Nonhost resistance to Magnaporthe oryzae in Arabidopsis thaliana

Rice blast, caused by Magnaporthe oryzae, is a devastating disease of rice (Oryza sativa). The mechanisms involved in resistance of rice to blast have been studied extensively and the rice—M. oryzae pathosystem has become a model for plant—microbe interaction studies. However, the mechanisms involved in nonhost resistance (NHR) of other plants to rice blast are still poorly understood. We have recently demonstrated that AGB1 and PMR5 contribute to PEN2-mediated preinvasion resistance to M. oryzae in Arabidopsis thaliana, suggesting a complex genetic network regulating the resistance. To determine whether other defense factors: RAR1, SGT1 and NHO1, affected the A. thaliana-M. oryzae interactions, double mutants were generated between pen2 and these defense-related mutants. All these double mutants exhibited a level of penetration resistance similar to that of the pen2 mutant, suggesting that none of these mutants significantly compromised resistance to M. oryzae in a pen2 background.Key words: nonhost resistance, PEN2, RAR1, SGT1, NHO1Plants face microbial attacks and have evolved innate immunity systems to defend against these threats. The initial step of the immunity signaling pathway is recognition of intra- or extracellular pathogen-derived molecules. Externally oriented transmembrane-type proteins containing leucine-rich repeat (LRR) domains detect extracellular molecules, whereas cytoplasmic sensors possess nucleotide-binding (NB) and LRR domains (NLR).^1,2 The LRR domain serves as a pattern-recognition receptor to detect pathogen-derived molecules or host proteins that are targeted by pathogen peptides that have entered the cell, effectors.³ NLR-type sensors are the substrates of a structurally and functionally conserved chaperone complex that consists of HEAT SHOCK PROTEIN 90 (HSP90) and its cochaperone SUPPRESSOR OF THE G2 ALLELE OF SKP1 (SGT1). REQUIRED FOR MLA12 RESISTANCE 1 (RAR1) regulated the HSP90-SGT1 complex, resulting in the stabilization of NLR proteins. Thus, SGT1 and RAR1 are required for the function of multiple and distinct R genes that encode NLR immune sensors in plants.⁴ Experiments in RAR1-silenced transgenic rice lines showed that RAR1 is not essential for Pib, which encodes an NLR against rice blast fungus.⁵ In contrast, basal resistance to normally virulent races of rice blast fungus or bacterial blight is significantly reduced in RAR1-silenced lines. This result is consistent with earlier reports that RAR1 is involved in basal resistance to virulent Pseudomonas bacteria in Arabidopsis or blast fungus in barley.^6,7 The requirement of SGT1 for immunity in plants is shown mostly by transient silencing of a number of NLR proteins.^8,9 In addition, SGT1 is also required for immune responses triggered by non-NLR-type sensors.¹⁰ This requirement indicates that either SGT1 function is not limited to the NLR sensors, or some unknown SGT1-dependent NLR proteins also operate downstream of non NLR-type sensors. Furthermore, SGT1 is involved in nonhost resistance, indicating that SGT1 may be a general factor of disease resistance.¹⁰ An Arabidopsis mutant, nho1 (nonhost resistance 1), has been isolated on which Pseudomonas syringae pv. phaseolicola grows and causes disease symptoms.^11,12 It is significant that this mutant is also compromised in R-gene-mediated resistance to P. syringae.¹¹ Although NHO1 is the flagellin-induced glycerol kinase, whose exact function in NHR remains elusive.^12,13 A possible explanation might be that altered plant glycerol pools either directly or indirectly affect nutrient availability for P. syringae. NHO1 is also required for resistance to the fungal pathogen Botrytis cinerea, indicating that NHO1 is not limited to bacterial resistance.¹² However, these contributions to NHR to M. oryzae in A. thaliana have not been understood.To determine whether these factors were necessary for the resistance to M. oryzae in A. thaliana, the following A. thaliana mutants were inoculated with M. oryzae and monitored by microscopy: rar1-21;¹⁴ edm1-1;¹⁵ nho1-1,¹¹ (all Col-0 background). All these mutants exhibited a level of penetration resistance similar to that of the wild-type plants (data not shown), suggesting that none of these mutants significantly compromised resistance to M. oryzae. We have recently shown that among the penetration (pen) mutants, only the pen2,¹⁶ mutant allowed increased penetration into epidermal cells by M. oryzae.¹⁷ Thus, double mutants were generated between pen2 and these mutants to determine whether these factors were necessary for the resistance to M. oryzae in a pen2 background: pen2 rar1-21; pen2 edm1-1; pen2 nho1-1. All these double mutants exhibited a level of penetration resistance similar to that of the pen2 mutant (Fig. 1), suggesting that none of these mutants significantly compromised resistance to M. oryzae in a pen2 background. This might indicate that NHR against M. oryzae may not be conferred by RAR1- and SGT1-dependent NLR immune sensors. Alternatively, since there has been no report that RAR1 is required for any known transmembrane sensors, such as FLS2, EFR or Xa21, RAR1- and SGT1-independent transmembrane-type immune sensors may be required for NHR against M. oryzae. Future studies will be required to reveal the genetic and mechanistic requirements for NHR in A. thaliana-M. oryzae interactions.Open in a separate windowFigure 1Double mutant analysis to evaluate the role of the defense related genes on resistance to Magnaporthe oryzae in Arabidopsis thaliana. The frequency of M. oryzae penetration on double mutants at 3 days post-inoculation was expressed as a percentage of total appressoria. Data were collected from six independent plants per line. A minimum of 100 infection sites was inspected per leaf. Results represent mean ± standard error of three independent experiments.     

The Blast Resistance Gene Pi54of Cloned from Oryza officinalis Interacts with Avr-Pi54 through Its Novel Non-LRR Domains

The dominant rice blast resistance gene Pi54 cloned by map-based cloning approach from indica rice cultivar Tetep confers broad spectrum resistance to Magnaporthe oryzae. In this investigation, an orthologue of Pi54 designated as Pi54of was cloned from Oryza officinalis conferring high degree of resistance to M. oryzae and is functionally validated. We have also characterized the Pi54of protein and demonstrate its interaction with AVR-Pi54 protein. The Pi54of encoded ∼43 kDa small and unique cytoplasmic LRR family of disease resistance protein having unique Zinc finger domain overlapped with the leucine rich repeat regions. Pi54of showed Magnaporthe-induced expression. The phylogenetic and western blot analysis confirmed orthologous nature of Pi54 and Pi54of genes, whereas the identity of protein was confirmed through MALDI-TOF analysis. The in silico analysis showed that Pi54of is structurally more stable than other cloned Pi54 proteins. The molecular docking revealed that Pi54of protein interacts with AVR-Pi54 through novel non-LRR domains such as STI1 and RhoGEF. The STI1 and GEF domains which interact with AVR-Pi54 are also components of rice defensome complex. The Pi54of protein showed differential domain specificity while interacting with the AVR protein. Functional complementation revealed that Pi54of transferred in two rice lines belonging to indica and japonica background imparts enhanced resistance against three highly virulent strains of M. oryzae. In this study, for the first time, we demonstrated that a rice blast resistance gene Pi54of cloned from wild species of rice provides high degree of resistance to M. oryzae and might display different molecular mechanism involved in AVRPi54-Pi54of interaction.     

Rice BiP3 regulates immunity mediated by the PRRs XA3 and XA21 but not immunity mediated by the NB-LRR protein,Pi5

Plant innate immunity is mediated by pattern recognition receptors (PRRs) and intracellular NB-LRR (nucleotide-binding domain and leucine-rich repeat) proteins. Overexpression of the endoplasmic reticulum (ER) chaperone, luminal-binding protein 3 (BiP3) compromises resistance to Xanthomonas oryzae pv. oryzae (Xoo) mediated by the rice PRR XA21 [12]. Here we show that BiP3 overexpression also compromises resistance mediated by rice XA3, a PRR that provides broad-spectrum resistance to Xoo. In contrast, BiP3 overexpression has no effect on resistance mediated by rice Pi5, an NB-LRR protein that confers resistance to the fungal pathogen Magnaporthe oryzae (M. oryzae). Our results suggest that rice BiP3 regulates membrane-resident PRR-mediated immunity.     

国外引进水稻种质资源的稻瘟病抗性基因检测与评价

为了筛选出福建省水稻稻瘟病重发区育种中可利用的新抗性资源,在福建省上杭县对156份外引水稻种质资源进行了2年田间自然诱发鉴定,并对Pi2、Pi9、Pi5、Pi54、Pikm、Pita、Pia和Pib等8个稻瘟病抗性基因做了分子检测。结果表明:156份资源对苗瘟、叶瘟、穗颈瘟和综合抗性表现抗病的分别有10份、14份、29份和26份,且苗瘟抗性级别与叶瘟抗性级别(r=0.816,P<0.01)、苗瘟抗性级别与穗颈瘟抗性级别(r=0.347,P<0.01)、以及叶瘟抗性级别与穗颈瘟抗性级别(r=0.344,P<0.01),均呈极显著正相关。分子标记检测到携带稻瘟病抗性基因Pi9、Pi2、Pi54、Pikm、Pi5、Pib、Pia和Pita的水稻资源分别有1、6、20、22、37、88、101和106份,其中携带稻瘟病抗性基因Pi9和Pi2的水稻资源的抗性表现较好,表现抗病的超过60%,携带其他稻瘟病抗性基因的水稻资源表现抗病的均在50%以下;水稻资源携带0~6个稻瘟病抗性基因,随着携带抗性基因数目增加,抗病率呈上升趋势,综合抗性等级呈下降趋势。进一步研究发现,携带Pi9+Pi5+Pikm+Pia、Pi5+Pib+Pita+Pikm+Pia和Pi2+Pi54+Pib+Pita+Pikm+Pia等3个基因型的水稻资源,稻瘟病抗性较好。最后,筛选了8份稻瘟病抗性较好的材料,提供育种者参考、利用。     

Functional complementation of rice blast resistance gene Pi-k h (Pi54) conferring resistance to diverse strains of Magnaporthe oryzae

Blast disease of rice, caused by Magnaporthe oryzae is an explosive disease that can spread rapidly in conducive conditions. R-gene mediated resistance offers an environmentally sustainable solution for management of this important disease of rice. We have earlier identified a unique R-gene of rice, on chromosome 11 of Oryza sativa ssp. indica cultivar Tetep. In this study we report functional validation of the Pi-k ^h (Pi54) gene using complementation assay. The blast resistance candidate gene Pi-k ^h (Pi54) was cloned into a plant transformation vector and the construct was used to transform a japonica cultivar of rice Taipei 309, which is susceptible to M. oryzae. Transgenic lines containing Pi-k ^h (Pi54) gene were found to confer high degree of resistance to diverse isolates of M. oryzae. The callose deposition was analyzed and compared between the transgenic and non-transgenic rice plants and widespread deposition was observed at the infection sites in plants showing incompatible interaction. Successful complementation of Pi-k ^h (Pi54) gene confirmed that the gene is responsible for resistance to M. oryzae in transgenic lines developed during this study. Expression analysis of the gene in resistant plants revealed that the gene is pathogen inducible in nature and is not expressed constitutively. Detection of callose deposition in resistant plants containing Pi-k ^h (Pi54) gene implicates its involvement in the initiation of defense response cascade.     

Rice Snl6, a Cinnamoyl-CoA Reductase-Like Gene Family Member,Is Required for NH1-Mediated Immunity to Xanthomonas oryzae pv. oryzae

Rice NH1 (NPR1 homolog 1) is a key mediator of innate immunity. In both plants and animals, the innate immune response is often accompanied by rapid cell death at the site of pathogen infection. Over-expression of NH1 in rice results in resistance to the bacterial pathogen, Xanthomonas oryzae pv. oryzae (Xoo), constitutive expression of defense related genes and enhanced benzothiadiazole (BTH)- mediated cell death. Here we describe a forward genetic screen that identified a suppressor of NH1-mediated lesion formation and resistance, snl6. Comparative genome hybridization and fine mapping rapidly identified the genomic location of the Snl6 gene. Snl6 is a member of the cinnamoyl-CoA reductase (CCR)-like gene family. We show that Snl6 is required for NH1-mediated resistance to Xoo. Further, we show that Snl6 is required for pathogenesis-related gene expression. In contrast to previously described CCR family members, disruption of Snl6 does not result in an obvious morphologic phenotype. Snl6 mutants have reduced lignin content and increased sugar extractability, an important trait for the production of cellulosic biofuels. These results suggest the existence of a conserved group of CCR-like genes involved in the defense response, and with the potential to alter lignin content without affecting development.     

Proteomic analysis of rice mutants susceptible to Magnaporthe oryzae

To identify genes involved in rice Pi5-mediated disease resistance to Magnaporthe oryzae, we compared the proteomes of the RIL260 rice strain carrying the Pi5 resistance gene with its susceptible mutants M5465 and M7023. Proteins were extracted from the leaf tissues of both RIL260 and the mutant lines at 0, 24, and 48 h after M. oryzae inoculation and separated by two-dimensional polyacrylamide gel electrophoresis (2-DE). Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis identified eight proteins that were differently expressed between the resistant and susceptible plants (three down- and five up-regulated proteins in the mutants). The down-regulated proteins included a triosephosphate isomerase (spot no. 2210), a 2,3-bisphosphoglycerate-independent phosphoglycerate mutase (no. 3611), and an unknown protein (no. 4505). In addition, the five up-regulated proteins in the mutants were predicted to be a fructokinase I (no. 313), a glutathione S-transferase (no. 2310), an atpB of chloroplast ATP synthase (no. 3616), an aminopeptidase N (no. 3724), and an unknown protein (no. 308). These results suggest that proteomic analysis of rice susceptible mutants is a useful method for identifying novel proteins involved in resistance to the M. oryzae pathogen.     

The RhoGAP SPIN6 Associates with SPL11 and OsRac1 and Negatively Regulates Programmed Cell Death and Innate Immunity in Rice

The ubiquitin proteasome system in plants plays important roles in plant-microbe interactions and in immune responses to pathogens. We previously demonstrated that the rice U-box E3 ligase SPL11 and its Arabidopsis ortholog PUB13 negatively regulate programmed cell death (PCD) and defense response. However, the components involved in the SPL11/PUB13-mediated PCD and immune signaling pathway remain unknown. In this study, we report that SPL11-interacting Protein 6 (SPIN6) is a Rho GTPase-activating protein (RhoGAP) that interacts with SPL11 in vitro and in vivo. SPL11 ubiquitinates SPIN6 in vitro and degrades SPIN6 in vivo via the 26S proteasome-dependent pathway. Both RNAi silencing in transgenic rice and knockout of Spin6 in a T-DNA insertion mutant lead to PCD and increased resistance to the rice blast pathogen Magnaporthe oryzae and the bacterial blight pathogen Xanthomonas oryzae pv. oryzae. The levels of reactive oxygen species and defense-related gene expression are significantly elevated in both the Spin6 RNAi and mutant plants. Strikingly, SPIN6 interacts with the small GTPase OsRac1, catalyze the GTP-bound OsRac1 into the GDP-bound state in vitro and has GAP activity towards OsRac1 in rice cells. Together, our results demonstrate that the RhoGAP SPIN6 acts as a linkage between a U-box E3 ligase-mediated ubiquitination pathway and a small GTPase-associated defensome system for plant immunity.     

Detection of novel blast resistance genes, Pi58(t) and Pi59(t), in a Myanmar rice landrace based on a standard differential system

The use of broad-spectrum R genes is an effective way to achieve durable resistance against rice blast (Magnaporthe oryzae Couch, anamorph: Pyricularia oryzae Cavara) in rice (Oryza sativa L.). We previously surveyed the diversity of blast resistance in 948 rice varieties and found a Myanmar rice landrace, Haoru (International Rice Research Institute genebank acc. no. IRGC33090), with broad-spectrum resistance against the standard differential blast isolates. Here, we examined the genetic basis of Haoru’s broad-spectrum resistance by using the standard blast differential system consisting of the standard isolates and differential varieties. For genetic analysis, we used a BC1F1 population and BC1F2 lines derived from crosses of Haoru with a susceptible variety, US-2. Co-segregation analysis of the reaction pattern in the BC1F1 population against the 20 standard isolates suggested that Haoru harbors three R genes. By using bulk-segregant and linkage analysis, we mapped two of the three R genes on chromosomes 12 and 6, and designated them as Pi58(t) and Pi59(t), respectively. Pi58(t) and Pi59(t) were differentiated from other reported R genes using the standard differential system. The estimated resistance spectrum of Pi58(t) corresponded with that of Haoru, suggesting that Pi58(t) is primarily responsible for Haoru’s broad-spectrum resistance. In addition, Pi59(t) and the third gene were also proven to be new and useful genetic resources for studying and improving blast resistance in rice.     

Combination Patterns of Major R Genes Determine the Level of Resistance to the M. oryzae in Rice (Oryza sativa L.)

Rice blast caused by Magnaporthe oryzae is the most devastating disease of rice and poses a serious threat to world food security. In this study, the distribution and effectiveness of 18 R genes in 277 accessions were investigated based on pathogenicity assays and molecular markers. The results showed that most of the accessions exhibited some degree of resistance (resistance frequency, RF >50%). Accordingly, most of the accessions were observed to harbor two or more R genes, and the number of R genes harbored in accessions was significantly positively correlated with RF. Some R genes were demonstrated to be specifically distributed in the genomes of rice sub-species, such as Pigm, Pi9, Pi5 and Pi1, which were only detected in indica-type accessions, and Pik and Piz, which were just harbored in japonica-type accessions. By analyzing the relationship between R genes and RF using a multiple stepwise regression model, the R genes Pid3, Pi5, Pi9, Pi54, Pigm and Pit were found to show the main effects against M. oryzae in indica-type accessions, while Pita, Pb1, Pik, Pizt and Pia were indicated to exhibit the main effects against M. oryzae in japonica-type accessions. Principal component analysis (PCA) and cluster analysis revealed that combination patterns of major R genes were the main factors determining the resistance of rice varieties to M. oryzae, such as ‘Pi9+Pi54’, ‘Pid3+Pigm’, ‘Pi5+Pid3+Pigm’, ‘Pi5+Pi54+Pid3+Pigm’, ‘Pi5+Pid3’ and ‘Pi5+Pit+Pid3’ in indica-type accessions and ‘Pik+Pib’, ‘Pik+Pita’, ‘Pik+Pb1’, ‘Pizt+Pia’ and ‘Pizt+Pita’ in japonica-type accessions, which were able to confer effective resistance against M. oryzae. The above results provide good theoretical support for the rational utilization of combinations of major R genes in developing rice cultivars with broad-spectrum resistance.     

Genotyping and development of single-nucleotide polymorphism (SNP) markers associated with blast resistance genes in rice using GoldenGate assay

Development and large-scale genotyping of single-nucleotide polymorphism (SNP) is required to use identified sequence variation in the alleles of different genes to determine their functional relevance to the candidate gene(s). In the present study, Illumina GoldenGate assay was used to validate and genotype SNPs in a set of six major rice blast resistance genes, viz. Pi-ta, Piz(t), Pi54, Pi9, Pi5(1) and Pib, distributed over five chromosomes, to understand their functional relevance and study the population structure in rice. All the selected SNPs loci (96) of six blast (Magnaporthe oryzae) resistance genes were genotyped successfully in 92 rice lines with an overall genotype call rate of 92.0 % and minimum GenTrain cutoff score of ≥0.448. The highest genotyped SNPs were found in japonica type (97.1 %) rice lines, followed by indica (92.12 %), indica basmati (91.84 %) and minimum in case of wild species (82.0 %). Among the genotyped loci, the highest score (98.68 %) was observed in case of Piz(t), followed by Pi-ta, Pi5(1), Pib, Pi54 and Pi9. Polymorphism was obtained in 87.5 % SNPs loci producing 7,728 genotype calls. Minor allele frequency ranged from 0.01 to 0.49 and has good differentiating power for distinguishing different rice accessions. Population structure analysis revealed that a set of genotypes from four rice subpopulations had “admix” ancestry (>26 %) with more than one genetic background of indica, japonica and wild types. SNPs markers were validated in a set of 92 rice lines and converted into CAPS markers which can be used in blast resistance breeding programme.     

Surface ��-1,3-Glucan Facilitates Fungal Stealth Infection by Interfering with Innate Immunity in Plants

Plants evoke innate immunity against microbial challenges upon recognition of pathogen-associated molecular patterns (PAMPs), such as fungal cell wall chitin. Nevertheless, pathogens may circumvent the host PAMP-triggered immunity. We previously reported that the ascomycete Magnaporthe oryzae, a famine-causing rice pathogen, masks cell wall surfaces with α-1,3-glucan during invasion. Here, we show that the surface α-1,3-glucan is indispensable for the successful infection of the fungus by interfering with the plant''s defense mechanisms. The α-1,3-glucan synthase gene MgAGS1 was not essential for infectious structure development but was required for infection in M. oryzae. Lack or degradation of surface α-1,3-glucan increased fungal susceptibility towards chitinase, suggesting the protective role of α-1,3-glucan against plants'' antifungal enzymes during infection. Furthermore, rice plants secreting bacterial α-1,3-glucanase (AGL-rice) showed strong resistance not only to M. oryzae but also to the phylogenetically distant ascomycete Cochlioborus miyabeanus and the polyphagous basidiomycete Rhizoctonia solani; the histocytochemical analysis of the latter two revealed that α-1,3-glucan also concealed cell wall chitin in an infection-specific manner. Treatment with α-1,3-glucanase in vitro caused fragmentation of infectious hyphae in R. solani but not in M. oryzae or C. miyabeanus, indicating that α-1,3-glucan is also involved in maintaining infectious structures in some fungi. Importantly, rapid defense responses were evoked (a few hours after inoculation) in the AGL-rice inoculated with M. oryzae, C. miyabeanus and R. solani as well as in non-transgenic rice inoculated with the ags1 mutant. Taken together, our results suggest that α-1,3-glucan protected the fungal cell wall from degradative enzymes secreted by plants even from the pre-penetration stage and interfered with the release of PAMPs to delay innate immune defense responses. Because α-1,3-glucan is nondegradable in plants, it is reasonable that many fungal plant pathogens utilize α-1,3-glucan in the innate immune evasion mechanism and some in maintaining the structures.     

CEBiP is the major chitin oligomer-binding protein in rice and plays a main role in the perception of chitin oligomers

CEBiP, a plasma membrane-localized glycoprotein of rice, directly binds with chitin elicitors (CE), and has been identified as a receptor for CE by using CEBiP-RNAi rice cells. To further clarify the function of CEBiP, we produced CEBiP-disrupted rice plants by applying an efficient Agrobacterium-mediated gene-targeting system based on homologous recombination, which has recently been developed for rice. Homologous recombination occurred at the CEBiP locus in ~0.5 % of the positive/negative selected calli. In the self-pollinated next generation, it was confirmed that the first exon of CEBiP was replaced with the hygromycin selection cassette as designed, and that the expression of CEBiP was completely deficient in homozygous cebip lines. Affinity-labeling analysis using biotinylated N-acetylchitooctaose demonstrated that CEBiP is the major CE-binding protein in rice cultured cells and leaves, which was consistent with the result that the response to CE in cebip cells was greatly diminished. Nevertheless, we observed a significant decrease in disease resistance against Magnaporthe oryzae, the causal agent of rice blast disease, only when the cebip leaf sheaths were inoculated with a weakly virulent strain, suggesting that CE perception during the infection process of M. oryzae is limited. The response to peptidoglycan and lipopolysaccharides in cebip cells was not affected, strongly suggesting that CEBiP is a CE-specific receptor.     

Rice XB15, a protein phosphatase 2C, negatively regulates cell death and XA21-mediated innate immunity

Perception of extracellular signals by cell surface receptors is of central importance to eukaryotic development and immunity. Kinases that are associated with the receptors or are part of the receptors themselves modulate signaling through phosphorylation events. The rice (Oryza sativa L.) XA21 receptor kinase is a key recognition and signaling determinant in the innate immune response. A yeast two-hybrid screen using the intracellular portion of XA21, including the juxtamembrane (JM) and kinase domain as bait, identified a protein phosphatase 2C (PP2C), called XA21 binding protein 15 (XB15). The interaction of XA21 and XB15 was confirmed in vitro and in vivo by glutathione-S-transferase (GST) pull-down and co-immunoprecipitation assays, respectively. XB15 fusion proteins purified from Escherichia coli and from transgenic rice carry PP2C activity. Autophosphorylated XA21 can be dephosphorylated by XB15 in a temporal- and dosage-dependent manner. A serine residue in the XA21 JM domain is required for XB15 binding. Xb15 mutants display a severe cell death phenotype, induction of pathogenesis-related genes, and enhanced XA21-mediated resistance. Overexpression of Xb15 in an XA21 rice line compromises resistance to the bacterial pathogen Xanthomonas oryzae pv. oryzae. These results demonstrate that Xb15 encodes a PP2C that negatively regulates the XA21-mediated innate immune response.     

Improvement of seedling and panicle blast resistance in <Emphasis Type="Italic">Xian</Emphasis> rice varieties following <Emphasis Type="Italic">Pish</Emphasis> introgression

Rice blast is a serious disease caused by the filamentous ascomycetous fungus Magnaporthe oryzae. Incorporating disease resistance genes in rice varieties and characterizing the distribution of M. oryzae isolates form the foundation for enhancing rice blast resistance. In this study, the blast resistance gene Pish was observed to be differentially distributed in the genomes of rice sub-species. Specifically, Pish was present in 80.5% of Geng varieties, but in only 2.3% of Xian varieties. Moreover, Pish conferred resistance against only 23.5% of the M. oryzae isolates from the Geng-planting regions, but against up to 63.2% of the isolates from the Xian-planting regions. Thus, Pish may be an elite resistance gene for improving rice blast resistance in Xian varieties. Therefore, near-isogenic lines (NILs) with Pish and the polygene pyramid lines (PPLs) PPL^Pish/Pi1, PPL^Pish/Pi54, and PPL^Pish/Pi33 in the Xian background Yangdao 6 were generated using a molecular marker-assisted selection method. The results suggested that (1) Pish significantly improved rice blast resistance in Xian varieties, which exhibited considerably improved seedling and panicle blast resistance after Pish was introduced; (2) PPLs with Pish were more effective than the NILs with Pish regarding seedling and panicle blast resistance; (3) the PPL seedling and panicle blast resistance was improved by the complementary and overlapping effects of different resistance genes; and (4) the stability of NIL and PPL resistance varied under different environmental conditions, with only PPL^Pish/Pi54 exhibiting highly stable resistance in three natural disease nurseries (Jianyang, Jinggangshan, and Huangshan). This study provides new blast resistance germplasm resources and describes a novel molecular strategy for enhancing rice blast resistance.     

Differential Gene Expression Reflects Morphological Characteristics and Physiological Processes in Rice Immunity against Blast Pathogen Magnaporthe oryzae

The rice blast fungus Magnaporthe oryzae is a serious pathogen that jeopardises the world’s most important food-security crop. Ten common Malaysian rice varieties were examined for their morphological, physiological and genomic responses to this rice blast pathogen. qPCR quantification was used to assess the growth of the pathogen population in resistant and susceptible rice varieties. The chlorophyll content and photosynthesis were also measured to further understand the disruptive effects that M. oryzae has on infected plants of these varieties. Real-time PCR was used to explore the differential expression of eight blast resistance genes among the ten local varieties. Blast disease has destructive effects on the growth of rice, and the findings of our study provide evidence that the Pikh, Pi9, Pi21, and Osw45 genes are involved in defence responses in the leaves of Malaysian rice at 31 h after inoculation with M. oryzae pathotype P7.2. Both the chlorophyll content and photosynthesis were reduced, but the levels of Pikh gene expression remained constant in susceptible varieties, with a developed pathogen population and mild or severe symptoms. The Pi9, Pi21, and Osw45 genes, however, were simultaneously upregulated in infected rice plants. Therefore, the presence of the Pikh, Pi9, Pi21, and Osw45 genes in the germplasm is useful for improving the resistance of rice varieties.