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The advancement of precision engineering for crop trait improvement is important in the face of rapid population growth, climate change, and disease. To this end, targeted double-stranded break technology using RNAguided Cas9 has been adopted widely for genome editing in plants. Agrobacterium or particle bombardment-based delivery of plasmids encoding Cas9 and guide RNA (gRNA) is common, but requires optimization of expression and often results in random integration of plasmid DNA into the plant genome. Recent advances have described gene editing by the delivery of Cas9 and gRNA as pre-assembled ribonucleoproteins (RNPs) into various plant tissues, but with moderate efficiency in resulting regenerated plants. In this report we describe significant improvements to Cas9-RNP mediated gene editing in wheat. We demonstrate that Cas9-RNP assays in protoplasts are a fast and effective tool for rational selection of optimal gRNAs for gene editing in regenerable immature embryos (IEs), and that high temperature treatment enhances gene editing rates in both tissue types. We also show that Cas9- mediated editing persists for at least 14 days in gold particle bombarded wheat IEs. The regenerated edited wheat plants in this work are recovered at high rates in the absence of exogenous DNA and selection. With this method, we produce knockouts of a set of three homoeologous genes and two pathogenic effector susceptibility genes, engineering insensitivity to corresponding necrotrophic effectors produced by Parastagonospora nodorum. The establishment of highly efficient, exogenous DNA-free gene editing technology holds promise for accelerated trait diversity production in an expansive array of crops. [ABSTRACT FROM AUTHOR]

Septoria nodorum blotch (SNB) is a foliar disease of wheat caused by the necrotrophic fungal pathogen Parastagonospora nodorum. Research over the last two decades has shown that the wheat-P. nodorum pathosystem mostly follows an inverse gene-for-gene model. The fungus produces necrotrophic effectors (NEs) that interact with specific host gene products encoded by dominant sensitivity (S) genes. When a compatible interaction occurs, a 'defense response' in the host leads to programmed cell death thereby provided dead/dying cells from which the pathogen, being a necrotroph, can acquire nutrients allowing it to grow and sporulate. To date, nine S gene-NE interactions have been characterized in this pathosystem. Five NE-encoding genes, SnTox1, SnTox3, SnToxA, SnTox5, and SnTox267, have been cloned along with three host S genes, Tsn1, Snn1, and Snn3-D1. Studies have shown that P. nodorum hijacks multiple and diverse host targets to cause disease. SNB resistance is often quantitative in nature because multiple compatible interactions usually occur concomitantly. NE gene expression plays a key role in disease severity, and the effect of each compatible interaction can vary depending on the other existing compatible interactions. Numerous SNB-resistance QTL have been identified in addition to the known S genes, and more research is needed to understand the nature of these resistance loci. Marker-assisted elimination of S genes through conventional breeding practices and disruption of S genes using gene editing techniques are both effective strategies for the development of SNB-resistant wheat cultivars, which will become necessary as the global demand for sustenance grows. [ABSTRACT FROM AUTHOR]

The stem rust (Puccinia graminis Pers.:Pers. f.sp. tritici Eriks. and Henn.) resistance gene Sr39, which confers resistance to TTKSK (Ug99), has been incorporated into the wheat (Triticum aestivum L.) genome from Aegilops speltoides in the form of a chromosome translocation but it has not been deployed into adapted cultivars. In this study, we characterized translocation lines carrying Sr39 in four different wheat backgrounds with fluorescent genomic in situ hybridization (GISH) and simple sequence repeat (SSR) markers. The results indicated that RL5711 and RL6082 had translocation chromosomes of comparable structure. The translocation chromosome in PI 600683 has lost Ae. speltoides chromatin in the telomeric end of the 2S long arm. Six translocation lines derived from the cross P1600683/3*HY438 had translocation chromosomes of corn parable structure to the one found in PI 600683. However, one line (P9714-AM03C51), showed a substantial reduction in Ae. speltoides chromatin in the short arm of the translocated chromosome. The study demonstrated that it is apparently feasible to shorten Ae. speltoides chromatin in some wheat-Ae, speltoides translocation lines. These results and the identification of diagnostic SSR markers will be useful in guiding chromosome manipulation efforts to further shorten the Ae. speltoides chromosome segments in these materials. Greenhouse inoculation of translocation lines with stem rust indicated that the Sr39 gene conditions resistance to at least seven stem rust races. doi: 10.2135/cropsci2009.10.0633

Abstract: The wheat tan spot fungus (Pyrenophora tritici-repentis) produces a well-characterized host-selective toxin (HST) known as Ptr ToxA, which induces necrosis in genotypes that harbor the Tsn1 gene on chromosome [...]

Tan spot, caused by the fungal pathogen Pyrenophora tritici-repentis (Died.) Drechs. causes severe yield losses in wheat (Triticum aestivum L, 2n = 6x = 42, AABBDD) and durum (T. turgidum L., 2n = 4x = 28, AABB). The Tsn1 gene acts dominantly to confer sensitivity to a host-selective proteinaceous toxin (Ptr ToxA) produced by the fungus. Our objectives were to: (i) target markers to the Tsn1 genomic region and (ii) develop a high-resolution map of the Tsn1 locus. The techniques of methylation-sensitive AFLP, traditional AFLP, and cDNA-AFLP were combined with bulked segregant analysis (BSA) using various wheat and durum cytogenetic stocks to target markers to the Tsn1 genomic region. Over 500 primer combinations were screened resulting in the identification of 18 low-copy markers closely linked to Tsn1. High-resolution mapping of the markers delineated the Tsn1 gene to a 0.2 cM interval in a hexaploid wheat population consisting of 1266 gametes, and to 0.8 cM in a durum wheat population consisting of 1860 gametes. Comparisons with rice BAC/PAC sequences indicated the lack of colinearity within the Tsn1 genomic region. Tsn1 was located within a gene-rich recombination hot spot region, and the physical distance separating the flanking markers may be as little as 200 kb. Therefore, these markers will serve as a basis for the map-based cloning of Tsn1. The isolation of Tsn1 will further our knowledge of wheat-tan spot interactions as well as host-pathogen interactions in general., SOME OF THE MOST FUNDAMENTAL QUESTIONS in plant biology concern the mechanisms underlying the phenomena of variable disease resistance among genotypes of a particular plant species. Much research has focused [...]

Summary: Parastagonospora nodorum is an economically important necrotrophic fungal pathogen of wheat. Parastagonospora nodorum secretes necrotrophic effectors that target wheat susceptibility genes to induce programmed cell death (PCD). In this study, we cloned and functionally validated SnTox5 and characterized its role in pathogenesis.We used whole genome sequencing, genome‐wide association study (GWAS) mapping, CRISPR‐Cas9‐based gene disruption, gain‐of‐function transformation, quantitative trait locus (QTL) analysis, haplotype and isoform analysis, protein modeling, quantitative PCR, and laser confocal microscopy to validate SnTox5 and functionally characterize SnTox5.SnTox5 is a mature 16.26 kDa protein with high structural similarity to SnTox3. Wild‐type and mutant P. nodorum strains and wheat genotypes of SnTox5 and Snn5, respectively, were used to show that SnTox5 not only targets Snn5 to induce PCD but also facilitates the colonization of the mesophyll layer even in the absence of Snn5.Here we show that SnTox5 facilitates the efficient colonization of the mesophyll tissue and elicits PCD specific to host lines carrying Snn5. The homology to SnTox3 and the ability of SnTox5 to facilitate the colonizing of the mesophyll also suggest a role in the suppression of host defense before PCD induction. See also the Commentary on this article by Kanyuka et al., 233: 11–14. [ABSTRACT FROM AUTHOR]

Summary: Parastagonospora nodorum is a fungal pathogen of wheat. As a necrotrophic specialist, it deploys effector proteins that target dominant host susceptibility genes to elicit programmed cell death (PCD). Here we identify and functionally validate the effector targeting the host susceptibility genes Snn2, Snn6 and Snn7.We utilized whole‐genome sequencing, association mapping, gene‐disrupted mutants, gain‐of‐function transformants, virulence assays, bioinformatics and quantitative PCR to characterize these interactions.A single proteinaceous effector, SnTox267, targeted Snn2, Snn6 and Snn7 to trigger PCD. Snn2 and Snn6 functioned cooperatively to trigger PCD in a light‐dependent pathway, whereas Snn7‐mediated PCD functioned in a light‐independent pathway. Isolates harboring 20 SnTox267 protein isoforms quantitatively varied in virulence. The diversity and distribution of isoforms varied between populations, indicating adaptation to local selection pressures. SnTox267 deletion resulted in the upregulation of effector genes SnToxA, SnTox1 and SnTox3.We validated a novel effector operating in an inverse‐gene‐for‐gene manner to target three genetically distinct host susceptibility genes and elicit PCD. The discovery of the complementary gene action of Snn2 and Snn6 indicates their potential function in a guard or decoy model. Additionally, differences in light dependency in the elicited pathways and upregulation of unlinked effectors sheds new light onto a complex fungal necrotroph–host interaction. See also the Commentary on this article by Kanyuka et al., 233: 11–14. [ABSTRACT FROM AUTHOR]

Disease lesion mimic mutants (DLMMs) are characterized by the spontaneous development of necrotic spots with various phenotypes designated as necrotic (nec) mutants in barley. The nec mutants were traditionally considered to have aberrant regulation of programmed cell death (PCD) pathways, which have roles in plant immunity and development. Most barley nec3 mutants express cream to orange necrotic lesions contrasting them from typical spontaneous DLMMs that develop dark pigmented lesions indicative of serotonin/phenolics deposition. Barley nec3 mutants grown under sterile conditions did not exhibit necrotic phenotypes until inoculated with adapted pathogens, suggesting that they are not typical DLMMs. The F2 progeny of a cross between nec3-γ1 and variety Quest segregated as a single recessive susceptibility gene post-inoculation with Bipolaris sorokiniana, the causal agent of the disease spot blotch. Nec3 was genetically delimited to 0.14 cM representing 16.5 megabases of physical sequence containing 149 annotated high confidence genes. RNAseq and comparative analysis of the wild type and five independent nec3 mutants identified a single candidate cytochrome P450 gene (HORVU.MOREX.r2.6HG0460850) that was validated as nec3 by independent mutations that result in predicted nonfunctional proteins. Histology studies determined that nec3 mutants had an unstable cutin layer that disrupted normal Bipolaris sorokiniana germ tube development. Author summary: At the site of pathogen infection, plant defense mechanisms rely on controlled programmed cell death (PCD) to sequester biotrophic pathogens that require living cells to extract nutrients from the host. However, these defense mechanisms are hijacked by necrotrophic plant pathogens that purposefully induce PCD to feed on the dead cells, thus facilitating further disease development. Thus, understanding PCD responses is important for resistance to both classes of pathogens. We characterized five independent disease lesion mimic mutants of barley designated necrotic 3 (nec3) that show aberrant regulation of PCD responses upon pathogen challenge. A cytochrome P450 gene was identified as Nec3 encoding a Tryptamine 5-Hydroxylase that functions as a terminal serotonin biosynthetic enzyme in the Tryptophan pathway of plants. We posit that nec3 mutants have disrupted serotonin biosynthesis resulting in expanded PCD, necrotrophic pathogen susceptibility and cutin layer instability. The nec3 mutants show expanded PCD and disease susceptibility of pathogen-induced necrotic lesions, suggesting a role of serotonin to sequester PCD and suppress pathogen colonization. The identification of Nec3 will facilitate functional analysis to elucidate the role that serotonin plays in the elicitation or suppression of PCD immunity responses to diverse pathogens and the effects it has on cutin layer biosynthesis. [ABSTRACT FROM AUTHOR]

The rapid and widespread evolution of fungicide resistance remains a challenge for crop disease management. The demethylation inhibitor (DMI) class of fungicides is a widely used chemistry for managing disease, but there has been a gradual decline in efficacy in many crop pathosystems. Reliance on DMI fungicides has increased resistance in populations of the plant pathogenic fungus Cercospora beticola worldwide. To better understand the genetic and evolutionary basis for DMI resistance in C. beticola , a genome-wide association study (GWAS) and selective sweep analysis were conducted for the first time in this species. We performed whole-genome resequencing of 190 C. beticola isolates infecting sugar beet (Beta vulgaris ssp. vulgaris). All isolates were phenotyped for sensitivity to the DMI tetraconazole. Intragenic markers on chromosomes 1, 4, and 9 were significantly associated with DMI fungicide resistance, including a polyketide synthase gene and the gene encoding the DMI target CbCYP51. Haplotype analysis of CbCYP51 identified a synonymous mutation (E170) and nonsynonymous mutations (L144F, I387M, and Y464S) associated with DMI resistance. Genome-wide scans of selection showed that several of the GWAS mutations for fungicide resistance resided in regions that have recently undergone a selective sweep. Using radial plate growth on selected media as a fitness proxy, we did not find a trade-off associated with DMI fungicide resistance. Taken together, we show that population genomic data from a crop pathogen can allow the identification of mutations conferring fungicide resistance and inform about their origins in the pathogen population. [ABSTRACT FROM AUTHOR]

Great strides have been made in defining the details of the plant defense response involving biotrophic fungal and bacterial pathogens. The groundwork for the current model was laid by H.H. Flor and others who defined the gene-for-gene hypothesis, which is now known to involve effector-triggered immunity (ETI). PAMP-triggered immunity (PTI) is also a highly effective response to most pathogens because of the recognition of common pathogen molecules by pattern recognition receptors. In this article, we consider the three pathogens that make up the foliar disease complex of wheat, Zymoseptoria tritici, Pyrenophora tritici-repentis, and Parastagonospora nodorum, to review the means by which necrotrophic pathogens circumvent, or outright hijack, the ETI and PTI pathways to cause disease. [ABSTRACT FROM AUTHOR]

SUMMARY: The resistance gene Sr13 is one of the most important genes in durum wheat for controlling stem rust caused by Puccinia graminis f. sp. tritici (Pgt). The Sr13 functional gene CNL13 has haplotypes R1, R2 and R3. The R1/R3 and R2 haplotypes were originally designated as alleles Sr13a and Sr13b, respectively. To detect additional Sr13 alleles, we developed Kompetitive allele specific PCR (KASP™) marker KASPSr13 and four semi‐thermal asymmetric reverse PCR markers, rwgsnp37–rwgsnp40, based on the CNL13 sequence. These markers were shown to detect R1, R2 and R3 haplotypes in a panel of diverse tetraploid wheat accessions. We also observed the presence of Sr13 in durum line CAT‐A1, although it lacked any of the known haplotypes. Sequence analysis revealed that CNL13 of CAT‐A1 differed from the susceptible haplotype S1 by a single nucleotide (C2200T) in the leucine‐rich repeat region and differed from the other three R haplotypes by one or two additional nucleotides, confirming that CAT‐A1 carries a new (R4) haplotype. Stem rust tests on the monogenic, transgenic and mutant lines showed that R1 differed from R3 in its susceptibility to races TCMJC and THTSC, whereas R4 differed from all other haplotypes for susceptibility to TTKSK, TPPKC and TCCJC. Based on these differences, we designate the R1, R3 and R4 haplotypes as alleles Sr13a, Sr13c and Sr13d, respectively. This study indicates that Sr13d may be the primitive functional allele originating from the S1 haplotype via a point mutation, with the other three R alleles probably being derived from Sr13d through one or two additional point mutations. Significance Statement: A new resistant haplotype (R4) of Sr13, designated as Sr13d, was identified in the genotype CAT‐A1, and based on sequencing studies this allele is hypothesized to be a primitive functional resistant allele of Sr13. We established from extensive rust tests that haplotypes R1 and R3 should be separated, and R3 was designated Sr13c and determined to be preferable to Sr13a for breeding, while Sr13d was undesirable for breeding due to susceptibility to stem rust Ug99. [ABSTRACT FROM AUTHOR]

SUMMARY: Septoria nodorum blotch (SNB), a disease caused by the necrotrophic fungal pathogen Parastagonospora nodorum, is a threat to wheat (Triticum aestivum) production worldwide. Multiple inverse gene‐for‐gene interactions involving the recognition of necrotrophic effectors (NEs) by wheat sensitivity genes play major roles in causing SNB. One interaction involves the wheat gene Snn3 and the P. nodorum NE SnTox3. Here, we used a map‐based strategy to clone the Snn3‐D1 gene from Aegilops tauschii, the D‐genome progenitor of common wheat. Snn3‐D1 contained protein kinase and major sperm protein domains, both of which were essential for function as confirmed by mutagenesis. As opposed to other characterized interactions in this pathosystem, a compatible Snn3‐D1–SnTox3 interaction was light‐independent, and Snn3‐D1 transcriptional expression was downregulated by light and upregulated by darkness. Snn3‐D1 likely emerged in Ae. tauschii due to an approximately 218‐kb insertion that occurred along the west bank of the Caspian Sea. The identification of this new class of NE sensitivity genes combined with the previously cloned sensitivity genes demonstrates that P. nodorum can take advantage of diverse host targets to trigger SNB susceptibility in wheat. Significance Statement: Necrotrophic effectors produced by the necrotrophic fungal pathogen Parastagonospora nodorum can be recognized by wheat (Triticum aestivum) sensitivity genes to cause septoria nodorum blotch (SNB), a fungal disease posing a threat to wheat production worldwide. Here, we cloned a new type of wheat sensitivity gene containing protein kinase and major sperm protein domains, which furthered our understanding of how P. nodorum takes advantage of diverse host targets to trigger SNB susceptibility in wheat. [ABSTRACT FROM AUTHOR]

Cercospora beticola is a hemibiotrophic fungus that causes cercospora leaf spot disease of sugar beet (Beta vulgaris). After an initial symptomless biotrophic phase of colonization, necrotic lesions appear on host leaves as the fungus switches to a necrotrophic lifestyle. The phytotoxic secondary metabolite cercosporin has been shown to facilitate fungal virulence for several Cercospora spp. However, because cercosporin production and subsequent cercosporin‐initiated formation of reactive oxygen species is light‐dependent, cell death evocation by this toxin is only fully ensured during a period of light. Here, we report the discovery of the effector protein CbNip1 secreted by C. beticola that causes enhanced necrosis in the absence of light and, therefore, may complement light‐dependent necrosis formation by cercosporin. Infiltration of CbNip1 protein into sugar beet leaves revealed that darkness is essential for full CbNip1‐triggered necrosis, as light exposure delayed CbNip1‐triggered host cell death. Gene expression analysis during host infection shows that CbNip1 expression is correlated with symptom development in planta. Targeted gene replacement of CbNip1 leads to a significant reduction in virulence, indicating the importance of CbNip1 during colonization. Analysis of 89 C. beticola genomes revealed that CbNip1 resides in a region that recently underwent a selective sweep, suggesting selection pressure exists to maintain a beneficial variant of the gene. Taken together, CbNip1 is a crucial effector during the C. beticola–sugar beet disease process. [ABSTRACT FROM AUTHOR]

Barley is an important cereal crop worldwide because of its use in the brewing and distilling industry. However, adequate supplies of quality malting barley are threatened by global climate change due to drought in some regions and excess precipitation in others, which facilitates epidemics caused by fungal pathogens. The disease net form net blotch caused by the necrotrophic fungal pathogen Pyrenophora teres f. teres (Ptt) has emerged as a global threat to barley production and diverse populations of Ptt have shown a capacity to overcome deployed genetic resistances. The barley line CI5791 exhibits remarkably effective resistance to diverse Ptt isolates from around the world that maps to two major QTL on chromosomes 3H and 6H. To identify genes involved in this effective resistance, CI5791 seed were γ-irradiated and two mutants, designated CI5791-γ3 and CI5791-γ8, with compromised Ptt resistance were identified from an M2 population. Phenotyping of CI5791-γ3 and -γ8 × Heartland F2 populations showed three resistant to one susceptible segregation ratios and CI5791-γ3 × -γ8 F1 individuals were susceptible, thus these independent mutants are in a single allelic gene. Thirty-four homozygous mutant (susceptible) CI5791-γ3 × Heartland F2 individuals, representing 68 recombinant gametes, were genotyped via PCR genotype by sequencing. The data were used for single marker regression mapping placing the mutation on chromosome 3H within an approximate 75 cM interval encompassing the 3H CI5791 resistance QTL. Sequencing of the mutants and wild-type (WT) CI5791 genomic DNA following exome capture identified independent mutations of the HvWRKY6 transcription factor located on chromosome 3H at ∼50.7 cM, within the genetically delimited region. Post transcriptional gene silencing of HvWRKY6 in barley line CI5791 resulted in Ptt susceptibility, confirming that it functions in NFNB resistance, validating it as the gene underlying the mutant phenotypes. Allele analysis and transcript regulation of HvWRKY6 from resistant and susceptible lines revealed sequence identity and upregulation upon pathogen challenge in all genotypes analyzed, suggesting a conserved transcription factor is involved in the defense against the necrotrophic pathogen. We hypothesize that HvWRKY6 functions as a conserved signaling component of defense mechanisms that restricts Ptt growth in barley. [ABSTRACT FROM AUTHOR]

Plant fungal pathogens can be classified according to their lifestyles. Biotrophs feed on living tissue and constitute an economically significant group of pathogens historically. Necrotrophs, which feed on dead tissue, have become economically significant over recent decades, especially those of the Dothideomycetes , which produce necrotrophic effectors (NEs) to modulate the host response. Some of these pathogens interact with their hosts in an inverse gene-for-gene manner, where NEs are recognized by specific dominant genes in the host leading to host-mediated programmed cell death allowing the pathogen to cause disease. Whereas the NE genes tend to be unique, several of the plant 'susceptibility' genes belong to the nucleotide-binding leucine-rich repeat class of disease 'resistance' genes, and one is a wall-associated kinase. These susceptible interactions exhibit hallmarks of defense responses to biotrophic pathogens. Therefore, there is now accumulating evidence that many necrotrophic specialists hijack the resistance mechanisms that are effective against biotrophic pathogens. [ABSTRACT FROM AUTHOR]

Summary: Pyrenophora teres f. teres and P. teres f. maculata are significant pathogens that cause net blotch of barley. An increased number of loci involved in P. teres resistance or susceptibility responses of barley as well as interacting P. teres virulence effector loci have recently been identified through biparental and association mapping studies of both the pathogen and host. Characterization of the resistance/susceptibility loci in the host and the interacting effector loci in the pathogen will provide a path for targeted gene validation for better‐informed release of resistant barley cultivars. This review assembles concise consensus maps for all loci published for both the host and pathogen, providing a useful resource for the community to be used in pathogen characterization and barley breeding for resistance to both forms of P. teres. [ABSTRACT FROM AUTHOR]

Filamentous fungi rapidly evolve in response to environmental selection pressures in part due to their genomic plasticity. Parastagonospora nodorum, a fungal pathogen of wheat and causal agent of septoria nodorum blotch, responds to selection pressure exerted by its host, influencing the gain, loss, or functional diversification of virulence determinants, known as effector genes. Whole genome resequencing of 197 P. nodorum isolates collected from spring, durum, and winter wheat production regions of the United States enabled the examination of effector diversity and genomic regions under selection specific to geographically discrete populations. 1,026,859 SNPs/InDels were used to identify novel loci, as well as SnToxA and SnTox3 as factors in disease. Genes displaying presence/absence variation, predicted effector genes, and genes localized on an accessory chromosome had significantly higher pN/pS ratios, indicating a higher rate of sequence evolution. Population structure analyses indicated two P. nodorum populations corresponding to the Upper Midwest (Population 1) and Southern/Eastern United States (Population 2). Prevalence of SnToxA varied greatly between the two populations which correlated with presence of the host sensitivity gene Tsn1 in the most prevalent cultivars in the corresponding regions. Additionally, 12 and 5 candidate effector genes were observed to be under diversifying selection among isolates from Population 1 and 2, respectively, but under purifying selection or neutrally evolving in the opposite population. Selective sweep analysis revealed 10 and 19 regions that had recently undergone positive selection in Population 1 and 2, respectively, involving 92 genes in total. When comparing genes with and without presence/absence variation, those genes exhibiting this variation were significantly closer to transposable elements. Taken together, these results indicate that P. nodorum is rapidly adapting to distinct selection pressures unique to spring and winter wheat production regions by rapid adaptive evolution and various routes of genomic diversification, potentially facilitated through transposable element activity. Parastagonospora nodorum is an economically important pathogen of wheat, employing proteinaceous effectors to cause disease. Recognition of effectors by host susceptibility genes often leads to the elicitation of programmed cell death. However, little is known on the correlation between effector diversity and the spatial distribution of host resistance/susceptibility or the genomic mechanisms of diversification. This research presents the genome resequencing of 197 P. nodorum isolates collected from spring, winter, and durum wheat production regions of the United States, enabling the investigation of genome dynamics and evolution. Results illustrate local adaptation to host resistance or susceptibility, as evidenced by population-specific evolution of predicted effector genes and positively selected selective sweeps. Predicted effector genes, genes exhibiting presence/absence variation, and genes residing on an accessory chromosome, were found to be diversifying more rapidly. Additionally, transposable elements were predicted to play a role in the maintenance or elimination of genes. A GWAS approach identified the previously reported SnToxA and SnTox3 as well as novel virulence candidates as major elicitors of disease. These results highlight the flexibility of the P. nodorum genome in response to population-specific selection pressures and illustrate the utility of whole genome resequencing for the identification of putative virulence mechanisms. [ABSTRACT FROM AUTHOR]

Synthetic hexaploid wheat (SHW) can serve as a bridge for the transfer of useful genes from Aegilops tauschii and tetraploid wheat (Triticum turgidum) into common wheat (T. aestivum). The objective of this study was to evaluate 149 SHW lines and their 74 tetraploid parents for their genetic diversity, breeding values and inter-genomic interactions for resistance to Fusarium head blight (FHB). The genetic diversity analysis was performed based on the population structure established using 4,674 and 3,330 polymorphic SNP markers among the SHW lines and tetraploid parents, respectively. The results showed that all T. carthlicum and most T. dicoccum accessions formed different clusters and subpopulations, respectively, whereas all the T. durum , T. polonicum , T. turgidum , and T. turanicum accessions were clustered together, suggesting that T. durum was more closely related to T. polonicum , T. turgidum , and T. turanicum than to T. dicoccum. The genetic diversity of the SHW lines mainly reflected that of the tetraploid parents. The SHW lines and their tetraploid parents were evaluated for reactions to FHB in two greenhouse seasons and at two field nurseries for 2 years. As expected, most of the SHW lines were more resistant than their tetraploid parents in all environments. The FHB severities of the SHW lines varied greatly depending on the Ae. tauschii and tetraploid genotypes involved. Most of the SHW lines with a high level of FHB resistance were generally derived from the tetraploid accessions with a high level of FHB resistance. Among the 149 SHW lines, 140 were developed by using three Ae. tauschii accessions CIae 26, PI 268210, and RL 5286. These SHW lines showed FHB severities reduced by 21.7%, 17.3%, and 11.5%, respectively, with an average reduction of 18.3%, as compared to the tetraploid parents, suggesting that the D genome may play a major role in reducing disease severity in the SHW lines. Thirteen SHW lines consistently showed a high level of FHB resistance compared to the resistant check, Sumai 3, in each environment. These SHW lines will be useful for the development of FHB-resistant wheat germplasm and populations for discovery of novel FHB resistance genes. [ABSTRACT FROM AUTHOR]

We report a fungal pan-genome study involving Parastagonospora spp. including 21 isolates of the wheat (Triticum aestivum) pathogen P arastagonospora nodorum, 10 of the grass-infecting Parastagonospora avenae, and 2 of a closely related undefined sister species. We observed substantial variation in the distribution of polymorphisms across the pan-genome, including repeat-induced point mutations, diversifying selection and gene gains and losses. We also discovered chromosome-scale inter and intraspecific presence/absence variation of some sequences, suggesting the occurrence of one or more accessory chromosomes or regions that may play a role in host–pathogen interactions. The presence of known pathogenicity effector loci SnToxA, SnTox1, and SnTox3 varied substantially among isolates. Three P. nodorum isolates lacked functional versions for all three loci, whereas three P. avenae isolates carried one or both of the SnTox1 and SnTox3 genes, indicating previously unrecognized potential for discovering additional effectors in the P. nodorum -wheat pathosystem. We utilized the pan-genomic comparative analysis to improve the prediction of pathogenicity effector candidates, recovering the three confirmed effectors among our top-ranked candidates. We propose applying this pan-genomic approach to identify the effector repertoire involved in other host–microbe interactions involving necrotrophic pathogens in the Pezizomycotina. [ABSTRACT FROM AUTHOR]