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Septoria tritici blotch (STB) caused by Zymoseptoria tritici is one of the most important foliar diseases of wheat causing significant yield losses worldwide. In this study, a panel of bread wheat genotypes comprised 185 globally diverse genotypes were tested against 10 Z. tritici isolates at the seedling stage. Genome-wide association study (GWAS) using high-throughput DArTseq markers was performed and further gene expression analysis of significant markers trait association (MTAs) associated with resistance to STB was analyzed. Disease severity level showed significant differences among wheat genotypes for resistance to different Z. tritici isolates. We found novel landrace genotypes that showed highly resistance spectra to all tested isolates. GWAS analysis resulted in 19 quantitative trait loci (QTLs) for resistance to STB that were located on 14 chromosomes. Overall, 14 QTLs were overlapped with previously known QTLs or resistance genes, as well as five potentially novel QTLs on chromosomes 1A, 4A, 5B, 5D and 6D. Identified novel resistance sources and also novel QTLs for resistance to different Z. tritici isolates can use for pyramiding and development of durable resistance cultivars in future wheat breeding programs.

Fire blight caused by the gram-negative bacterium Erwinia amylovora is the most destructive disease of pome fruit trees. The pathogen induces the disease by secreting HrpN, HrpW, and DspA/E effector proteins into host tissues triggering oxidative burst. To study the role of these effectors in the course of interaction with host plants, hrpN−, hrpW− and dspA/E− mutants were inoculated on two pear cultivars under in situ (greenhouse and immature fruit) and in vitro systems. In addition, the determinantal role of these effectors on host chloroplasts was studied under activated and inactivated electron cascade of chloroplasts. Results showed that the lack of HrpN and DspA/E effectors postponed the initiation of necrosis and also decreased necrosis progress rate in comparison with the wild type strain. In contrast, hrpW− had the least decline in the pathogenicity of fire blight. The comparison of chloroplast activity confirmed the role of effectors in in situ experiments and also revealed that HrpN could be the main factor for the interaction of the pathogen with host cell chloroplasts.

Proteases are key components of the hydrolytic enzyme arsenal employed by fungal pathogens to invade their host plants. The recent advances in -omics era have facilitated identification of functional proteases involved in plant-fungus interactions. By comparison of the publically available sequences of fungal genomes we found that the number of protease genes present in the genome of Cladosporium fulvum, a biotrophic tomato pathogen, is comparable with that of hemibiotrophs. To identify host plant inducible protease genes and their products, we performed transcriptome and proteome analyses of C. fulvumin vitro and in planta by means of RNA-Seq/RT-qPCR and mass spectrometry. Transcriptome data showed that 14 out of the 59 predicted proteases are expressed during in vitro and in planta growth of C. fulvum, of which nine belong to serine proteases S8 and S10 and the rest belong to metallo- and aspartic proteases. Mass spectrometry confirmed the presence of six proteases at proteome level during plant infection. Expression of limited number of proteases by C. fulvum might sustain biotrophic growth and benefits its stealth pathogenesis.

Ascochyta blight caused by Ascochyta rabiei is one of the most important foliar diseases of chickpea that can lead to significant yield losses worldwide. A genome-wide association study (GWAS) was applied on a panel of 165 chickpea genotypes against A. rabiei isolates belonging to six different pathotypes. Large phenotypic variation in disease reaction was observed, as the frequency of resistance to pathotypes-I and III was comparatively higher than resistance to other pathotypes. Overall, among the all genotype × pathotype interactions, 259 resistance responses were identified and genotypes were grouped into four clusters based on their phenotypic reactions. The chickpea panel genotyped using DArTseq assay was used for genome-wide association mapping (GWAS). The GWAS revealed 30 marker-trait associations (MTAs) representing 17 quantitate trait loci (QTL) across all chickpea chromosomes. Among them, 11 R-QTLs were associated with resistance to specific pathotype, and 6 R-QTLs were associated with resistance to two or more pathotypes. Most of these R-QTLs overlapped with the previously reported R-QTLs for resistance to A. rabiei, whereas identified R-QTLs on chromosomes Ca1, Ca2, Ca6 and Ca7 are most likely novel. Meanwhile, annotation within the R-QTL intervals revealed genes involved in salicylic acid (SA) and jasmonic acid (JA) regulation pathways as well as cell death programming. The known and novel R-QTL identified in this study may be targeted for use in future chickpea breeding programs to recombine different loci to improve the durability of resistance to multiple A. rabiei isolates/pathotypes.

Ascochyta blight caused by Ascochyta rabiei (Pass.) Lab. (Telomorph: Didymella rabiei) (Kov.) is one of the most important fungal diseases in chickpea worldwide. Knowledge about pathogen aggressiveness and identification resistance sources to different pathotypes is very useful for proper decisions in breeding programs. In this study, virulence of 32 A. rabiei isolates from different part of Iran were analyzed on seven chickpea differentials and grouped into six races based on 0–9 rating scale and susceptibility/resistant pattern of chickpea differentials. The least and most frequent races were race V and race I, respectively. Race V and VI showed highly virulence on most of differential, while race I showed least aggressiveness. Resistance pattern of 165 chickpea genotypes also were tested against six different A. rabiei races. ANOVA analysis showed high significant difference for isolate, chickpea genotypes and their interactions. Overall chickpea × isolate (race) interactions, 259 resistance responses (disease severity ≤ 4) were identified. Resistance spectra of chickpea genotypes showed more resistance rate to race I (49.70%) and race III (35.15%), while there were no resistance genotypes to race VI. Cluster analysis based on disease severity rate, grouped chickpea genotypes into four distinct clusters. Interactions between isolates or races used in this study, showed the lack of a genotype with complete resistance. Our finding for virulence pattern of A. rabiei and newly identified resistance sources could be considerably important for integration of ascochyta blight resistance genes into chickpea breeding programs and proper decision in future for germplasm conservation and diseases management.

Background: MicroRNAs, as small non-coding RNAs, are recently reported to be involved in plant defense system against pathogens including fungi. Objective: In this research, it was intended to investigate candidate susceptible rice (Oryza Sativa) Osa-miRNA expression alteration following the infection by Rhizoctonia solani. Materials and Methods: To this aim, literature review suggested eight conserved plant miRNAs that are involved in other plant-pathogen interactions. Then, sixty days old rice plants (Hashemi, susceptible cultivar) were inoculated with R. solani and candidate miRNA expression alterations were investigated 2 hpi (hours post inoculation), 2 dpi (days post inoculation) and 6 dpi. Results: RT-qPCR analysis suggested four subgroups of candidate miRNAs based on the time of their responses to the pathogenesis of R. solani. While Osa-miR-156 was early-responsive, Osa-miR159 was the last-responsive and Osa-miR167, Osa-miR171, Osa-miR408, and Osa-miR444 were late responsive to R. solani infection. Osa-miR166 and Osa-miR393 were non-responsive to this infection, compared to the mock-inoculated control group. Consistently, Os-SPL3 and Os-MADS known target genes were expressed in reverse correlation to Osa-miR156 and Osa-miR444, respectively. Conclusions: From these data, it is suggested that both early (Osa-miR-156) and late (Osa-miR167, Osa-miR171, Osa- miR408, Osa-miR444) responsive miRNAs might be involved in R. solani infection in rice plants.

Septoria tritici blotch (STB) caused by Zymoseptoria tritici is one of the most important foliar diseases of wheat that can lead to significant yield losses worldwide. In this study, we analyzed STB resistance pattern of 185 diverse bread wheat genotypes at the seedling stage in order to confirm known resistance genes or identifying novel putative QTLs conferring resistance to STB. The phenotyping data using 10 Z. tritici isolates showed that large genetic variance exists for STB infection in wheat germplasm. Genome-wide association mapping (GWAS) using CMLM algorithm identified 37 quantitative-trait loci (QTL) for resistance to STB on 17 chromosomes. Most of QTLs overlapped with known STB resistance genes, while GWAS scan revealed several newly identified QTLs on chromosomes 4A, 5D and 6D for resistance to STB. Genome annotation of marker-trait association (MTA) against wheat reference genome revealed 29 MTAs corresponded to the putative genes, which their functional descriptions identifiers included diseases resistance proteins in plants. These MTAs and novel QTLs associated with STB resistance found here, can be used in future wheat breeding programs to recombine different loci for durable resistance to STB.

Ascochyta blight (caused by Ascochyta rabiei) is an important disease of chickpea. Mating type distribution, genetic diversity and population structure A. rabiei isolates from western Iran, using specific matting type primers, and ISSR and SSR molecular markers. Two mating types were identified, with the 57% of isolates belonging to MAT1-1. Ten ISSR markers produced 78 polymorphic bands with an average polymorphism information content (PIC) value of 0.33. Seven SSR markers showed high allelic variation (four to seven alleles) with the average PIC value of 0.61. The generated dendrogram using neighbor joining approach with ISSR and SSR marker data grouped isolates in three clusters. Combined dendrogram and model-based population structure analysis divided the isolates into two distinct populations. No significant correlation was found between geographical origins of isolates and their genetic diversity patterns, although the isolates from North Kermanshah and Kurdistan were closely grouped, and most of isolates from Lorestan and Kermanshah were clustered in a separate group. This relative spatial correlation between geographical locations and A. rabiei grouping indicated high genetic diversity within populations and no significant gene flow between distinctly geographical regions. This suggests the nece0ssity of continuous monitoring of A. rabiei populations in order to design effective chickpea breeding strategies to control the disease.

Background Sugar beet (Beta vulgaris subsp. vulgaris) is an economically important crop that provides nearly one third of the global sugar production. The beet cyst nematode (BCN), Heterodera schachtii, causes major yield losses in sugar beet and other crops worldwide. The most effective and economic approach to control this nematode is growing tolerant or resistant cultivars. To identify candidate genes involved in susceptibility and resistance, the transcriptome of sugar beet and BCN in compatible and incompatible interactions at two time points was studied using mRNA-seq. Results In the susceptible cultivar, most defense-related genes were induced at 4 dai while suppressed at 10 dai but in the resistant cultivar Nemakill, induction of genes involved in the plant defense response was observed at both time points. In the compatible interaction, alterations in phytohormone-related genes were detected. The effect of exogenous application of Methyl Jasmonate and ET-generator ethephon on susceptible plants was therefore investigated and the results revealed significant reduction in plant susceptibility. Genes putatively involved in the resistance of Nemakill were identified, such as genes involved in phenylpropanoid pathway and genes encoding CYSTM domain-containing proteins, F-box proteins, chitinase, galactono-1,4-lactone dehydrogenase and CASP-like protein. Also, the transcriptome of the BCN was analyzed in infected root samples and several novel potential nematode effector genes were found. Conclusions Our data provides detailed insights into the plant and nematode transcriptional changes occurring during compatible and incompatible interactions between sugar beet and BCN. Many important genes playing potential roles in susceptibility or resistance of sugar beet against BCN, as well as some BCN effectors with a potential role as avr proteins were identified. In addition, our findings indicate the effective role of jasmonate and ethylene in enhancing sugar beet defense response against BCN. This research provides new molecular insights into the plant-nematode interactions that can be used to design novel management strategies against BCN.

Zymoseptoria tritici is one of the most economically destructive wheat diseases all over the world and is a model fungal plant pathogen within the ascomycetes. In this study, the instrumental role of the ZtRlm1 gene encoding a MADS-box transcription factor (TF) in the infection process of Z. tritici was functionally characterized as these proteins play critical roles in the global gene regulation required for various developmental and physiological processes. Our infection assays showed that ZtRlm1 mutants were attenuated in disease development as a 30 and 90% reduction in chloro-necrotic lesions and pycnidia formation, respectively, were observed in plants inoculated with ZtRlm1 mutant strains demonstrating that ZtRlm1 is a crucial factor playing a significant role in the late stage of infection corresponding with pycnidial formation. Our expression analysis demonstrated that the transcript level of ZtRlm1 is induced at 2 and 20 days post-inoculation, coinciding with pycnidial sporulation. In addition, microscopic analyses showed that branch intensity and biomass production were significantly reduced, indicating that impaired pycnidia formation is a result of impaired differentiation and biomass production in the ZtRlm1 mutants. Furthermore, melanization, a phenomenon required for fruiting body formation, was significantly hampered in ZtRlm1 mutants as they were not melanized under all tested temperature and media conditions. Overall, our data showed that impaired disease development of the ZtRlm1 mutants is mainly due to the significant impact of ZtRlm1 in different cellular processes, including differentiation, branching, fungal biomass production, and melanization, in which identification of downstream genes are of interest to increase our understanding of this pathosystem.

The wheat leaf rust fungus, Puccinia triticina, has widespread geographical distribution in Iran within the Fertile Crescent region of the Middle East where wheat was domesticated and P. triticina originated. Therefore, it is of great importance to identify the prevalence and distribution of P. triticina pathotypes in this area. From 2010 to 2017, 241 single-uredinium isolates of P. triticina were purified from 175 collections of P. triticina made from various hosts in 14 provinces of Iran, and they were tested on 20 Thatcher near-isogenic lines carrying single-leaf rust resistance genes. In total, 86 pathotypes were identified, of which the pathotypes FDTTQ, FDKPQ, FDKTQ, and FDTNQ were most prevalent. No virulence for Lr2a was detected, whereas virulence for Lr1 was found only on bread wheat in a few provinces in 2016. Only isolates from durum wheat and wild barley were virulent to Lr28. Although virulence for Lr9, Lr20, and Lr26 was observed in some years, the virulence frequency for these genes was lower than that of the other Lr genes. P. triticina collections from host plants with different ploidy levels or genetically dissimilar backgrounds were grouped individually according to genetic distance. Based on these results, collections from barley, durum wheat, oat, triticale, and wild barley were different from those of bread wheat.

In this study, 129 wheat genotypes from globally diverse origins were genotyped using DArTseq (SilicoDArT and SNP) markers. After filtering markers for quality-filtering, 14,270 SilicoDArTs and 6484 SNPs were retained and used for genetic diversity, population structure and linkage disequilibrium analyses. The highest number of SilicoDArT and SNP markers mapped on genome A and B compared to genome D. In both marker types, polymorphism information content (PIC) values ranged from 0.1 to 0.5, while > 0.80% of SilicoDArTs and > 0.44% SNPs showed PIC value more than median (0.25%). Un-weighted Neighbor Joining cluster analysis and Bayesian-based model population structure grouped wheat genotypes into three and four clusters, respectively. Principal component analysis and discriminant analysis of principal component results showed highly match with cluster and population structure analysis. Linkage disequilibrium (LD) was more extensive in both marker types, while graphical display of LD decay for both marker types showed that LD declined in the region close to 15 kbp, where r(2)-values corresponded to r(2) = 0.16. Overall, our genetic diversity analysis showed high level of variation in studied wheat genotypes, even though there was no relationship between wheat grouping and origins. This might be attributed to admixture level that occurred during long-term natural selection of wheat genotypes in different parts of the world. Highly diverse wheat genotypes used in this study may possess unique genes and are useful sources in breeding programs to improve grain yield and quality. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s13205-019-2014-z) contains supplementary material, which is available to authorized users.

Characterization of genetic diversity, population structure, and linkage disequilibrium is a prerequisite for proper management of breeding programs and conservation of genetic resources. In this study, 186 chickpea genotypes, including advanced “Kabuli” breeding lines and Iranian landrace “Desi” chickpea genotypes, were genotyped using DArTseq-Based single nucleotide polymorphism (SNP) markers. Out of 3339 SNPs, 1152 markers with known chromosomal position were selected for genome diversity analysis. The number of mapped SNP markers varied from 52 (LG8) to 378 (LG4), with an average of 144 SNPs per linkage group. The chromosome size that was covered by SNPs varied from 16,236.36 kbp (LG8) to 67,923.99 kbp (LG5), while LG4 showed a higher number of SNPs, with an average of 6.56 SNPs per Mbp. Polymorphism information content (PIC) value of SNP markers ranged from 0.05 to 0.50, with an average of 0.32, while the markers on LG4, LG6, and LG8 showed higher mean PIC value than average. Unweighted neighbor joining cluster analysis and Bayesian-based model population structure grouped chickpea genotypes into four distinct clusters. Principal component analysis (PCoA) and discriminant analysis of principal component (DAPC) results were consistent with that of the cluster and population structure analysis. Linkage disequilibrium (LD) was extensive and LD decay in chickpea germplasm was relatively low. A few markers showed r2 ≥ 0.8, while 2961 pairs of markers showed complete LD (r2 = 1), and a huge LD block was observed on LG4. High genetic diversity and low kinship value between pairs of genotypes suggest the presence of a high genetic diversity among the studied chickpea genotypes. This study also demonstrates the efficiency of DArTseq-based SNP genotyping for large-scale genome analysis in chickpea. The genotypic markers provided in this study are useful for various association mapping studies when combined with phenotypic data of different traits, such as seed yield, abiotic, and biotic stresses, and therefore can be efficiently used in breeding programs to improve chickpea.

Characterization of genetic diversity, population structure and linkage disequilibrium is prerequisite for proper management of breeding programs and conservation of genetic resources. In this study, 186 chickpea genotypes including advanced “Kabuli” breeding lines and Iranian landrace “Desi” chickpea genotypes were genotyped using DArTseq-Based SNP markers. Out of 3339 SNPs, 1152 markers with known chromosomal position were selected for genome diversity analysis. The number of mapped SNP markers varied from 52 (LG8) to 378 (LG4), with an average of 144 SNPs per linkage group. The chromosome size that covered by SNPs varied from 16236.36 kbp (LG8) to 67923.99 kbp (LG5), while LG4 showed higher number of SNPs, with an average of 6.56 SNPs per Mbp. Polymorphism information content (PIC) value of SNP markers ranged from 0.05 to 0.50, with an average of 0.32, while the markers on LG4, LG6 and LG8 showed higher mean PIC value than average. Un-weighted Neighbor Joining cluster analysis and Bayesian-based model population structure grouped chickpea genotypes into four distinct clusters. Principal component analysis (PCoA) and Discriminant Analysis of Principal Component (DAPC) results were consistent with that of the cluster and population structure analysis. Linkage disequilibrium (LD) was extensive and LD decay in chickpea germplasm was relatively low. A few markers showed r2≥0.8, while 2961 pairs of markers showed complete LD (r2=1) and a huge LD block was observed on LG4. High genetic diversity and low kinship value between pairs of genotypes suggesting the presence of a high genetic diversity among studied chickpea genotypes. This study also demonstrated the efficiency of DArTseq-based SNP genotyping for large scale genome analysis in chickpea. The genotypic markers provided in this study are useful for various association mapping studies when combined with phenotypic data of different traits such as seed yield, abiotic and biotic stresses and therefore can be efficiently used in breeding programs to improve chickpea.

Zymoseptoria tritici is the causal agent of Septoria tritici blotch (STB) disease of wheat. Z. tritici is an apoplastic fungal pathogen, which does not penetrate plant cells at any stage of infection, and has a long initial period of symptomless leaf colonisation. During this phase it is unclear to what extent the fungus can access host plant nutrients or communicate with plant cells. Several important primary and secondary metabolite pathways in fungi are regulated by the post-translational activator phosphopantetheinyl transferase (Ppt) which provides an essential co-factor for lysine biosynthesis and the activities of non-ribosomal peptide synthases (NRPS) and polyketide synthases (PKS). To investigate the relative importance of lysine biosynthesis, NRPS-based siderophore production and PKS-based DHN melanin biosynthesis, we generated deletion mutants of ZtPpt. The ∆ZtPpt strains were auxotrophic for lysine and iron, non-melanised and non-pathogenic on wheat. Deletion of the three target genes likely affected by ZtPpt loss of function (Aar- lysine; Nrps1-siderophore and Pks1- melanin), highlighted that lysine auxotrophy was the main contributing factor for loss of virulence, with no reduction caused by loss of siderophore production or melanisation. This reveals Ppt, and the lysine biosynthesis pathway, as potential targets for fungicides effective against Z. tritici.

Fungal plant pathogens, such as Zymoseptoria tritici (formerly known as Mycosphaerella graminicola), secrete repertoires of effectors to facilitate infection or trigger host defence mechanisms. The discovery and functional characterization of effectors provides valuable knowledge that can contribute to the design of new and effective disease management strategies. Here, we combined bioinformatics approaches with expression profiling during pathogenesis to identify candidate effectors of Z.?tritici. In addition, a genetic approach was conducted to map quantitative trait loci (QTLs) carrying putative effectors, enabling the validation of both complementary strategies for effector discovery. In?planta expression profiling revealed that candidate effectors were up-regulated in successive waves corresponding to consecutive stages of pathogenesis, contrary to candidates identified by QTL mapping that were, overall, expressed at low levels. Functional analyses of two top candidate effectors (SSP15 and SSP18) showed their dispensability for Z.?tritici pathogenesis. These analyses reveal that generally adopted criteria, such as protein size, cysteine residues and expression during pathogenesis, may preclude an unbiased effector discovery. Indeed, genetic mapping of genomic regions involved in specificity render alternative effector candidates that do not match the aforementioned criteria, but should nevertheless be considered as promising new leads for effectors that are crucial for the Z.?tritici–wheat pathosystem

Zymoseptoria tritici is an economically important pathogen of wheat. However, the molecular basis of pathogenicity on wheat is still poorly understood. Here, we present a global survey of the proteins secreted by this fungus in the apoplast of resistant (cv. Shafir) and susceptible (cv. Obelisk) wheat cultivars after inoculation with reference Z. tritici strain IPO323. The fungal proteins present in apoplastic fluids were analyzed by gel electrophoresis and by data-independent acquisition liquid chromatography/mass spectrometry (LC/MSE) combined with data-dependent acquisition LC–MS/MS. Subsequent mapping mass spectrometry-derived peptide sequence data against the genome sequence of strain IPO323 identified 665 peptides in the MSE and 93 in the LC–MS/MS mode that matched to 85 proteins. The identified fungal proteins, including cell-wall degrading enzymes and proteases, might function in pathogenicity, but the functions of many remain unknown. Most fungal proteins accumulated in cv. Obelisk at the onset of necrotrophy. This inventory provides an excellent basis for future detailed studies on the role of these genes and their encoded proteins during pathogenesis in wheat.

The tan spot disease of wheat caused byHere we used ISSR and RAPD markers to study the genetic diversity of 34The leaves having the typical symptoms of tan spot disease were collected and after fungal isolation, purification and identification, DNAs were extracted. After PCR amplification using each primer, PCR products were run in agarose gels, and the resulting bands were scored. Cluster analysis was performed using Un-Wighted Nighbor Joining method.A total of 178 reproducible bands were scored. Out of which 115 (65%) were polymorphic corresponding to an average of 8 polymorphic bands per primer. The average PIC values for ISSR and RAPD markers were 0.38 and 0.43, respectively. A high degree of genetic variability among IranianGenetic variability of

Recent advances in genetic and molecular technologies gradually paved the way for the transition from traditional fungal karyotyping to more comprehensive chromosome biology studies. Extensive chromosomal polymorphisms largely resulting from chromosomal rearrangements (CRs) are widely documented in fungal genomes. These extraordinary CRs in fungi generate substantial genome plasticity compared to other eukaryotic organisms. Here, we review the most recent findings on fungal CRs and their underlying mechanisms and discuss the functional consequences of CRs for adaptation, fungal evolution, host range, and pathogenicity of fungal plant pathogens in the context of chromosome biology. In addition to a complement of permanent chromosomes called core chromosomes, the genomes of many fungal pathogens comprise distinct unstable chromosomes called dispensable chromosomes (DCs) that also contribute to chromosome polymorphisms. Compared to the core chromosomes, the structural features of DCs usually differ for gene density, GC content, housekeeping genes, and recombination frequency. Despite their dispensability for normal growth and development, DCs have important biological roles with respect to pathogenicity in some fungi but not in others. Therefore, their evolutionary origin is also reviewed in relation to overall fungal physiology and pathogenicity.

Septoria tritici blotch (STB), caused by Zymoseptoria tritici, is an important disease of wheat worldwide. This study was performed to investigate the efficacy of Stb genes against 10 isolates of Z. tritici and to elucidate the resistance response pattern of 185 wheat genotypes to STB. The 10 isolates showed different virulence patterns on the Stb differentials. Effectiveness determination showed that most Stb genes (particularly Stb1–Stb9) were ineffective against the 10 isolates, whereas Kavkaz-K4500 (possessing Stb10, Stb12), Arina and Riband (possessing Stb15), and M3 (possessing Stb16 and Stb17) were resistant to all isolates tested. Of the 185 wheat genotypes, 72% were susceptible to all isolates, indicating that these genotypes lacked any effective resistance genes at the seedling stage against the isolates used. The remaining 51 genotypes showed specific resistance to one or more isolates, suggesting that they contain at least one effective resistance gene. Six genotypes were resistant to all isolates, indicating that they may possess broad-spectrum resistance gene(s) or a combination of diverse uncharacterised Stb genes that could be effectively used in breeding programs.

Septoria tritici blotch (STB), caused by Zymoseptoria tritici has been considered one of the important diseases of wheat in Europe and elsewhere including Iran. The most economical strategy to control the disease is the use of genetic resistance. To this aim, this study was conducted to identify new sources of resistance to STB and to investigate the effectiveness of known Stb genes against six Iranian isolates of Z. tritici. Genetic variability analysis using random amplified polymorphic DNA (RAPD) markers revealed high polymorphic banding patterns indicating that a high genetic variability existed among the isolates. Inoculation assays identified 116 isolate-specific resistances among all interactions (n = 420). Of 52 genotypes, 26 wheat genotypes showed no isolate-specific responses and were susceptible to all isolates, which may explain increasing epidemics of Z. tritici occurring in major wheat-growing regions in Iran. Five genotypes were highly resistant to all isolates. Effectiveness determination of Stb genes revealed that Iranian isolates have a broad virulence spectrum against most of the known Stb genes. Among all Stb genes, Stb15 and Stb16/Stb17 in cultivars Arina, Riband and M3 were the most effective resistance genes and conferred resistance to all isolates tested. Hence, these genes should be effective in breeding programs in Iran to control the disease.

Graphical abstract Drought stress increased the amount of glycyrrhizin in the stolons of Glycyrrhiza glabra. The expression of sequalene synthase (SQS) and β-amyrin synthase (bAS) genes were increased under drought stress, whereas expression of cycloartenol synthase (CAS) was not changed., Highlights • We applied various drought stresses in liquorice. • Drought stress increased the expression of SQS and bAS, whereas expression of CAS was not changed. • Drought stress increased the amount of glycyrrhizin in the stolons of G. glabra. • Applying special irrigation regimes can principally improve secondary metabolite production., Glycyrrhiza glabra is an important medicinal plant throughout the world. Glycyrrhizin is a triterpenoid that is among the most important secondary metabolites produced by liquorice. Drought stress is proposed to enhance the levels of secondary metabolites. In this study, the effect of drought stress on the expression of important genes involved in the glycyrrhizin biosynthetic pathway was examined. Drought stress at the seedling stage was applied to 8-day-old plants using polyethylene glycol. Subsequently, the samples were collected 0, 4, 8 or 24 h post-treatment. At the adult plant stage, 10-month-old plants were subjected to drought stress by discontinuing irrigation. Subsequently, samples were collected at 2, 16 and 28 days after drought imposition (S2d, S16d and S28d, respectively). We performed semi-quantitative RT-PCR assays to evaluate the gene expression levels of sequalene synthase (SQS), β-amyrin synthase (bAS), lupeol synthase (LUS) and cycloartenol synthase (CAS) during stress. Finally, the glycyrrhizin content of stolons was determined via HPLC. The results revealed that due to osmotic stress, the gene expression levels of SQS and bAS were increased, whereas those of CAS were relatively unchanged at the seedling stage. At the adult plant stage, the expression levels of SQS and bAS were increased under drought stress conditions, whereas the gene expression level of CAS remained relatively constant. The glycyrrhizin content in stolons was increased only under severe drought stress conditions (S28d). Our results indicate that application of controlled drought stress up-regulates the expression of key genes involved in the biosynthesis of triterpenoid saponins and directly enhances the production of secondary metabolites, including glycyrrhizin, in liquorice plants.

Twenty-nine synthetic hexaploid wheats (SHWs) were evaluated for resistance to five isolates of Zymoseptoria tritici, a devastating wheat pathogen worldwide. The five Z. tritici isolates varied in their virulence spectra towards wheat genotypes, indicating that they have distinct set of avirulence genes. New isolate-specific resistances were identified that could be used in wheat breeding programmes. Comparing with the previous studies, the number of specific resistances identified in this study is considerable. Among 150 interactions, 78 isolate-specific resistances were identified. Interestingly, 21 wheat genotypes showed specific responses to one or more isolates tested. Of these, 12 genotypes were highly resistant to all isolates, indicating that they possess known or novel effective resistance genes. The Stb15 and Stb16/Stb17 are effective resistance genes towards isolates used in this study, indicating that the conferred resistance in these genotypes is due to the presence of either of these genes in combination or individually. Alternatively, they may carry novel broad-spectrum resistance gene(s) that their identification is of interest. Our data suggest that the presence of complete resistance to various Z. tritici isolates in SHWs justifies the need for more in-depth research to characterize the likely novel genes.

Characterization of genetic diversity, population structure, and linkage disequilibrium is a prerequisite for proper management of breeding programs and conservation of genetic resources. In this study, 186 chickpea genotypes, including advanced &ldquo, Kabuli&rdquo, breeding lines and Iranian landrace &ldquo, Desi&rdquo, chickpea genotypes, were genotyped using DArTseq-Based single nucleotide polymorphism (SNP) markers. Out of 3339 SNPs, 1152 markers with known chromosomal position were selected for genome diversity analysis. The number of mapped SNP markers varied from 52 (LG8) to 378 (LG4), with an average of 144 SNPs per linkage group. The chromosome size that was covered by SNPs varied from 16,236.36 kbp (LG8) to 67,923.99 kbp (LG5), while LG4 showed a higher number of SNPs, with an average of 6.56 SNPs per Mbp. Polymorphism information content (PIC) value of SNP markers ranged from 0.05 to 0.50, with an average of 0.32, while the markers on LG4, LG6, and LG8 showed higher mean PIC value than average. Unweighted neighbor joining cluster analysis and Bayesian-based model population structure grouped chickpea genotypes into four distinct clusters. Principal component analysis (PCoA) and discriminant analysis of principal component (DAPC) results were consistent with that of the cluster and population structure analysis. Linkage disequilibrium (LD) was extensive and LD decay in chickpea germplasm was relatively low. A few markers showed r2 &ge, 0.8, while 2961 pairs of markers showed complete LD (r2 = 1), and a huge LD block was observed on LG4. High genetic diversity and low kinship value between pairs of genotypes suggest the presence of a high genetic diversity among the studied chickpea genotypes. This study also demonstrates the efficiency of DArTseq-based SNP genotyping for large-scale genome analysis in chickpea. The genotypic markers provided in this study are useful for various association mapping studies when combined with phenotypic data of different traits, such as seed yield, abiotic, and biotic stresses, and therefore can be efficiently used in breeding programs to improve chickpea.

Septoria tritici blotch (STB) caused by Mycosphaerella graminicola is a serious disease in some wheat-growing regions of Iran. Continuous monitoring of the fungus populations is indispensable to study the efficiency of known resistant cultivars and Stb genes. In this study, resistance of 49 wheat genotypes, including 20 wheat differential cultivars worldwide and 29 other wheat genotypes, were studied to 19 M. graminicola isolates. Among 931 genotype–isolate interactions, 224 isolate-specific resistances (26%) were observed by comparing the means of disease severity data using LSD1% and LSD5%. Iranian M. graminicola isolates showed new virulence patterns. All isolates were virulent to Stb2, Stb5, Stb6, Stb7, Stb13 and Stb14. Among wheat differentials, M3 was resistant to all isolates, indicating that it has gene(s) with the most efficiency against Iranian isolates. Among other genotypes, two genotypes were resistant to all isolates. This suggests that these genotypes have efficient resistance gene(s) to Ir...

Zymoseptoria tritici causes the major fungal wheat disease septoria tritici blotch, and is increasingly being used as a model for transmission and population genetics, as well as host–pathogen interactions. Here, we study the biological function of ZtWor1, the orthologue of Wor1 in the fungal human pathogen Candida albicans, as a representative of a superfamily of regulatory proteins involved in dimorphic switching. In Z.?tritici, this gene is pivotal for pathogenesis, as ZtWor1 mutants were nonpathogenic and complementation restored the wild-type phenotypes. In?planta expression analyses showed that ZtWor1 is up-regulated during the initiation of colonization and fructification, and regulates candidate effector genes, including one that was discovered after comparative proteome analysis of the Z.?tritici wild-type strain and the ZtWor1 mutant, which was particularly expressed in?planta. Cell fusion and anastomosis occur frequently in ZtWor1 mutants, reminiscent of mutants of MgGpb1, the s-subunit of the heterotrimeric G protein. Comparative expression of ZtWor1 in knock-out strains of MgGpb1 and MgTpk2, the catalytic subunit of protein kinase A, suggests that ZtWor1 is downstream of the cyclic adenosine monophosphate (cAMP) pathway that is crucial for pathogenesis in many fungal plant pathogens

Black Sigatoka or black leaf streak disease, caused by the Dothideomycete fungus Pseudocercospora fijiensis (previously: Mycosphaerella fijiensis), is the most significant foliar disease of banana worldwide. Due to the lack of effective host resistance, management of this disease requires frequent fungicide applications, which greatly increase the economic and environmental costs to produce banana. Weekly applications in most banana plantations lead to rapid evolution of fungicide-resistant strains within populations causing disease-control failures throughout the world. Given its extremely high economic importance, two strains of P. fijiensis were sequenced and assembled with the aid of a new genetic linkage map. The 74-Mb genome of P. fijiensis is massively expanded by LTR retrotransposons, making it the largest genome within the Dothideomycetes. Melting-curve assays suggest that the genomes of two closely related members of the Sigatoka disease complex, P. eumusae and P. musae, also are expanded. Electrophoretic karyotyping and analyses of molecular markers in P. fijiensis field populations showed chromosome-length polymorphisms and high genetic diversity. Genetic differentiation was also detected using neutral markers, suggesting strong selection with limited gene flow at the studied geographic scale. Frequencies of fungicide resistance in fungicide-treated plantations were much higher than those in untreated wild-type P. fijiensis populations. A homologue of the Cladosporium fulvum Avr4 effector, PfAvr4, was identified in the P. fijiensis genome. Infiltration of the purified PfAVR4 protein into leaves of the resistant banana variety Calcutta 4 resulted in a hypersensitive-like response. This result suggests that Calcutta 4 could carry an unknown resistance gene recognizing PfAVR4. Besides adding to our understanding of the overall Dothideomycete genome structures, the P. fijiensis genome will aid in developing fungicide treatment schedules to combat this pathogen and in improving the efficiency of banana breeding programs., Author Summary Black Sigatoka or black leaf streak disease, caused by the ascomycete fungus Pseudocercospora fijiensis, inflicts huge costs on banana producers, due to crop losses and expenses for disease control. The global banana export trade relies on Cavendish clones that are highly susceptible to P. fijiensis. Sustainable production of the world’s number one fruit requires a better understanding of host resistance and sophisticated management of fungicide resistance in the pathogen. In the P. fijiensis genome sequence we identified an effector that induced an HR-like necrosis on a resistant banana accession but not on a susceptible cultivar. If validated, this assay may be useful for identifying resistance in banana breeding programs. We also used the genomic sequence to develop highly polymorphic molecular markers for analyzing P. fijiensis field populations and identified a strong enrichment (nearly 100%) for fungicide resistance markers in fungicide-treated banana plantations compared to untreated wild-type populations. This rapid evolution of fungicide resistance poses an immediate threat to sustainable banana production.

Septoria tritici blotch (STB), caused by Mycosphaerella graminicola has recently become one of the devastating diseases in Iran causing significant yield losses on most commercial bread and durum wheat cultivars. Iran is located in the Fertile Crescent, a region where wheat was historically domesticated; and, thus, landraces derived from this region are of considerable global interest for identification of new sources of resistance to various stresses. Here, we report on the resistance responses of 45 tetraploid wheat landraces collected from different provinces of Iran to eight M. graminicola isolates. In total 138 isolate-specific resistances were found among all interactions (n = 360). The highest number of specific resistances (30 out of 32 interactions) was found in wheat landraces collected from West Azarbaijan. In contrast, all landraces from Kordestan were highly susceptible to M. graminicola isolates and only one isolate-specific resistance was identified among 106 isolate-wheat interactions. Kermanshah landraces showed the highest resistance variation against different isolates. About 57 isolate-specific resistances were identified among 104 interactions. Ilam landraces were highly resistant to STB as 28 specific resistances were observed among 32 interactions. Markazi (n = 2) and Sistan-Baluchestan (n = 1) were susceptible to all isolates tested. Landraces from Lorestan were generally susceptible to isolates tested as 26 susceptible responses were observed out of 32 interactions. Our results indicate that landraces collected from the Fertile Crescent region may possess diverse effective resistance genes or valuable broad spectrum resistance genes, and that their identification is of interest and can be exploited in breeding programs.

Co-evolution of wheat and its devastating pathogen Mycosphaerella graminicola (anamorph Septoria tritici), the causal agent of septoria tritici blotch, a foliar disease of wheat, is suggested to occur in Fertile Crescent as their center of origin and, thus, interaction between pathogen virulence and host resistance is important subject to be addressed. We have investigated resistance spectra of 54 wheat genotypes including a set of differentials carrying known resistance genes and virulence patterns of 14 M. graminicola isolates at seedling stage under controlled environmental conditions. The isolates were collected in Iran from five provinces. Diversity in virulence and aggressiveness was observed among the isolates from four provinces. Isolates collected from Golestan province were virulent to all wheat genotypes from germplasm of Iran, while specific resistances were identified to the isolates from other provinces. Among wheat genotypes, cvs. Chamran, Morvarid and Hirmand had the greatest number of specific resistances as well as partial resistance. Wheat genotypes of the differential set also differed in their reactions to the isolates. Arina, Flame and TE 9111 were specifically resistant to the greatest number of isolates from different provinces. Most isolates were virulent to the other differentials such as cvs. Shafir, Estanzuela federal and Courtot indicating that extensive adaption of virulence to most of the known resistance genes (Stb) has occurred in these regions. The new sources of resistance to highly virulent isolates from Iran may also be utilized in wheat breeding programs to develop resistant cultivars against pathogen populations in other countries.

The Mycosphaerella complex is both poly- and paraphyletic, containing several different families and genera. The genus Mycosphaerella is restricted to species with Ramularia anamorphs, while Septoria is restricted to taxa that cluster with the type species of Septoria, S. cytisi, being closely related to Cercospora in the Mycosphaerellaceae. Species that occur on graminicolous hosts represent an as yet undescribed genus, for which the name Zymoseptoria is proposed. Based on the 28S nrDNA phylogeny derived in this study, Zymoseptoria is shown to cluster apart from Septoria. Morphologically species of Zymoseptoria can also be distinguished by their yeast-like growth in culture, and the formation of different conidial types that are absent in Septoria s.str. Other than the well-known pathogens such as Z. tritici, the causal agent of septoria tritici blotch on wheat, and Z. passerinii, the causal agent of septoria speckled leaf blotch of barley, both for which epitypes are designated, two leaf blotch pathogens are also described on graminicolous hosts from Iran. Zymoseptoria brevis sp. nov. is described from Phalaris minor, and Z. halophila comb. nov. from leaves of Hordeum glaucum. Further collections are now required to elucidate the relative importance, host range and distribution of these species.

To provide insight into the genetic structure of Mycosphaerella graminicola populations in Iran, a total of 221 isolates were collected from naturally infected wheat fields of five major wheat-growing provinces and analysed using AFLP markers and mating-type loci. All populations showed intermediate to high genotypic diversity. In the Golestan and Ardabil populations two mating types were found at near-equal frequencies, whilst all populations were in gametic disequilibrium. Moreover, clonal haplotypes were identified in different sampling sites within a field in both the Khuzestan and Fars provinces, demonstrating that pycnidia are probably the primary source of inoculum. All five populations had low levels of gene diversity and had private bands. Low levels of gene flow and high genetic differentiation were observed among populations and different clustering methods revealed five genetically distinct groups in accordance with the sampling areas. The Golestan and East Azarbaijan populations were more genetically differentiated than the others. Random genetic drift, selection and geographic barriers may account for the differentiation of the populations. The results of this study indicate a population structure of M. graminicola in Iran contrasting to that of most other countries studied.

Head blight or scab caused by Fusarium graminearum is a destructive disease of wheat and barley. The pathogen can cause severe yield losses and contaminates infested kernels with harmful mycotoxins. In this study, we characterized the CID1 gene in F. graminearum that is an ortholog of the Fusarium verticilloidesFCC1 and yeast UME3 genes. The protein encoded by CID1 has typical structural features of C-type cyclins. Deletion of CID1 resulted in a reduction in conidiation and vegetative growth but an increase in pigmentation. The Deltacid1 mutant was female sterile but could outcross as a male. It was significantly reduced in DON production and virulence on wheat heads and corn stalks. Only about 50% of inoculated spikelets developed scab symptoms and scab disease rarely extended to nearby florets, suggesting that the Deltacid1 mutant was defective in colonizing and spreading in wheat heads. Deletion of CID1 resulted in reduced expression levels of TRI5 and TRI101 but increased PKS12 expression. When expressed in F. verticillioides, the CID1 gene complemented the defects of the Deltafcc1 mutant in conidiation, hyphal growth, and fumonisin production. Our data indicate that the CID1 C-type cyclin gene plays multiple roles in the regulation of vegetative growth, sexual development, conidiation, DON production, and pathogenicity.

The pioneering research of Harold Flor on flax and the flax rust fungus culminated in his gene‐for‐gene hypothesis. It took nearly 50 years before the first fungal avirulence (Avr) gene in support of his hypothesis was cloned. Initially, fungal Avr genes were identified by reverse genetics and map‐based cloning from model organisms, but, currently, the availability of many sequenced fungal genomes allows their cloning from additional fungi by a combination of comparative and functional genomics. It is believed that most Avr genes encode effectors that facilitate virulence by suppressing pathogen‐associated molecular pattern‐triggered immunity and induce effector‐triggered immunity in plants containing cognate resistance proteins. In resistant plants, effectors are directly or indirectly recognized by cognate resistance proteins that reside either on the plasma membrane or inside the plant cell. Indirect recognition of an effector (also known as the guard model) implies that the virulence target of an effector in the host (the guardee) is guarded by the resistance protein (the guard) that senses manipulation of the guardee, leading to activation of effector‐triggered immunity. In this article, we review the literature on fungal effectors and some pathogen‐associated molecular patterns, including those of some fungi for which no gene‐for‐gene relationship has been established.