Plant disease resistance (R) genes control the recognition of specific pathogens and activate subsequent defense responses. We show that the Arabidopsis thaliana locus RESISTANCE TO POWDERY MILDEW8 (RPW8) contains two naturally polymorphic, dominant R genes, RPW8.1 and RPW8.2, which individually control resistance to a broad range of powdery mildew pathogens. Although the predicted RPW8.1 and RPW8.2 proteins are different from the previously characterized R proteins, they induce localized, salicylic acid-dependent defenses similar to those induced by R genes that control specific resistance. Apparently, broad-spectrum resistance mediated by RPW8 uses the same mechanisms as specific resistance.

Two types of symptom are caused by P. teres. These are net-type lesions caused by P. teres f. teres and spot-type lesions caused by P. teres f. maculata. The net-like symptom, for which the disease was originally named, has characteristic narrow, dark-brown, longitudinal and transverse striations on infected leaves. The spot form symptom consists of dark-brown, circular to elliptical lesions surrounded by a chlorotic or necrotic halo of varying width.

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.

BackgroundThe 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.ResultsCodingQuarry 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.ConclusionsWe 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.

Pyrenophora teres f. sp. teres is the cause of net form of net blotch (NFNB), an economically important foliar disease in barley (Hordeum vulgare). Net and spot forms of net blotch are widely controlled using site-specific systemic fungicides. Although resistance to succinate dehydrogenase inhibitors and quinone outside inhibitors has been addressed before in net blotches, mechanisms controlling demethylation inhibitor resistance have not yet been reported at the molecular level. Here we report the isolation of strains of NFNB in Australia since 2013 resistant to a range of demethylase inhibitor fungicides. Cyp51A:KO103-A1, an allele with the mutation F489L, corresponding to the archetype F495I in Aspergillus fumigatus, was only present in resistant strains and was correlated with resistance factors to various demethylase inhibitors ranging from 1.1 for epoxiconazole to 31.7 for prochloraz. Structural in silico modeling of the sensitive and resistant CYP51A proteins docked with different demethylase inhibitor fungicides showed how the interaction of F489L within the heme cavity produced a localized constriction of the region adjacent to the docking site that is predicted to result in lower binding affinities. Resistant strains also displayed enhanced induced expression of the two Cyp51A paralogs and of Cyp51B genes. While Cyp51B was found to be constitutively expressed in the absence of fungicide, Cyp51A was only detected at extremely low levels. Under fungicide induction, expression of Cyp51B, Cyp51A2, and Cyp51A1 was shown to be 1.6-, 3,- and 5.3-fold higher, respectively in the resistant isolate compared to the wild type. These increased levels of expression were not supported by changes in the promoters of any of the three genes. The implications of these findings on demethylase inhibitor activity will require current net blotch management strategies to be reconsidered in order to avoid the development of further resistance and preserve the lifespan of fungicides in use.

We report the first gene-based linkage map of Lupinus angustifolius (narrow-leafed lupin) and its comparison to the partially sequenced genome of Medicago truncatula. The map comprises 382 loci in 20 major linkage groups, two triplets, three pairs and 11 unlinked loci and is 1,846 cM in length. The map was generated from the segregation of 163 RFLP markers, 135 gene-based PCR markers, 75 AFLP and 4 AFLP-derived SCAR markers in a mapping population of 93 recombinant inbred lines, derived from a cross between domesticated and wild-type parents. This enabled the mapping of five major genes controlling key domestication traits in L. angustifolius. Using marker sequence data, the L. angustifolius genetic map was compared to the partially completed M. truncatula genome sequence. We found evidence of conserved synteny in some regions of the genome despite the wide evolutionary distance between these legume species. We also found new evidence of widespread duplication within the L. angustifolius genome.

We report the first genetic linkage map of white lupin (Lupinus albus L.). An F8 recombinant inbred line population developed from Kiev mutant × 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.

Background: The development of genetic markers is complex and costly in species with little preexisting 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 F 6 population developed from a cross between the lines Vf6 (equina type) and Vf27 (paucijuga type).