initiation process is cessation of vegetative growth at the germ tube tip, which is achieved through the timely synthesis and action of a novel signaling moiety, Jasmonic acid (JA; fungal oxylipin), in M. oryzae. Such intrinsically produced JA is an important signal for appressorium formation and acts in concert with the MAPK cascade in M. oryzae. Interestingly, the cAMP signaling itself is necessary for the synthesis of JA in the blast fungus, which subsequently utilizes the ABM monooxygenase to produce a novel metabolite effector ie 12-hydroxy jasmonic acid. Typically, a biotrophic phase precedes necrotrophy during Magnaporthe-Rice interaction and disease development. The blast fungus secretes 12OH-JA into the host tissue and directly inhibits the phytohormone signaling involved in pathogen recognition and defense in rice. We propose a critical role for intrinsic jasmonates in pathogenic differentiation in M. oryzae, and a highly intricate chemical communication system during biotrophic interactions in Rice Blast.
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MagNet: the integrated gene network of the rice blast fungus Magnaporthe oryzae.

Magnaporthe oryzae, the rice blast fungus, plays a role as a model organism in the area of molecular plant-microbe interaction research. Studies on pathogenic mechanism of this fungus revealed many genes in signaling pathways are essential for appressorium formation and penetration. As multi-omics data are being available, genomic-level researches have been conducted to uncover the underlying biological process during the pathogenesis of M. oryzae. Identifying genome-wide protein-protein interaction (PPI) network is one of omics-level approaches which can help to understand signaling and regulatory pathways. However, existing biological network resources of M. oryzae are not sufficient to decipher molecular mechanisms of pathogenesis. In this study, PPI network analysis platform of M. oryzae, MagNet, was constructed with three methods: homology-based ‘Interolog’ search, co-expression network construction, and domain-domain interaction based prediction. Interologs within M. oryzae proteins which have orthologues in model species were predicted with six PPI repositories. Co-expression networks were built with RNA sequencing data from the infection stages and those from the vegetative stage. In addition, we gathered the information of domain-domain interactions (DDIs) from two integrated DDI databases, and predicted interactions between domain-annotated proteins. With three approaches all together, the pan-network with 6,600,976 interactions was generated including highly confident 215,731 interactions found in more than three sub-networks. Experimental data on M. oryzae PPIs support that our highly confident PPI network can predict PPIs with higher sensitivity (89.65%) and specificity (78.57%) compared to the previously constructed databases. MagNet would provide integrated biological network data which can help to understand the molecular mechanisms of the rice blast fungus and beyond.
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Characterization of Magnaporthe oryzae effectors and their interacting partners in rice.

The first line of defense against fungal pathogens by plants is the detection of pathogen-associated molecular patterns by pattern recognition receptors to trigger PAMP-triggered immunity (PTI). Pathogens that overcome PTI and successfully invade host cells do so by delivering effectors that interfere with PTI, leading to effector-triggered susceptibility (ETS). We have identified a family of effector-encoding genes in the rice blast fungus Magnaporthe oryzae. We first using RNA-seq to identify genes that were differentially expressed during plant infection and which were temporally co-regulated during the early stages of tissue colonisation by the fungus. The putative effector genes were then characterized by targeted gene deletion and their protein products localized using live-cell imaging. We found a group of apoplastic effectors that outlined invasive hyphae while cytoplasmic effectors accumulated at the biotrophic interfacial complex (BIC). We then identified potential effector targets in rice using yeast two hybrid analysis and co-immunoprecipitation in infected plant tissue. Putative protein-protein interactions were independently confirmed using Bimolecular Fluorescence Complementation (BiFC) assays. In this way, we have characterized a set of 32 MEP (Magnaporthe effector protein) genes. We will report progress on their characterization and, specifically, the role of MEP1 and MEP3 in fungal virulence and host immune suppression, respectively.
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Multiscale Phenotyping and Decision Strategies in Breeding for Resistance

Multiscale Phenotyping and Decision Strategies in Breeding for Resistance | Rice Blast | Scoop.it
Advances in biotechnology have rendered tracking of quantitative trait loci (QTLs) a much easier task, making phenotyping, and not genotyping, the main bottleneck to integrating quantitative host plant resistance into breeding programs. The relevance of phenotyping methods is conditioned by their ability to predict the performance of a genotype at the field scale. Components of resistance represent the keystone hierarchy level between resistance expression in the field (the breeder’s scale) and QTLs (the geneticist’s scale). We describe approaches for upscaling processes to identify components of resistance that best predict field resistance, and for decision making for selection in breeding programs. We further highlight avenues for future research considering specific processes: disease transmission, defoliation, disease escape, polyetic processes, and interactions between components of resistance.
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Cloning and sequence analysis of Avr-Pik gene from Magnaporthe oryzae isolates from Sarawak - UNIMAS Institutional Repository

Avr-Pik gene found in Magnaporthe oryzae is one of the specific effector protein that interact with host resistance (R) gene that result in effective activation of innate immunity. M. oryzae is an ascomycete fungal pathogen that has been found infecting cultivated rice (Oryzae sativa) and become the major constraint in rice global production. The aim of this research is to isolate, clone and sequence Avr-Pik gene from M. oryzae originated from Sarawak. Prior to isolation and cloning, a set of primer consist of forward primer (5'- ATGCGTGTTACCACTTTTAACA-3') and reverse primer (5'- TTAAAAGCCGGGCCTTTT-3') was designed based on Avr-Pik sequences obtained from database. Gradient Polymerase Chain Reaction (PCR) was performed to optimize the annealing temperature. DNA fragments of -342 bp were obtained from the amplification and were purified. Blue and white screening was conducted to grow colonies and distinguish between the recombinant and non-recombinant colonies. Subsequently, plasmid miniprep was conducted to obtain plasmid and it was sent for sequencing. The sequencing result was corroborated by using BLAST in which showed highest similarity with M. oryzae isolates from Japan. Based on this study, future identification and sequence analysis of this gene in understanding gene-for-gene resistance can be carried out, hence establishing Moryzae as the fungal model to study fungal pathogenicity.
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Biological Mechanisms of Plant Interactions With a Combination of Biotic and Abiotic Stresses | Frontiers Research Topic

This Research Topic addresses the way plants respond to a combination of different types of stresses, for instance, how a plant deals with simultaneous attacks by pathogens (biotic stress) and environmental stresses (abiotic stress). While most plant research has been focused on the understanding of individual biotic or abiotic stress responses, we propose to encourage submissions on a broader approach that better represents stress conditions encountered in the field. Indeed, in nature or under field conditions plants are not just dealing with one environmental stress: they are often coping with several simultaneously occurring biotic and abiotic stresses. Recent findings indicate that predicting the plant response to a combination of stresses cannot be made from the knowledge of an individual stress, and therefore remains a major challenge.It remains to be determined what biological systems respond to combined stresses. Hormones, with their complex regulation and cross-talk, as well as molecules like calcium or reactive oxygen species, are likely to play a key role in combined stresses as potential integrators of multiple signaling pathways. Moreover, given the importance of epigenetic changes in response to a given stress, the role of epigenetic modifications during combined stresses needs further attention. Combined stresses may not only be concomitant but be successive, either during a plant’s life or across generations. In that respect, trans-generational, epigeneti
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MST50 Is Involved in Multiple MAP Kinase Signaling Pathways in Magnaporthe oryzae

MST50 Is Involved in Multiple MAP Kinase Signaling Pathways in Magnaporthe oryzae | Rice Blast | Scoop.it
Appressorium formation plays a critical role in Magnaporthe oryzae. Mst50 is an adapter protein of the Mst11-Mst7-Pmk1 cascade that is essential for appressorium formation. To further characterize its functions, affinity purification was used to identify Mst50-interacting proteins (MIPs) in this study. Two of the MIPs are Mst11 and Mst7 that are known to interact with Mst50 for Pmk1 activation. Surprisingly, two other MIPs are Mck1 and Mkk2 that are the upstream kinases of the Mps1 pathway. Domain deletion analysis showed that the sterile alpha-motif of Mst50 but not the Ras-association domain was important for its interaction with Mck1 and responses to cell wall and oxidative stresses. The mst50 mutant was reduced in Mps1 activation under stress conditions. MIP11 encodes a RACK1 protein that also interacted with Mck1. Deletion of MIP11 resulted in defects in cell wall integrity, Mps1 phosphorylation, and plant infection. Furthermore, Mst50 interacted with histidine kinase Hik1, and the mst50 mutant was reduced in Osm1 phosphorylation. These results indicated that Mst50 is involved in all three MAPK pathways in M. oryzae although its functions differ in each pathway. Several MIPs are conserved hypothetical proteins and may be involved in responses to various signals and crosstalk among signaling pathways. This article is protected by copyright. All rights reserved.
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