RESUMO
Pyrenophora tritici-repentis (tan spot) is a destructive foliar pathogen of wheat with global impact. This ascomycete fungus possesses a highly plastic open pangenome shaped by the gain and loss of effector genes. This study investigated the allelic variations in the chlorosis-encoding gene ToxB across 422 isolates representing all identified pathotypes and worldwide origins. To gain better insights into ToxB evolution, we examined its presence and variability in other Pyrenophora spp. A ToxB haplotype network was constructed, revealing the evolutionary relationships of this gene (20 haplotypes) across four Pyrenophora species. Notably, toxb, the homolog of ToxB, was detected for the first time in the barley pathogen Pyrenophora teres. The ToxB/toxb genes display evidence of selection that is characterized by loss of function, duplication, and diverse mutations. Within the ToxB/toxb open reading frame, 72 mutations were identified, including 14 synonymous, 55 nonsynonymous, and 3 indel mutations. Remarkably, a, â¼5.6-kb Copia-like retrotransposon, named Copia-1_Ptr, was found inserted in the toxb gene of a race 3 isolate. This insert disrupted the ToxB gene's function, a first case of effector gene disruption by a transposable element in P. tritici-repentis. Additionally, a microsatellite with 25 nucleotide repeats (0 to 10) in the upstream region of ToxB suggested a potential mechanism influencing ToxB expression and regulation. Exploring ToxB-like protein distribution in other ascomycetes revealed the presence of ToxB-like proteins in 19 additional species, including the Leotiomycetes class for the first time. The presence/absence pattern of ToxB-like proteins defied species relatedness compared with a phylogenetic tree, suggesting a past horizontal gene transfer event during the evolution of the ToxB gene. [Formula: see text] Copyright © 2024 His Majesty the King in Right of Canada, as represented by the Minister of Agriculture and Agri-Food. This is an open access article distributed under the CC BY-NC-ND 4.0 International license.
Assuntos
Ascomicetos , Proteínas Fúngicas , Filogenia , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Ascomicetos/genética , Ascomicetos/metabolismo , Triticum/genética , Triticum/microbiologiaRESUMO
The global wheat disease tan spot is caused by the necrotrophic fungal pathogen Pyrenophora tritici-repentis (Ptr) which secretes necrotrophic effectors to facilitate host plant colonization. We previously reported a role of the Zn2 Cys6 binuclear cluster transcription factor Pf2 in the regulation of the Ptr effector ToxA. Here, we show that Pf2 is also a positive regulator of ToxB, via targeted deletion of PtrPf2 which resulted in reduced ToxB expression and defects in conidiation and pathogenicity. To further investigate the function of Ptr Pf2 in regulating protein secretion, the secretome profiles of two Δptrpf2 mutants of two Ptr races (races 1 and 5) were evaluated using a SWATH-mass spectrometry (MS) quantitative approach. Analysis of the secretomes of the Δptrpf2 mutants from in vitro culture filtrate identified more than 500 secreted proteins, with 25% unique to each race. Of the identified proteins, less than 6% were significantly differentially regulated by Ptr Pf2. Among the downregulated proteins were ToxA and ToxB, specific to race 1 and race 5 respectively, demonstrating the role of Ptr Pf2 as a positive regulator of both effectors. Significant motif sequences identified in both ToxA and ToxB putative promoter regions were further explored via GFP reporter assays.
Assuntos
Ascomicetos , Micotoxinas , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Secretoma , Ascomicetos/metabolismo , Triticum/metabolismo , Triticum/microbiologia , Doenças das Plantas/microbiologia , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Micotoxinas/metabolismoRESUMO
The necrotrophic effector ToxA is a well-studied virulence factor produced by several fungal necrotrophs. Initially cloned from the wheat tan spot pathogen Pyrenophora tritici-repentis in 1996, ToxA was found almost a decade later in another fungal pathogen, Parastagonospora nodorum, and its sister species, Parastagonospora pseudonodorum. In 2018, ToxA was detected in a third wheat fungal pathogenic species, Bipolaris sorokiniana, which causes spot blotch disease. However, unlike the case with P. tritici-repentis and P. nodorum, the ToxA in B. sorokiniana has only been investigated in recent years. In this report, five Australian B. sorokiniana isolates were assessed for the presence of ToxA. Four isolates were found to contain ToxA. While one isolate harbored the previously reported ToxA haplotype sequence (ToxA19), three isolates contain a different haplotype, designated herein as ToxA25, which has a nonsynonymous mutation resulting in an amino acid change of glycine to arginine at position 168. Both B. sorokiniana ToxA isoforms, when heterologously expressed in Escherichia coli, exhibited the classic ToxA necrosis-inducing activity on ToxA-sensitive Tsn1 cultivars. Preliminary analysis of the B. sorokiniana isolates in Australian wheat cultivars showed that isolates with ToxA19, ToxA25, or ToxA-deficient displayed various degrees of virulence, with the most aggressive isolates observed for those producing ToxA. Differences in spot blotch disease severity between Tsn1 and tsn1 cultivars were observed; however, this was not limited to the ToxA-producing isolates. The overall results suggest that the virulence of the Australian B. sorokiniana isolates is diverse, with the significance of ToxA-Tsn1 interactions depending on individual isolates.
Assuntos
Bipolaris , Proteínas Fúngicas , Haplótipos , Micotoxinas , Doenças das Plantas , Triticum , Triticum/microbiologia , Doenças das Plantas/microbiologia , Micotoxinas/genética , Micotoxinas/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Bipolaris/genética , Ascomicetos/genética , Ascomicetos/patogenicidade , Austrália , Fatores de Virulência/genética , Virulência/genéticaRESUMO
KEY MESSAGE: Novel sources of genetic resistance to tan spot in Australia have been discovered using one-step GWAS and genomic prediction models that accounts for additive and non-additive genetic variation. Tan spot is a foliar disease in wheat caused by the fungal pathogen Pyrenophora tritici-repentis (Ptr) and has been reported to generate up to 50% yield losses under favourable disease conditions. Although farming management practices are available to reduce disease, the most economically sustainable approach is establishing genetic resistance through plant breeding. To further understand the genetic basis for disease resistance, we conducted a phenotypic and genetic analysis study using an international diversity panel of 192 wheat lines from the Maize and Wheat Improvement Centre (CIMMYT), the International Centre for Agriculture in the Dry Areas (ICARDA) and Australian (AUS) wheat research programmes. The panel was evaluated using Australian Ptr isolates in 12 experiments conducted in three Australian locations over two years, with assessment for tan spot symptoms at various plant development stages. Phenotypic modelling indicated high heritability for nearly all tan spot traits with ICARDA lines displaying the greatest average resistance. We then conducted a one-step whole-genome analysis of each trait using a high-density SNP array, revealing a large number of highly significant QTL exhibiting a distinct lack of repeatability across the traits. To better summarise the genetic resistance of the lines, a one-step genomic prediction of each tan spot trait was conducted by combining the additive and non-additive predicted genetic effects of the lines. This revealed multiple CIMMYT lines with broad genetic resistance across the developmental stages of the plant which can be utilised in Australian wheat breeding programmes to improve tan spot disease resistance.
Assuntos
Locos de Características Quantitativas , Triticum , Triticum/genética , Triticum/microbiologia , Mapeamento Cromossômico , Resistência à Doença/genética , Melhoramento Vegetal , Austrália , Doenças das Plantas/genética , Doenças das Plantas/microbiologiaRESUMO
Tan spot disease is caused by Pyrenophora tritici-repentis (Ptr), one of the major necrotrophic fungal pathogens that affects wheat crops globally. Extensive research has shown that the necrotrophic fungal effectors ToxA, ToxB, and ToxC underlie the genetic interactions of Ptr race classification. ToxA and ToxB are both small proteins secreted during infection; however, the structure of ToxC remains unknown. In line with the recent discovery of the ToxC1 gene that is involved in ToxC production, a subset of 68 isolates collected from the Australian wheat cropping regions were assessed for the presence of all three effectors by pathotyping against four tan spot wheat differential lines and PCR amplification of ToxA, ToxB, and ToxC1. Based on the disease phenotypes, the 68 isolates were grouped into two races with 63 classified as race 1 and five as race 2. A representative selection of each race was tested against eight Australian commercial wheat cultivars and showed no distinction between the virulence levels. Sequencing of ToxA showed that both races had identical gene sequences of haplotype PtrA1. All the race 1 isolates possessed ToxC1 but three race 2 isolates also contained ToxC1 despite being unable to induce a spreading chlorotic symptom on the ToxC differential line. Quantitative trait loci mapping confirmed the absence of the ToxC-Tsc1 association in disease response caused by the ToxC1-containing race 2 isolate; however, ToxC1 expression was detected during plant infection. Altogether, these results suggest that there is a complex regulatory process involved in the production of ToxC within the Australian race 2 isolates.
Assuntos
Ascomicetos , Doenças das Plantas , Doenças das Plantas/microbiologia , Austrália , Locos de Características Quantitativas , Ascomicetos/genética , Triticum/genética , Triticum/microbiologiaRESUMO
ToxA is one of the most studied proteinaceous necrotrophic effectors produced by plant pathogens. It has been identified in four pathogens (Pyrenophora tritici-repentis, Parastagonospora nodorum, Parastagonospora pseudonodorum [formerly Parastagonospora avenaria f. sp. tritici], and Bipolaris sorokiniana) causing leaf spot diseases on cereals worldwide. To date, 24 different ToxA haplotypes have been identified. Some P. tritici-repentis and related species also express ToxB, another small protein necrotrophic effector. We present here a revised and standardized nomenclature for these effectors, which could be extended to other poly-haplotypic genes found across multiple species.
Assuntos
Proteínas Fúngicas , Micotoxinas , Haplótipos , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Doenças das Plantas/microbiologia , Micotoxinas/genéticaRESUMO
The fungus Pyrenophora tritici-repentis causes tan spot, an important foliar disease of wheat worldwide. The fungal pathogen produces three necrotrophic effectors, namely Ptr ToxA, Ptr ToxB, and Ptr ToxC to induce necrosis or chlorosis in wheat. Both Ptr ToxA and Ptr ToxB are proteins, and their encoding genes have been cloned. Ptr ToxC was characterized as a low-molecular weight molecule 20 years ago but the one or more genes controlling its production in P. tritici-repentis are unknown. Here, we report the genetic mapping, molecular cloning, and functional analysis of a fungal gene that is required for Ptr ToxC production. The genetic locus controlling the production of Ptr ToxC, termed ToxC, was mapped to a subtelomeric region using segregating biparental populations, genome sequencing, and association analysis. Additional marker analysis further delimited ToxC to a 173-kb region. The predicted genes in the region were examined for presence/absence polymorphism in different races and isolates leading to the identification of a single candidate gene. Functional validation showed that this gene was required but not sufficient for Ptr ToxC production, thus it is designated as ToxC1. ToxC1 encoded a conserved hypothetical protein likely located on the vacuole membrane. The gene was highly expressed during infection, and only one haplotype was identified among 120 isolates sequenced. Our work suggests that Ptr ToxC is not a protein and is likely produced through a cascade of biosynthetic pathway. The identification of ToxC1 is a major step toward revealing the Ptr ToxC biosynthetic pathway and studying its molecular interactions with host factors.[Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.
Assuntos
Ascomicetos , Doenças das Plantas , Ascomicetos/genética , Mapeamento Cromossômico , Doenças das Plantas/microbiologia , Triticum/genética , Triticum/microbiologiaRESUMO
Pyrenophora tritici-repentis is an ascomycete fungus that causes tan spot of wheat. The disease has a worldwide distribution and can cause significant yield and quality losses in wheat production. The fungal pathogen is homothallic in nature, which means it can undergo sexual reproduction by selfing to produce pseudothecia on wheat stubble for seasonal survival. Since homothallism precludes the development of bi-parental fungal populations, no genetic linkage map has been developed for P. tritici-repentis for mapping and map-based cloning of fungal virulence genes. In this work, we created two heterothallic strains by deleting one of the mating type genes in each of two parental isolates 86-124 (race 2) and AR CrossB10 (a new race) and developed a bi-parental fungal population between them. The draft genome sequences of the two parental isolates were aligned to the Pt-1C-BFP reference sequence to mine single nucleotide polymorphisms (SNPs). A total of 225 SNP markers were developed for genotyping the entire population. Additionally, 75 simple sequence repeat, and two gene markers were also developed and used in the genotyping. The resulting linkage map consisted of 13 linkage groups spanning 5,075.83 cM in genetic distance. Because the parental isolate AR CrossB10 is a new race and produces Ptr ToxC, it was sequenced using long-read sequencing platforms and de novo assembled into contigs. The majority of the contigs were further anchored into chromosomes with the aid of the linkage maps. The whole genome comparison of AR CrossB10 to the reference genome of M4 revealed a few chromosomal rearrangements. The genetic linkage map and the new AR CrossB10 genome sequence are valuable tools for gene cloning in P. tritici-repentis.
Assuntos
Ascomicetos/genética , Proteínas Fúngicas/genética , Ligação Genética , Micotoxinas/genética , Mapeamento Cromossômico , Marcadores Genéticos , Interações Hospedeiro-Patógeno/genética , Doenças das Plantas/microbiologia , Polimorfismo de Nucleotídeo Único , Virulência/genéticaRESUMO
KEY MESSAGE: QTL mapping identified key genomic regions associated with adult-plant resistance to tan spot, which are effective even in the presence of the sensitivity gene Tsn1, thus serving as a new genetic solution to develop disease-resistant wheat cultivars. Improving resistance to tan spot (Pyrenophora tritici-repentis; Ptr) in wheat by eliminating race-specific susceptibility genes is a common breeding approach worldwide. The potential to exploit variation in quantitative forms of resistance, such as adult-plant resistance (APR), offers an alternative approach that could lead to broad-spectrum protection. We previously identified wheat landraces in the Vavilov diversity panel that exhibited high levels of APR despite carrying the sensitivity gene Tsn1. In this study, we characterised the genetic control of APR by developing a recombinant inbred line population fixed for Tsn1, but segregating for the APR trait. Linkage mapping using DArTseq markers and disease response phenotypes identified a QTL associated with APR to Ptr race 1 (producing Ptr ToxA- and Ptr ToxC) on chromosome 2B (Qts.313-2B), which was consistently detected in multiple adult-plant experiments. Additional loci were also detected on chromosomes 2A, 3D, 5A, 5D, 6A, 6B and 7A at the seedling stage, and on chromosomes 1A and 5B at the adult stage. We demonstrate that Qts.313-2B can be combined with other adult-plant QTL (i.e. Qts.313-1A and Qts.313-5B) to strengthen resistance levels. The APR QTL reported in this study provide a new genetic solution to tan spot in Australia and could be deployed in wheat cultivars, even in the presence of Tsn1, to decrease production losses and reduce the application of fungicides.
Assuntos
Ascomicetos/fisiologia , Cromossomos de Plantas/genética , Resistência à Doença/imunologia , Doenças das Plantas/imunologia , Proteínas de Plantas/metabolismo , Locos de Características Quantitativas , Triticum/genética , Mapeamento Cromossômico/métodos , Resistência à Doença/genética , Regulação da Expressão Gênica de Plantas , Interações Hospedeiro-Patógeno , Fenótipo , Melhoramento Vegetal , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Proteínas de Plantas/genética , Triticum/crescimento & desenvolvimento , Triticum/microbiologiaRESUMO
BACKGROUND: Necrotrophic effector proteins secreted by fungal pathogens are important virulence factors that mediate the development of disease in wheat. Pyrenophora tritici-repentis (Ptr), the causal agent of wheat tan spot, has a race structure dependent on the combination of effectors. In Ptr, ToxA and ToxB are known proteinaceous effectors responsible for necrosis and chlorosis respectively. While Ptr ToxA is encoded by the single gene ToxA, ToxB has multiple loci in the Ptr genome, which is postulated to be directly related to the level of ToxB production and leaf chlorosis. Although previous analysis has indicated that the majority of the ToxB loci lie on a single chromosome, the exact number and chromosomal locations for all the ToxB loci have not been fully identified. RESULTS: In this study, we have sequenced the genome of a race 5 ToxB-producing isolate (DW5), using PacBio long read technology, and found that ToxB duplications are nested in the complex subtelomeric chromosomal regions. A total of ten identical ToxB gene copies were identified and based on flanking sequence identity, nine loci appeared associated with chromosome 10 and a single copy with chromosome 5. Chromosome 10 multiple ToxB gene loci were separated by large sequence regions between 31 and 66 kb within larger segmental duplications in an alternating pattern related to loci strand, and flanked by transposable elements. CONCLUSION: This work provides for the first time the full accompaniment of ToxB loci and surrounding regions, and identifies the organization and distribution of ten ToxB loci to subtelomeric regions. To our knowledge, this is the first report of an interwoven strand-related duplication pattern event. This study further highlights the importance of resolving the highly complex distal chromosomal regions, that remain difficult to assemble, and can harbour important effectors and virulence factors.
Assuntos
Ascomicetos/genética , Proteínas Fúngicas/genética , Genoma Fúngico , Dosagem de GenesRESUMO
KEY MESSAGE: Genetic mapping of sensitivity to the Pyrenophora tritici-repentis effector ToxB allowed development of a diagnostic genetic marker, and investigation of wheat pedigrees allowed transmission of sensitive alleles to be tracked. Tan spot, caused by the necrotrophic fungal pathogen Pyrenophora tritici-repentis, is a major disease of wheat (Triticum aestivum). Secretion of the P. tritici-repentis effector ToxB is thought to play a part in mediating infection, causing chlorosis of plant tissue. Here, genetic analysis using an association mapping panel (n = 480) and a multiparent advanced generation intercross (MAGIC) population (n founders = 8, n progeny = 643) genotyped with a 90,000 feature single nucleotide polymorphism (SNP) array found ToxB sensitivity to be highly heritable (h2 ≥ 0.9), controlled predominantly by the Tsc2 locus on chromosome 2B. Genetic mapping of Tsc2 delineated a 1921-kb interval containing 104 genes in the reference genome of ToxB-insensitive variety 'Chinese Spring'. This allowed development of a co-dominant genetic marker for Tsc2 allelic state, diagnostic for ToxB sensitivity in the association mapping panel. Phenotypic and genotypic analysis in a panel of wheat varieties post-dated the association mapping panel further supported the diagnostic nature of the marker. Combining ToxB phenotype and genotypic data with wheat pedigree datasets allowed historic sources of ToxB sensitivity to be tracked, finding the variety 'Maris Dove' to likely be the historic source of sensitive Tsc2 alleles in the wheat germplasm surveyed. Exploration of the Tsc2 region gene space in the ToxB-sensitive line 'Synthetic W7984' identified candidate genes for future investigation. Additionally, a minor ToxB sensitivity QTL was identified on chromosome 2A. The resources presented here will be of immediate use for marker-assisted selection for ToxB insensitivity and the development of germplasm with additional genetic recombination within the Tsc2 region.
Assuntos
Ascomicetos , Resistência à Doença/genética , Interações Hospedeiro-Patógeno/genética , Micotoxinas/toxicidade , Doenças das Plantas/genética , Triticum/genética , Mapeamento Cromossômico , Ligação Genética , Marcadores Genéticos , Genômica , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Fenótipo , Doenças das Plantas/microbiologia , Polimorfismo de Nucleotídeo Único , Locos de Características QuantitativasRESUMO
The economically important necrotrophic fungal pathogen, Pyrenophora tritici-repentis (Ptr), causes tan spot of wheat, a disease typified by foliar necrosis and chlorosis. The culture filtrate of an Australian Ptr isolate, M4, possesses phytotoxic activity and plant bioassay guided discovery led to the purification of necrosis inducing toxins called triticone A and B. High-resolution LC-MS/MS analysis of the culture filtrate identified an additional 37 triticone-like compounds. The biosynthetic gene cluster responsible for triticone production (the Ttc cluster) was identified and deletion of TtcA, a hybrid polyketide synthase (PKS)-nonribosomal peptide synthase (NRPS), abolished production of all triticones. The pathogenicity of mutant (ttcA) strains was not visibly affected in our assays. We hypothesize that triticones possess general antimicrobial activity important for competition in multi-microbial environments.
Assuntos
Ascomicetos/enzimologia , Proteínas Fúngicas/metabolismo , Lactamas/metabolismo , Peptídeo Sintases/metabolismo , Doenças das Plantas/microbiologia , Policetídeo Sintases/metabolismo , Triticum/microbiologia , Ascomicetos/química , Ascomicetos/genética , Ascomicetos/metabolismo , Austrália , Cromatografia Líquida , Proteínas Fúngicas/genética , Deleção de Genes , Lactamas/química , Peptídeo Sintases/genética , Policetídeo Sintases/genética , Espectrometria de Massas em TandemRESUMO
BACKGROUND: Pyrenophora tritici-repentis (Ptr) is a necrotrophic fungal pathogen that causes the major wheat disease, tan spot. We set out to provide essential genomics-based resources in order to better understand the pathogenicity mechanisms of this important pathogen. RESULTS: Here, we present eight new Ptr isolate genomes, assembled and annotated; representing races 1, 2 and 5, and a new race. We report a high quality Ptr reference genome, sequenced by PacBio technology with Illumina paired-end data support and optical mapping. An estimated 98% of the genome coverage was mapped to 10 chromosomal groups, using a two-enzyme hybrid approach. The final reference genome was 40.9 Mb and contained a total of 13,797 annotated genes, supported by transcriptomic and proteogenomics data sets. CONCLUSIONS: Whole genome comparative analysis revealed major chromosomal segmental rearrangements and fusions, highlighting intraspecific genome plasticity in this species. Furthermore, the Ptr race classification was not supported at the whole genome level, as phylogenetic analysis did not cluster the ToxA producing isolates. This expansion of available Ptr genomics resources will directly facilitate research aimed at controlling tan spot disease.
Assuntos
Ascomicetos/genética , Ascomicetos/fisiologia , Cromossomos Fúngicos/genética , Variação Genética , Genoma Fúngico/genética , Genômica , Triticum/microbiologia , Transferência Genética Horizontal , Genoma Mitocondrial/genética , Anotação de Sequência Molecular , Filogenia , Homologia de Sequência do Ácido NucleicoRESUMO
Tan spot or yellow (leaf) spot disease of wheat (Triticum spp.) is caused by Pyrenophora tritici-repentis, a necrotrophic fungal pathogen that is widespread throughout the main wheat-growing regions in the world. This disease is currently the single most economically important crop disease in Australia. In this study, a real-time quantitative polymerase chain reaction (qPCR) assay was developed as a diagnostic tool to detect the pathogen on wheat foliar tissue. A multicopy locus (PtrMulti) present in the P. tritici-repentis genome was assessed for its suitability as a qPCR probe. The primer pair PtrMulti_F/R that targets the region was evaluated with respect to species specificity and sensitivity. A PtrMulti SYBR qPCR assay was developed and proved to be suitable for the identification and relative quantification of P. tritici-repentis with a detection limit of DNA levels at <0.1 pg. Variation of the PtrMulti copy number between the geographical representatives of P. tritici-repentis strains examined was minimal, with the range of 63 to 85 copies per genome. For naturally infected wheat field samples, the incidence of P. tritici-repentis DNA on leaves quantified by qPCR varied up to 1,000-fold difference in the concentration, with a higher incidence of DNA occurring on the lower canopy for most of the growth stages examined. At the early growth stages, qPCR assay was able to detect P. tritici-repentis DNA on the younger leaves in the absence of visible tan spot lesions. These results demonstrate the potential of PtrMulti probe to be used for early detection and rapid screening of tan spot disease on wheat plants.
RESUMO
Three of the most important fungal pathogens of cereals are Pyrenophora tritici-repentis, the cause of tan spot on wheat, and Pyrenophora teres f. teres and Pyrenophora teres f. maculata, the cause of spot form and net form of net blotch on barley, respectively. Orthologous intergenic regions were used to examine the genetic relationships and divergence times between these pathogens. Mean divergence times were calculated at 519 kya (±30) between P. teresf. teres and P. teresf. maculata, while P. tritici-repentis diverged from both Pyrenophora teresforms 8.04 Mya (±138 ky). Individual intergenic regions showed a consistent pattern of co-divergence of the P. teresforms from P. tritici-repentis, with the pattern supported by phylogenetic analysis of conserved genes. Differences in calculated divergence times between individual intergenic regions suggested that they are not entirely under neutral selection, a phenomenon shared with higher Eukaryotes. P. tritici-repentis regions varied in divergence time approximately 5-12 Mya from the P. teres lineage, compared to the separation of wheat and barley some 12 Mya, while the P. teresf. teres and P. teresf. maculata intergenic region divergences correspond to the middle Pleistocene. The data suggest there is no correlation between the divergence of these pathogens the domestication of wheat and barley, and show P. teresf. teres and P. teresf. maculata are closely related but autonomous. The results are discussed in the context of speciation and the evolution of intergenic regions.
Assuntos
Ascomicetos/genética , Evolução Molecular , Hordeum/microbiologia , Doenças das Plantas/microbiologia , Triticum/microbiologia , Ascomicetos/classificação , Sequência de Bases , DNA Fúngico/genética , DNA Intergênico/genética , Especiação Genética , Genoma Fúngico/genética , Sequenciamento de Nucleotídeos em Larga Escala , Dados de Sequência Molecular , Tipagem de Sequências Multilocus , Técnicas de Tipagem Micológica , Filogenia , Análise de Sequência de DNA , Fatores de TempoRESUMO
⢠Arabidopsis SENSITIVE TO FREEZING6 (SFR6) controls cold- and drought-inducible gene expression and freezing- and osmotic-stress tolerance. Its identification as a component of the MEDIATOR transcriptional co-activator complex led us to address its involvement in other transcriptional responses. ⢠Gene expression responses to Pseudomonas syringae, ultraviolet-C (UV-C) irradiation, salicylic acid (SA) and jasmonic acid (JA) were investigated in three sfr6 mutant alleles by quantitative real-time PCR and susceptibility to UV-C irradiation and Pseudomonas infection were assessed. ⢠sfr6 mutants were more susceptible to both Pseudomonas syringae infection and UV-C irradiation. They exhibited correspondingly weaker PR (pathogenesis-related) gene expression than wild-type Arabidopsis following these treatments or after direct application of SA, involved in response to both UV-C and Pseudomonas infection. Other genes, however, were induced normally in the mutants by these treatments. sfr6 mutants were severely defective in expression of plant defensin genes in response to JA; ectopic expression of defensin genes was provoked in wild-type but not sfr6 by overexpression of ERF5. ⢠SFR6/MED16 controls both SA- and JA-mediated defence gene expression and is necessary for tolerance of Pseudomonas syringae infection and UV-C irradiation. It is not, however, a universal regulator of stress gene transcription and is likely to mediate transcriptional activation of specific regulons only.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Ciclopentanos/metabolismo , Regulação da Expressão Gênica de Plantas , Oxilipinas/metabolismo , Ácido Salicílico/metabolismo , Transativadores/metabolismo , Arabidopsis/efeitos dos fármacos , Arabidopsis/metabolismo , Arabidopsis/microbiologia , Arabidopsis/efeitos da radiação , Proteínas de Arabidopsis/genética , Ciclopentanos/farmacologia , Complexo Mediador/genética , Complexo Mediador/metabolismo , Oxilipinas/farmacologia , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Pseudomonas syringae/patogenicidade , Ácido Salicílico/farmacologia , Transdução de Sinais , Transativadores/genética , Raios UltravioletaRESUMO
The adaptive potential of plant fungal pathogens is largely governed by the gene content of a species, consisting of core and accessory genes across the pathogen isolate repertoire. To approximate the complete gene repertoire of a globally significant crop fungal pathogen, a pan genomic analysis was undertaken for Pyrenophora tritici-repentis (Ptr), the causal agent of tan (or yellow) spot disease in wheat. In this study, 15 new Ptr genomes were sequenced, assembled and annotated, including isolates from three races not previously sequenced. Together with 11 previously published Ptr genomes, a pangenome for 26 Ptr isolates from Australia, Europe, North Africa and America, representing nearly all known races, revealed a conserved core-gene content of 57â% and presents a new Ptr resource for searching natural homologues (orthologues not acquired by horizontal transfer from another species) using remote protein structural homology. Here, we identify for the first time a non-synonymous mutation in the Ptr necrotrophic effector gene ToxB, multiple copies of the inactive toxb within an isolate, a distant natural Pyrenophora homologue of a known Parastagonopora nodorum necrotrophic effector (SnTox3), and clear genomic break points for the ToxA effector horizontal transfer region. This comprehensive genomic analysis of Ptr races includes nine isolates sequenced via long read technologies. Accordingly, these resources provide a more complete representation of the species, and serve as a resource to monitor variations potentially involved in pathogenicity.
Assuntos
Micotoxinas , Triticum , Ascomicetos , Interações Hospedeiro-Patógeno/genética , Micotoxinas/genética , Micotoxinas/metabolismo , Doenças das Plantas/microbiologia , Homologia Estrutural de Proteína , Triticum/genética , Triticum/metabolismo , Triticum/microbiologiaRESUMO
OBJECTIVES: The assembly of fungal genomes using short-reads is challenged by long repetitive and low GC regions. However, long-read sequencing technologies, such as PacBio and Oxford Nanopore, are able to overcome many problematic regions, thereby providing an opportunity to improve fragmented genome assemblies derived from short reads only. Here, a necrotrophic fungal pathogen Pyrenophora tritici-repentis (Ptr) isolate 134 (Ptr134), which causes tan spot disease on wheat, was sequenced on a MinION using Oxford Nanopore Technologies (ONT), to improve on a previous Illumina short-read genome assembly and provide a more complete genome resource for pan-genomic analyses of Ptr. RESULTS: The genome of Ptr134 sequenced on a MinION using ONT was assembled into 28 contiguous sequences with a total length of 40.79 Mb and GC content of 50.81%. The long-read assembly provided 6.79 Mb of new sequence and 2846 extra annotated protein coding genes as compared to the previous short-read assembly. This improved genome sequence represents near complete chromosomes, an important resource for large scale and pan genomic comparative analyses.