A unique resistance mechanism is associated with RBgh2 barley powdery mildew adult plant resistance.

KEY MESSAGE: Gene expression at the RBgh2 locus indicates involvement in cAMP/G-protein-coupled signalling and innate immunity in barley powdery mildew adult plant resistance. Barley powdery mildew is a globally significant disease, responsible for reduced grain yield and quality. A major effect adult plant resistance gene, RBgh2, was previously found in a landrace from Azerbaijan. The atypical phenotype suggested different underlying genetic factors compared to conventional resistance genes and to investigate this, genome-wide gene expression was compared between sets of heterogeneous doubled haploids. RBgh2 resistance is recessive and induces both temporary genome-wide gene expression changes during powdery mildew infection together with constitutive changes, principally at the RBgh2 locus. Defence-related genes significantly induced included homologues of genes associated with innate immunity and pathogen recognition. Intriguingly, RBgh2 resistance does not appear to be dependent on salicylic acid signalling, a key pathway in plant resistance to biotrophs. Constitutive co-expression of resistance gene homologues was evident at the 7HS RBgh2 locus, while no expression was evident for a 6-transmembrane gene, predicted in silico to contain both G-protein- and calmodulin-binding domains. The gene was disrupted at the 5' end, and G-protein-binding activity was suppressed. RBgh2 appears to operate through a unique mechanism that co-opts elements of innate immunity.

Major Susceptibility Gene Epistasis over Minor Gene Resistance to Spot Form Net Blotch in a Commercial Barley Cultivar.

Spot form net blotch, caused by Pyrenophora teres f. maculata, is a significant global disease of barley (Hordeum vulgare). Baudin, a barley cultivar that was until recently extensively grown in Western Australia, was reported as having minor seedling resistance. However, Baudin was highly susceptible to a local isolate, M3, suggesting that this isolate had gained virulence against a major susceptibility gene. M3 causes atypical lesions with pale centers early in the infection, with initial screens of a segregating population indicating that this was determined by a single locus in the Baudin genome. The susceptibility was semidominant in F1 progeny and the susceptibility gene, designated Spm1 (Susceptibility to P. teres f. maculata 1), mapped to a 190-kb section of the resistance gene-rich Mla region of chromosome 1H. Phenotyping with Ptm SP1, a non-M3 pathotype, identified a seedling resistance locus on 2H. Minor gene resistance is generally regarded as potentially durable, but our findings suggest the resistance to spot form net blotch in Baudin is nullified by strong susceptibility conferred by a separate locus on 1H. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Potentiators of Disease During Barley Infection by Pyrenophora teres f. teres in a Susceptible Interaction.

Pyrenophora teres f. teres is a necrotrophic fungal pathogen and causal agent of net form net blotch (NFNB), a significant disease of barley. RNA-seq data encompassing asymptomatic and subsequent necrotrophic phases of the pathogen was obtained for P. teres f. teres isolate W1-1 in NFNB-sensitive cultivar Baudin. Host genes notably regulated during infection included concerted induction of over half the repertoire of disease resistance genes, together with genes involved in oxidation-reduction processes, characteristic of a hypersensitive response. Several systemic acquired resistance response genes were suppressed and there was a complete absence of defense-related thionin gene expression. In P. teres f. teres, genes involved in hydrolase activities and cell-wall catabolic processes were induced during infection, while nitrate assimilation and response to oxidative stress processes were suppressed. Timecourse data allowed a number of predicted P. teres f. teres effector genes with differing expression profiles to be identified that may underlie barley sensitivity to NFNB. Candidate genes involved in the host-pathogen interaction provide a basis for functional characterization and control strategies based on fungicide or mutation targets, which will facilitate further research aimed at controlling NFNB disease.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Parallel evolution of multiple mechanisms for demethylase inhibitor fungicide resistance in the barley pathogen Pyrenophora teres f. sp. maculata.

The fungal pathogen Pyrenophora teres f. sp. maculata (Ptm), responsible for spot-form of net blotch (SFNB), is currently the most significant disease of barley in Australia and a major disease worldwide. Management of SFNB relies heavily on fungicides and in Australia the demethylase inhibitors (DMIs) predominate. There have been sporadic reports of resistance to DMIs in Ptm but the mechanisms remain obscure. Ptm isolates collected from 1996 to 2019 in Western Australia were tested for fungicide sensitivity levels. Decreased sensitivity to DMIs was observed in isolates collected after 2015. Resistance factors to tebuconazole fell into two classes; moderate resistance (MR; RF 6-11) and high resistance (HR; RFs 30-65). Mutations linked to resistance were detected in the promoter region and coding sequence of the DMI target gene Cyp51A. Solo-LTR insertion elements were found at 5 different locations in the promoter region. Three different non-synonymous mutations encoded an altered protein with a phenylalanine to leucine substitution at position 489, F489L (F495I in the archetype CYP51A of Aspergillus fumigatus). F489L mutations have also been found in DMI-resistant strains of P. teres f. sp. teres. Ptm isolates carrying either a LTR insertion element or a F489L allele displayed the MR1 or MR2 phenotypes, respectively. Isolates carrying both an insertion element and a F489L mutation displayed the HR phenotype. Multiple mechanisms acting both alone and in concert were found to contribute to DMI resistance in Ptm. Moreover, these mutations have emerged repeatedly in Western Australian Ptm populations by a process of parallel evolution.

Genomic Regions Associated with Virulence in Pyrenophora teres f. teres Identified by Genome-Wide Association Analysis and Biparental Mapping.

Net form net blotch (NFNB), caused by the fungal pathogen Pyrenophora teres f. teres, is an important foliar disease present in all barley-producing regions of the world. This fungus is a hemibiotrophic and heterothallic ascomycete, where sexual recombination can lead to changes in disease expression in the host. Knowledge of the genetic architecture and genes involved in virulence is vital to increase the durability of NFNB resistance in barley cultivars. We used a genome-wide association mapping approach to characterize P. teres f. teres genomic regions associated with virulence in Australian barley cultivars. One hundred eighty-eight P. teres f. teres isolates collected across five Australian states were genotyped using Diversity Arrays Technology sequence markers and phenotyped across 20 different barley genotypes. Association mapping identified 14 different genomic regions associated with virulence, with the majority located on P. teres f. teres chromosomes 3 and 5 and one each present on chromosomes 1, 6, and 9. Four of the regions identified were confirmed by quantitative trait loci (QTL) mapping. The QTL regions are discussed in the context of their genomic architecture together with examination of their gene contents, which identified 20 predicted effectors. The number of QTL shown in this study at the population level clearly illustrates a complex genetic basis of P. teres f. teres virulence compared with pure necrotrophs, such as the wheat pathogens Parastagonospora nodorum and Parastagonospora tritici-repentis.

Physiological Changes in Barley mlo-11 Powdery Mildew Resistance Conditioned by Tandem Repeat Copy Number.

Wild barley accessions have evolved broad-spectrum defence against barley powdery mildew through recessive mlo mutations. However, the mlo defence response is associated with deleterious phenotypes with a cost to yield and fertility, with implications for natural fitness and agricultural productivity. This research elucidates the mechanism behind a novel mlo allele, designated mlo-11(cnv2), which has a milder phenotype compared to standard mlo-11. Bisulphite sequencing and histone ChIP-seq analyses using near-isogenic lines showed pronounced repression of the Mlo promoter in standard mlo-11 compared to mlo-11(cnv2), with repression governed by 24 nt heterochromatic small interfering RNAs. The mlo-11(cnv2) allele appears to largely reduce the physiological effects of mlo while still endorsing a high level of powdery mildew resistance. RNA sequencing showed that this is achieved through only partly restricted expression of Mlo, allowing adequate temporal induction of defence genes during infection and expression close to wild-type Mlo levels in the absence of infection. The two mlo-11 alleles showed copy number proportionate oxidase and peroxidase expression levels during infection, but lower amino acid and aromatic compound biosynthesis compared to the null allele mlo-5. Examination of highly expressed genes revealed a common WRKY W-box binding motif (consensus ACCCGGGACTAAAGG) and a transcription factor more highly expressed in mlo-11 resistance. In conclusion, mlo-11(cnv2) appears to significantly mitigate the trade-off between mlo defence and normal gene expression.

Allelic barley MLA immune receptors recognize sequence-unrelated avirulence effectors of the powdery mildew pathogen.

Disease-resistance genes encoding intracellular nucleotide-binding domain and leucine-rich repeat proteins (NLRs) are key components of the plant innate immune system and typically detect the presence of isolate-specific avirulence (AVR) effectors from pathogens. NLR genes define the fastest-evolving gene family of flowering plants and are often arranged in gene clusters containing multiple paralogs, contributing to copy number and allele-specific NLR variation within a host species. Barley mildew resistance locus a (Mla) has been subject to extensive functional diversification, resulting in allelic resistance specificities each recognizing a cognate, but largely unidentified, AVRa gene of the powdery mildew fungus, Blumeria graminis f. sp. hordei (Bgh). We applied a transcriptome-wide association study among 17 Bgh isolates containing different AVRa genes and identified AVRa1 and AVRa13, encoding candidate-secreted effectors recognized by Mla1 and Mla13 alleles, respectively. Transient expression of the effector genes in barley leaves or protoplasts was sufficient to trigger Mla1 or Mla13 allele-specific cell death, a hallmark of NLR receptor-mediated immunity. AVRa1 and AVRa13 are phylogenetically unrelated, demonstrating that certain allelic MLA receptors evolved to recognize sequence-unrelated effectors. They are ancient effectors because corresponding loci are present in wheat powdery mildew. AVRA1 recognition by barley MLA1 is retained in transgenic Arabidopsis, indicating that AVRA1 directly binds MLA1 or that its recognition involves an evolutionarily conserved host target of AVRA1 Furthermore, analysis of transcriptome-wide sequence variation among the Bgh isolates provides evidence for Bgh population structure that is partially linked to geographic isolation.

Rare Pyrenophora teres Hybridization Events Revealed by Development of Sequence-Specific PCR Markers.

Pyrenophora teres f. teres and P. teres f. maculata cause net form and spot form, respectively, of net blotch on barley (Hordeum vulgare). The two forms reproduce sexually, producing hybrids with genetic and pathogenic variability. Phenotypic identification of hybrids is challenging because lesions induced by hybrids on host plants resemble lesions induced by either P. teres f. teres or P. teres f. maculata. In this study, 12 sequence-specific polymerase chain reaction markers were developed based on expressed regions spread across the genome. The primers were validated using 210 P. teres isolates, 2 putative field hybrids (WAC10721 and SNB172), 50 laboratory-produced hybrids, and 7 isolates collected from barley grass (H. leporinum). The sequence-specific markers confirmed isolate WAC10721 as a hybrid. Only four P. teres f. teres markers amplified on DNA of barley grass isolates. Amplified fragment length polymorphism markers suggested that P. teres barley grass isolates are genetically different from P. teres barley isolates and that the second putative hybrid (SNB172) is a barley grass isolate. We developed a suite of markers which clearly distinguish the two forms of P. teres and enable unambiguous identification of hybrids.

CodingQuarry: highly accurate hidden Markov model gene prediction in fungal genomes using RNA-seq transcripts.

BACKGROUND: The impact of gene annotation quality on functional and comparative genomics makes gene prediction an important process, particularly in non-model species, including many fungi. Sets of homologous protein sequences are rarely complete with respect to the fungal species of interest and are often small or unreliable, especially when closely related species have not been sequenced or annotated in detail. In these cases, protein homology-based evidence fails to correctly annotate many genes, or significantly improve ab initio predictions. Generalised hidden Markov models (GHMM) have proven to be invaluable tools in gene annotation and, recently, RNA-seq has emerged as a cost-effective means to significantly improve the quality of automated gene annotation. As these methods do not require sets of homologous proteins, improving gene prediction from these resources is of benefit to fungal researchers. While many pipelines now incorporate RNA-seq data in training GHMMs, there has been relatively little investigation into additionally combining RNA-seq data at the point of prediction, and room for improvement in this area motivates this study. RESULTS: CodingQuarry is a highly accurate, self-training GHMM fungal gene predictor designed to work with assembled, aligned RNA-seq transcripts. RNA-seq data informs annotations both during gene-model training and in prediction. Our approach capitalises on the high quality of fungal transcript assemblies by incorporating predictions made directly from transcript sequences. Correct predictions are made despite transcript assembly problems, including those caused by overlap between the transcripts of adjacent gene loci. Stringent benchmarking against high-confidence annotation subsets showed CodingQuarry predicted 91.3% of Schizosaccharomyces pombe genes and 90.4% of Saccharomyces cerevisiae genes perfectly. These results are 4-5% better than those of AUGUSTUS, the next best performing RNA-seq driven gene predictor tested. Comparisons against whole genome Sc. pombe and S. cerevisiae annotations further substantiate a 4-5% improvement in the number of correctly predicted genes. CONCLUSIONS: We demonstrate the success of a novel method of incorporating RNA-seq data into GHMM fungal gene prediction. This shows that a high quality annotation can be achieved without relying on protein homology or a training set of genes. CodingQuarry is freely available ( https://sourceforge.net/projects/codingquarry/ ), and suitable for incorporation into genome annotation pipelines.

Construction of integrated linkage map of a recombinant inbred line population of white lupin (Lupinus albus L.).

We report the development of a Diversity Arrays Technology (DArT) marker panel and its utilisation in the development of an integrated genetic linkage map of white lupin (Lupinus albus L.) using an F8 recombinant inbred line population derived from Kiev Mutant/P27174. One hundred and thirty-six DArT markers were merged into the first genetic linkage map composed of 220 amplified fragment length polymorphisms (AFLPs) and 105 genic markers. The integrated map consists of 38 linkage groups of 441 markers and spans a total length of 2,169 cM, with an average interval size of 4.6 cM. The DArT markers exhibited good genome coverage and were associated with previously identified genic and AFLP markers linked with quantitative trait loci for anthracnose resistance, flowering time and alkaloid content. The improved genetic linkage map of white lupin will aid in the identification of markers for traits of interest and future syntenic studies.

Widespread genetic heterogeneity and genotypic grouping associated with fungicide resistance among barley spot form net blotch isolates in Australia.

Spot form net blotch, caused by Pyrenophora teres f. maculata, is a major foliar disease of barley worldwide. Knowledge of the pathogen's genetic diversity and population structure is critical for a better understanding of inherent evolutionary capacity and for the development of sustainable disease management strategies. Genome-wide, single nucleotide polymorphism data of 254 Australian isolates revealed genotypic diversity and an absence of population structure, either between states, or between fields and cultivars in different agro-ecological zones. This indicates there is little geographical isolation or cultivar directional selection and that the pathogen is highly mobile across the continent. However, two cryptic genotypic groups were found only in Western Australia, predominantly associated with genes involved in fungicide resistance. The findings in this study are discussed in the context of current cultivar resistance and the pathogen's adaptive potential.

Evolution of three Pyrenophora cereal pathogens: recent divergence, speciation and evolution of non-coding DNA.

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.

Global genomic analyses of wheat powdery mildew reveal association of pathogen spread with historical human migration and trade.

The fungus Blumeria graminis f. sp. tritici causes wheat powdery mildew disease. Here, we study its spread and evolution by analyzing a global sample of 172 mildew genomes. Our analyses show that B.g. tritici emerged in the Fertile Crescent during wheat domestication. After it spread throughout Eurasia, colonization brought it to America, where it hybridized with unknown grass mildew species. Recent trade brought USA strains to Japan, and European strains to China. In both places, they hybridized with local ancestral strains. Thus, although mildew spreads by wind regionally, our results indicate that humans drove its global spread throughout history and that mildew rapidly evolved through hybridization.

Adult resistance genes to barley powdery mildew confer basal penetration resistance associated with broad-spectrum resistance.

Powdery mildew isa major disease of barley (Hordeum vulgare L.) for which breeders have traditionally relied on dominant, pathogen race-specific resistance genes for genetic control. Directional selection pressures in extensive monocultures invariably result in such genes being overcome as the pathogen mutates to evade recognition. This has led to a widespread reliance on fungicides and a single broad-spectrum recessive resistance provided by the mlo gene. The range of resistance genes and alleles found in wild crop relatives and landraces has been reduced in agricultural cultivars through an erosion of genetic diversity during domestication and selective breeding. Three novel major-effect adult plant resistance (APR) genes from landraces, designated Resistance to Blumeria graminis f. sp. hordei (Rbgh1 to Rbgh3), were identified in the terminal regions of barley chromosomes 5HL, 7HS, and 1HS, respectively. The phenotype of the new APR genes showed neither pronounced penetration resistance, nor the spontaneous necrosis and mesophyll cell death typical of mlo resistance, nor a whole epidermal cell hypersensitive response, typical of race-specific resistance. Instead, resistance was localized to the site of attempted penetration in an epidermal cell and was associated with cell wall appositions and cytosolic vesicle-like bodies, and lacked strong induction of reactive oxygen species. The APR genes exhibited differences in vesicle-like body sizes, their distribution, and the extent of localized 3,3-diaminobenzidine staining in individual doubled haploid lines. The results revealed a set of unique basal penetration resistance genes that offer opportunities for combining different resistance mechanisms in breeding programs for robust mildew resistance.

Expansion and Conservation of Biosynthetic Gene Clusters in Pathogenic Pyrenophora spp.

Pyrenophora is a fungal genus responsible for a number of major cereal diseases. Although fungi produce many specialised or secondary metabolites for defence and interacting with the surrounding environment, the repertoire of specialised metabolites (SM) within Pyrenophora pathogenic species remains mostly uncharted. In this study, an in-depth comparative analysis of the P. teres f. teres, P teres f. maculata and P. tritici-repentis potential to produce SMs, based on in silico predicted biosynthetic gene clusters (BGCs), was conducted using genome assemblies from PacBio DNA reads. Conservation of BGCs between the Pyrenophora species included type I polyketide synthases, terpene synthases and the first reporting of a type III polyketide synthase in P teres f. maculata. P. teres isolates exhibited substantial expansion of non-ribosomal peptide synthases relative to P. tritici-repentis, hallmarked by the presence of tailoring cis-acting nitrogen methyltransferase domains. P. teres isolates also possessed unique non-ribosomal peptide synthase (NRPS)-indole and indole BGCs, while a P. tritici-repentis phytotoxin BGC for triticone production was absent in P. teres. These differences highlight diversification between the pathogens that reflects their different evolutionary histories, host adaption and lifestyles.

Construction of a comparative genetic map in faba bean (Vicia faba L.); conservation of genome structure with Lens culinaris.

BACKGROUND: The development of genetic markers is complex and costly in species with little pre-existing genomic information. Faba bean possesses one of the largest and least studied genomes among cultivated crop plants and no gene-based genetic maps exist. Gene-based orthologous markers allow chromosomal regions and levels of synteny to be characterised between species, reveal phylogenetic relationships and chromosomal evolution, and enable targeted identification of markers for crop breeding. In this study orthologous codominant cross-species markers have been deployed to produce the first exclusively gene-based genetic linkage map of faba bean (Vicia faba), using an F6 population developed from a cross between the lines Vf6 (equina type) and Vf27 (paucijuga type). RESULTS: Of 796 intron-targeted amplified polymorphic (ITAP) markers screened, 151 markers could be used to construct a comparative genetic map. Linkage analysis revealed seven major and five small linkage groups (LGs), one pair and 12 unlinked markers. Each LG was comprised of three to 30 markers and varied in length from 23.6 cM to 324.8 cM. The map spanned a total length of 1685.8 cM. A simple and direct macrosyntenic relationship between faba bean and Medicago truncatula was evident, while faba bean and lentil shared a common rearrangement relative to M. truncatula. One hundred and four of the 127 mapped markers in the 12 LGs, which were previously assigned to M. truncatula genetic and physical maps, were found in regions syntenic between the faba bean and M. truncatula genomes. However chromosomal rearrangements were observed that could explain the difference in chromosome numbers between these three legume species. These rearrangements suggested high conservation of M. truncatula chromosomes 1, 5 and 8; moderate conservation of chromosomes 2, 3, 4 and 7 and no conservation with M. truncatula chromosome 6. Multiple PCR amplicons and comparative mapping were suggestive of small-scale duplication events in faba bean. This study also provides a preliminary indication for finer scale macrosynteny between M. truncatula, lentil and faba bean. Markers originally designed from genes on the same M. truncatula BACs were found to be grouped together in corresponding syntenic areas in lentil and faba bean. CONCLUSION: Despite the large size of the faba bean genome, comparative mapping did not reveal evidence for polyploidisation, segmental duplication, or significant rearrangements compared to M. truncatula, although a bias in the use of single locus markers may have limited the detection of duplications. Non-coding repetitive DNA or transposable element content provides a possible explanation for the difference in genome sizes. Similar patterns of rearrangements in faba bean and lentil compared to M. truncatula support phylogenetic studies dividing these species into the tribes Viceae and Trifoliae. However, substantial macrosynteny was apparent between faba bean and M. truncatula, with the exception of chromosome 6 where no orthologous markers were found, confirming previous investigations suggesting chromosome 6 is atypical. The composite map, anchored with orthologous markers mapped in M. truncatula, provides a central reference map for future use of genomic and genetic information in faba bean genetic analysis and breeding.

Two alternative recessive quantitative trait loci influence resistance to spring black stem and leaf spot in Medicago truncatula.

BACKGROUND: Knowledge of the genetic basis of plant resistance to necrotrophic pathogens is incomplete and has been characterised in relatively few pathosystems. In this study, the cytology and genetics of resistance to spring black stem and leaf spot caused by Phoma medicaginis, an economically important necrotrophic pathogen of Medicago spp., was examined in the model legume M. truncatula. RESULTS: Macroscopically, the resistant response of accession SA27063 was characterised by small, hypersensitive-like spots following inoculation while the susceptible interaction with accessions A17 and SA3054 showed necrotic lesions and spreading chlorosis. No unique cytological differences were observed during early infection (<48 h) between the resistant and susceptible genotypes, except pathogen growth was restricted to one or a few host cells in SA27063. In both interactions reactive oxygen intermediates and phenolic compounds were produced, and cell death occurred. Two F2 populations segregating for resistance to spring black stem and leaf spot were established between SA27063 and the two susceptible accessions, A17 and SA3054. The cross between SA27063 and A17 represented a wider cross than between SA27063 and SA3054, as evidenced by higher genetic polymorphism, reduced fertility and aberrant phenotypes of F2 progeny. In the SA27063 x A17 F2 population a highly significant quantitative trait locus (QTL, LOD = 7.37; P < 0.00001) named resistance to the necrotroph Phoma medicaginis one (rnpm1) genetically mapped to the top arm of linkage group 4 (LG4). rnpm1 explained 33.6% of the phenotypic variance in the population's response to infection depicted on a 1-5 scale and was tightly linked to marker AW256637. A second highly significant QTL (LOD = 6.77; P < 0.00001), rnpm2, was located on the lower arm of LG8 in the SA27063 x SA3054 map. rnpm2 explained 29.6% of the phenotypic variance and was fine mapped to a 0.8 cM interval between markers h2_16a6a and h2_21h11d. rnpm1 is tightly linked to a cluster of Toll/Interleukin1 receptor-nucleotide binding site-leucine-rich repeat (TIR-NBS-LRR) genes and disease resistance protein-like genes, while no resistance gene analogues (RGAs) are apparent in the genomic sequence of the reference accession A17 at the rnpm2 locus. CONCLUSION: The induction of defence responses and cell death in the susceptible interaction following infection by P. medicaginis suggested this pathogen is not negatively affected by these responses and may promote them. A QTL for resistance was revealed in each of two populations derived from crosses between a resistant accession and two different susceptible accessions. Both loci are recessive in nature, and the simplest explanation for the existence of two separate QTLs is the occurrence of host genotype-specific susceptibility loci that may interact with undetermined P. medicaginis virulence factors.

Transposable Element Genomic Fissuring in Pyrenophora teres Is Associated With Genome Expansion and Dynamics of Host-Pathogen Genetic Interactions.

Pyrenophora teres, P. teres f. teres (PTT) and P. teres f. maculata (PTM) cause significant diseases in barley, but little is known about the large-scale genomic differences that may distinguish the two forms. Comprehensive genome assemblies were constructed from long DNA reads, optical and genetic maps. As repeat masking in fungal genomes influences the final gene annotations, an accurate and reproducible pipeline was developed to ensure comparability between isolates. The genomes of the two forms are highly collinear, each composed of 12 chromosomes. Genome evolution in P. teres is characterized by genome fissuring through the insertion and expansion of transposable elements (TEs), a process that isolates blocks of genic sequence. The phenomenon is particularly pronounced in PTT, which has a larger, more repetitive genome than PTM and more recent transposon activity measured by the frequency and size of genome fissures. PTT has a longer cultivated host association and, notably, a greater range of host-pathogen genetic interactions compared to other Pyrenophora spp., a property which associates better with genome size than pathogen lifestyle. The two forms possess similar complements of TE families with Tc1/Mariner and LINE-like Tad-1 elements more abundant in PTT. Tad-1 was only detectable as vestigial fragments in PTM and, within the forms, differences in genome sizes and the presence and absence of several TE families indicated recent lineage invasions. Gene differences between P. teres forms are mainly associated with gene-sparse regions near or within TE-rich regions, with many genes possessing characteristics of fungal effectors. Instances of gene interruption by transposons resulting in pseudogenization were detected in PTT. In addition, both forms have a large complement of secondary metabolite gene clusters indicating significant capacity to produce an array of different molecules. This study provides genomic resources for functional genetics to help dissect factors underlying the host-pathogen interactions.

The first genetic and comparative map of white lupin (Lupinus albus L.): identification of QTLs for anthracnose resistance and flowering time, and a locus for alkaloid content.

We report the first genetic linkage map of white lupin (Lupinus albus L.). An F8 recombinant inbred line population developed from Kiev mutant x P27174 was mapped with 220 amplified fragment length polymorphism and 105 gene-based markers. The genetic map consists of 28 main linkage groups (LGs) that varied in length from 22.7 cM to 246.5 cM and spanned a total length of 2951 cM. There were seven additional pairs and 15 unlinked markers, and 12.8% of markers showed segregation distortion at P < 0.05. Syntenic relationships between Medicago truncatula and L. albus were complex. Forty-five orthologous markers that mapped between M. truncatula and L. albus identified 17 small syntenic blocks, and each M. truncatula chromosome aligned to between one and six syntenic blocks in L. albus. Genetic mapping of three important traits: anthracnose resistance, flowering time, and alkaloid content allowed loci governing these traits to be defined. Two quantitative trait loci (QTLs) with significant effects were identified for anthracnose resistance on LG4 and LG17, and two QTLs were detected for flowering time on the top of LG1 and LG3. Alkaloid content was mapped as a Mendelian trait to LG11.

Identification of Sources of Resistance to Phoma medicaginis Isolates in Medicago truncatula SARDI Core Collection Accessions, and Multigene Differentiation of Isolates.

ABSTRACT Phoma medicaginis is a necrotrophic fungal pathogen, commonly found infecting the annual medic Medicago truncatula. To differentiate eight P. medicaginis isolates, five gene regions were examined: actin, beta-tubulin, calmodulin, translation elongation factor 1-alpha (EF-1alpha), and the internal transcribed spacer ribosomal DNA. Sequence comparisons showed that specimens isolated from M. truncatula in Western Australia formed a group that was consistently different from, but allied to, a P. medicaginis var. medicaginis type specimen. EF-1alpha contained a hyper-variable 55-bp repeat unit, which forms the basis of a rapid polymerase chain reaction-based method of reliably distinguishing isolates. Characterization of three isolates showed that all exhibited a narrow host range, causing disease only in M. sativa and M. truncatula among eight commonly cultivated legume species sampled. Infection of 86 M. truncatula single-seeded accessions showed a continuous distribution in disease phenotypes, with the majority of accessions susceptible. On a 1-to-5 disease reaction scale increasing in severity, individual fungal isolates showed means of 2.6 to 3.2, and scores ranged from 1 to 4.8 among accessions. The results presented here suggest that M. truncatula harbors specific and diverse sources of resistance to individual P. medicaginis genotypes.