Mapping crown rust resistance in the oat diploid accession PI 258731 (Avena strigosa).

Oat crown rust, caused by Puccinia coronata Corda f. sp. avenae Eriks. (Pca), is a major biotic impediment to global oat production. Crown rust resistance has been described in oat diploid species A. strigosa accession PI 258731 and resistance from this accession has been successfully introgressed into hexaploid A. sativa germplasm. The current study focuses on 1) mapping the location of QTL containing resistance and evaluating the number of quantitative trait loci (QTL) conditioning resistance in PI 258731; 2) understanding the relationship between the original genomic location in A. strigosa and the location of the introgression in the A. sativa genome; 3) identifying molecular markers tightly linked with PI 258731 resistance loci that could be used for marker assisted selection and detection of this resistance in diverse A. strigosa accessions. To achieve this, A. strigosa accessions, PI 258731 and PI 573582 were crossed to produce 168 F5:6 recombinant inbred lines (RILs) through single seed descent. Parents and RILs were genotyped with the 6K Illumina SNP array which generated 168 segregating SNPs. Seedling reactions to two isolates of Pca (races TTTG, QTRG) were conditioned by two genes (0.6 cM apart) in this population. Linkage mapping placed these two resistant loci to 7.7 (QTRG) to 8 (TTTG) cM region on LG7. Field reaction data was used for QTL analysis and the results of interval mapping (MIM) revealed a major QTL (QPc.FD-AS-AA4) for field resistance. SNP marker assays were developed and tested in 125 diverse A. strigosa accessions that were rated for crown rust resistance in Baton Rouge, LA and Gainesville, FL and as seedlings against races TTTG and QTRG. Our data proposed SNP marker GMI_ES17_c6425_188 as a candidate for use in marker-assisted selection, in addition to the marker GMI_ES02_c37788_255 suggested by Rine's group, which provides an additional tool in facilitating the utilization of this gene in oat breeding programs.

Virulence Patterns of Oat Crown Rust in Australia - Season 2022.

Puccinia coronata f. sp. avenae (Pca) is an important foliar pathogen of oat which causes crown rust disease. The virulence profile of 48 Pca isolates derived from different locations in Australia was characterized using a collection of oat lines often utilized in rust surveys in the United States and Australia. This analysis indicates that Pca populations in Eastern Australia are broadly virulent, which contrasts with the population in Western Australia (WA). Several oat lines/Pc genes are effective against all rust samples collected from WA, suggesting they may provide useful resistance in this region if deployed in combination. We identified 19 lines from the United States oat differential set that display disease resistance to Pca in WA, with some in agreement with previous rust survey reports. We adopted the 10-letter nomenclature system to define oat crown rust races in Australia and compare the frequency of those virulence traits to published data from the United States. Based on this nomenclature, 42 unique races were detected among the 48 isolates, reflecting the high diversity of virulence phenotypes for Pca in Australia. Nevertheless, the Pca population in the United States is substantially more broadly virulent than that of Australia. Close examination of resistance profiles for the oat differential set lines after infection with Pca supports hypotheses of allelism or redundancy among Pc genes or the presence of several resistance genes in some oat differential lines. These findings illustrate the need to deconvolute the oat differential set using molecular tools.[Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Genome-Enabled Analysis of Population Dynamics and Virulence-Associated Loci in the Oat Crown Rust Fungus Puccinia coronata f. sp. avenae.

Puccinia coronata f. sp. avenae (Pca) is an important fungal pathogen causing crown rust that impacts oat production worldwide. Genetic resistance for crop protection against Pca is often overcome by the rapid virulence evolution of the pathogen. This study investigated the factors shaping adaptive evolution of Pca using pathogen populations from distinct geographic regions within the United States and South Africa. Phenotypic and genome-wide sequencing data of these diverse Pca collections, including 217 isolates, uncovered phylogenetic relationships and established distinct genetic composition between populations from northern and southern regions from the United States and South Africa. The population dynamics of Pca involve a bidirectional movement of inoculum between northern and southern regions of the United States and contributions from clonality and sexuality. The population from South Africa is solely clonal. A genome-wide association study (GWAS) employing a haplotype-resolved Pca reference genome was used to define 11 virulence-associated loci corresponding to 25 oat differential lines. These regions were screened to determine candidate Avr effector genes. Overall, the GWAS results allowed us to identify the underlying genetic factors controlling pathogen recognition in an oat differential set used in the United States to assign pathogen races (pathotypes). Key GWAS findings support complex genetic interactions in several oat lines, suggesting allelism among resistance genes or redundancy of genes included in the differential set, multiple resistance genes recognizing genetically linked Avr effector genes, or potentially epistatic relationships. A careful evaluation of the composition of the oat differential set accompanied by the development or implementation of molecular markers is recommended. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Genotypic and Resistance Profile Analysis of Two Oat Crown Rust Differential Sets Urge Coordination and Standardization.

Puccinia coronata f. sp. avenae is the causal agent of the disease known as crown rust, which represents a bottleneck in oat production worldwide. Characterization of pathogen populations often involves race (pathotype) assignments using differential sets, which are not uniform across countries. This study compared the virulence profiles of 25 P. coronata f. sp. avenae isolates from Australia using two host differential sets, one from Australia and one from the United States. These differential sets were also genotyped using diversity arrays technology sequencing technology. Phenotypic and genotypic discrepancies were detected on 8 out of 29 common lines between the two sets, indicating that pathogen race assignments based on those lines are not comparable. To further investigate molecular markers that could assist in the stacking of rust resistance genes important for Australia, four published Pc91-linked markers were validated across the differential sets and then screened across a collection of 150 oat cultivars. Drover, Aladdin, and Volta were identified as putative carriers of the Pc91 locus. This is the first report to confirm that the cultivar Volta carries Pc91 and demonstrates the value of implementing molecular markers to characterize materials in breeding pools of oat. Overall, our findings highlight the necessity of examining seed stocks using pedigree and molecular markers to ensure seed uniformity and bring robustness to surveillance methodologies. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Three decades of rust surveys in the United States reveal drastic virulence changes in oat crown rust.

To better understand how the pathogenicity of the oat crown rust pathogen, Puccinia coronata f. sp. avenae (Pca), has changed in the United States, 30 years of USDA survey isolates (n=5,456) tested on 30-40 differential lines were analyzed for overall and Pc resistance gene specific virulence trends and correlations. Pca is incredibly pathologically diverse with 88% of races represented by a single isolate. There are a slightly higher proportion of unique races from the Northern region of the United States and for one fourth of the years, Northern region isolates were significantly more virulent than Southern isolates which supports the idea that sexual recombination in this region is mediated by the alternate host as a major factor in creating new races. However, there is also support for regular isolate movement between North and South regions as isolates in the United States are steadily accumulating virulences at a rate of 0.35 virulences per year. Virulence significantly increased for 23 and decreased for 4 of the 40 differential lines. In the past few years, virulence has reached 90% or greater for 16 differential lines. There were also strong correlations in virulence for certain Pc genes that are likely identical, allelic, or target the same or closely linked pathogen effectors (e.g. Pc39, Pc55, and Pc71), and the results were largely in concordance with recent GWAS effector studies using USDA isolate subsets. Understanding changes in Pca pathogenicity is essential for the responsible deployment and management of Pc resistance genes for sustainable and profitable oat production.

A geminivirus crosses the monocot-dicot boundary and acts as a viral vector for gene silencing and genome editing.

INTRODUCTION: Members of the family Geminiviridae have been reported to infect either a monocot plant or a dicot plant, but not both. This study reports a geminivirus, Wheat Dwarf India Virus (WDIV), first identified in wheat, that is capable of infecting both monocot and dicot plants and acting as a viral vector. OBJECTIVES: This study was aimed at developing a broad host range viral vector system for reverse genetics and genome editing. METHODS: Here we used a wheat isolate of WDIV and Ageratum yellow leaf curl betasatellite (AYLCB) for infectivity assays and vector development. We performed Agrobacterium-mediated inoculation of WDIV and AYLCB in wheat, oat, barley, corn, soybean, and tobacco. To examine the potential of WDIV to act as a viral vector, we modified the WDIV genome and cloned DNA fragments of the phytoene desaturase (PDS) genes from wheat and tobacco, separately. For gene editing experiments, tobacco lines expressing Cas9 were infiltrated with a WDIV-based vector carrying gRNA targeting the PDS gene. RESULTS: About 80 to 90% of plants inoculated with infectious clones of WDIV alone or WDIV together with AYLCB showed mild symptoms, whereas some plants showed more prominent symptoms. WDIV and AYLCB were detected in the systemically infected leaves of all the plant species. Furthermore, the inoculation of the WDIV vector carrying PDS fragments induced silencing of the PDS gene in both wheat and tobacco plants. We also observed high-efficiency genome editing in the Cas9-expressing tobacco plants that were inoculated with WDIV vector-carrying gRNA. CONCLUSION: Detection of WDIV in naturally infected wheat, barley, and sugarcane in the field and its ability to systemically infect wheat, oat, barley, corn, soybean, and tobacco under laboratory conditions, provides compelling evidence that WDIV is the first geminivirus identified with the capability of infecting both monocot and dicot plant species. The wide host range of WDIV can be exploited for developing a single vector system for high-throughput genome editing in many plant species.

Sequencing 4.3 million mutations in wheat promoters to understand and modify gene expression.

Wheat is an important contributor to global food security, and further improvements are required to feed a growing human population. Functional genetics and genomics tools can help us to understand the function of different genes and to engineer beneficial changes. In this study, we used a promoter capture assay to sequence 2-kb regions upstream of all high-confidence annotated genes from 1,513 mutagenized plants from the tetraploid wheat variety Kronos. We identified 4.3 million induced mutations with an accuracy of 99.8%, resulting in a mutation density of 41.9 mutations per kb. We also remapped Kronos exome capture reads to Chinese Spring RefSeq v1.1, identified 4.7 million mutations, and predicted their effects on annotated genes. Using these predictions, we identified 59% more nonsynonymous substitutions and 49% more truncation mutations than in the original study. To show the biological value of the promoter dataset, we selected two mutations within the promoter of the VRN-A1 vernalization gene. Both mutations, located within transcription factor binding sites, significantly altered VRN-A1 expression, and one reduced the number of spikelets per spike. These publicly available sequenced mutant datasets provide rapid and inexpensive access to induced variation in the promoters and coding regions of most wheat genes. These mutations can be used to understand and modulate gene expression and phenotypes for both basic and commercial applications, where limited governmental regulations can facilitate deployment. These mutant collections, together with gene editing, provide valuable tools to accelerate functional genetic studies in this economically important crop.

Evaluation of Stem Rust Disease in Wheat Fields by Drone Hyperspectral Imaging.

Detecting plant disease severity could help growers and researchers study how the disease impacts cereal crops to make timely decisions. Advanced technology is needed to protect cereals that feed the increasing population using fewer chemicals; this may lead to reduced labor usage and cost in the field. Accurate detection of wheat stem rust, an emerging threat to wheat production, could inform growers to make management decisions and assist plant breeders in making line selections. A hyperspectral camera mounted on an unmanned aerial vehicle (UAV) was utilized in this study to evaluate the severity of wheat stem rust disease in a disease trial containing 960 plots. Quadratic discriminant analysis (QDA) and random forest classifier (RFC), decision tree classification, and support vector machine (SVM) were applied to select the wavelengths and spectral vegetation indices (SVIs). The trial plots were divided into four levels based on ground truth disease severities: class 0 (healthy, severity 0), class 1 (mildly diseased, severity 1-15), class 2 (moderately diseased, severity 16-34), and class 3 (severely diseased, highest severity observed). The RFC method achieved the highest overall classification accuracy (85%). For the spectral vegetation indices (SVIs), the highest classification rate was recorded by RFC, and the accuracy was 76%. The Green NDVI (GNDVI), Photochemical Reflectance Index (PRI), Red-Edge Vegetation Stress Index (RVS1), and Chlorophyll Green (Chl green) were selected from 14 SVIs. In addition, binary classification of mildly diseased vs. non-diseased was also conducted using the classifiers and achieved 88% classification accuracy. This highlighted that hyperspectral imaging was sensitive enough to discriminate between low levels of stem rust disease vs. no disease. The results of this study demonstrated that drone hyperspectral imaging can discriminate stem rust disease levels so that breeders can select disease-resistant varieties more efficiently. The detection of low disease severity capability of drone hyperspectral imaging can help farmers identify early disease outbreaks and enable more timely management of their fields. Based on this study, it is also possible to build a new inexpensive multispectral sensor to diagnose wheat stem rust disease accurately.

Genetic Analysis and Physical Mapping of Oat Adult Plant Resistance Loci Against Puccinia coronata f. sp. avenae.

Six quantitative trait loci (QTLs) for adult plant resistance against oat crown rust (Puccinia coronata f. sp. avenae) were identified from mapping three recombinant inbred populations. Using genotyping-by-sequencing with markers called against the OT3098 v1 reference genome, the QTLs were mapped on six different chromosomes: Chr1D, Chr4D, Chr5A, Chr5D, Chr7A, and Chr7C. Composite interval mapping with marker cofactor selection showed that the phenotypic variance explained by all identified QTLs for coefficient of infection range from 12.2 to 46.9%, whereas heritability estimates ranged from 0.11 to 0.38. The significant regions were narrowed down to intervals of 3.9 to 25 cM, equivalent to physical distances of 11 to 133 Mb. At least two flanking single-nucleotide polymorphism markers were identified within 10 cM of each QTL that could be used in marker-assisted introgression, pyramiding, and selection. The additive effects of the QTLs in each population were determined using single-nucleotide polymorphism haplotype data, which showed a significantly lower coefficient of infection in lines homozygous for the resistant alleles. Analysis of pairwise linkage disequilibrium also revealed high correlation of markers and presence of linkage blocks in the significant regions. To further facilitate marker-assisted breeding, polymerase chain reaction allelic competitive extension (PACE) markers for the adult plant resistance loci were developed. Putative candidate genes were also identified in each of the significant regions, which include resistance gene analogs that encode for kinases, ligases, and predicted receptors of avirulence proteins from pathogens.

Archetypes of inflorescence: genome-wide association networks of panicle morphometric, growth, and disease variables in a multiparent oat population.

There is limited information regarding the morphometric relationships of panicle traits in oat (Avena sativa) and their contribution to phenology and growth, physiology, and pathology traits important for yield. To model panicle growth and development and identify genomic regions associated with corresponding traits, 10 diverse spring oat mapping populations (n = 2,993) were evaluated in the field and 9 genotyped via genotyping-by-sequencing. Representative panicles from all progeny individuals, parents, and check lines were scanned, and images were analyzed using manual and automated techniques, resulting in over 60 unique panicle, rachis, and spikelet variables. Spatial modeling and days to heading were used to account for environmental and phenological variances, respectively. Panicle variables were intercorrelated, providing reproducible archetypal and growth models. Notably, adult plant resistance for oat crown rust was most prominent for taller, stiff stalked plants having a more open panicle structure. Within and among family variance for panicle traits reflected the moderate-to-high heritability and mutual genome-wide associations (hotspots) with numerous high-effect loci. Candidate genes and potential breeding applications are discussed. This work adds to the growing genetic resources for oat and provides a unique perspective on the genetic basis of panicle architecture in cereal crops.

Comparison of meiotic transcriptomes of three maize inbreds with different origins reveals differences in cell cycle and recombination.

BACKGROUND: Cellular events during meiosis can differ between inbred lines in maize. Substantial differences in the average numbers of chiasmata and double-strand breaks (DSBs) per meiotic cell have been documented among diverse inbred lines of maize: CML228, a tropical maize inbred line, B73 and Mo17, temperate maize lines. To determine if gene expression might explain these observed differences, an RNA-Seq experiment was performed on CML228 male meiocytes which was compared to B73 and Mo17 male meiocytes, where plants were grown in the same controlled environment. RESULTS: We found that a few DSB-repair/meiotic genes which promote class I crossovers (COs) and the Zyp1 gene which limits newly formed class I COs were up-regulated, whereas Mus81 homolog 2 which promotes class II COs was down-regulated in CML228. Although we did not find enriched gene ontology (GO) categories directly related to meiosis, we found that GO categories in membrane, localization, proteolysis, energy processes were up-regulated in CML228, while chromatin remodeling, epigenetic regulation, and cell cycle related processes including meiosis related cell cycle processes were down-regulated in CML228. The degree of similarity in expression patterns between the three maize lines reflect their genetic relatedness: B73 and Mo17 had similar meiotic expressions and CML228 had a more distinct expression profile. CONCLUSIONS: We found that meiotic related genes were mostly conserved among the three maize inbreds except for a few DSB-repair/meiotic genes. The findings that the molecular players in limiting class I CO formation (once CO assurance is achieved) were up-regulated and those involved in promoting class II CO formation were down-regulated in CML228 agree with the lower chiasmata number observed in CML228 previously. In addition, epigenetics such as chromatin remodeling and histone modification might play a role. Transport and energy-related processes was up-regulated and Cyclin13 was down-regulated in CML228. The direction of gene expression of these processes agree with that previously found in meiotic tissues compared with vegetative tissues. In summary, we used different natural maize inbred lines from different climatic conditions and have shown their differences in expression landscape in male meiocytes.

A reference-anchored oat linkage map reveals quantitative trait loci conferring adult plant resistance to crown rust (Puccinia coronata f. sp. avenae).

KEY MESSAGE: We mapped three adult plant resistance (APR) loci on oat chromosomes 4D and 6C and developed flanking KASP/PACE markers for marker-assisted selection and gene pyramiding. Using sequence orthology search and the available oat genomic and transcriptomic data, we surveyed these genomic regions for genes that may control disease resistance. Sources of durable disease resistance are needed to minimize yield losses in cultivated oat caused by crown rust (Puccinia coronata f. sp. avenae). In this study, we developed five oat recombinant inbred line mapping populations to identify sources of adult plant resistance from crosses between five APR donors and Otana, a susceptible variety. The preliminary bulk segregant mapping based on allele frequencies showed two regions in linkage group Mrg21 (Chr4D) that are associated with the APR phenotype in all five populations. Six markers from these regions in Chr4D were converted to high-throughput allele specific PCR assays and were used to genotype all individuals in each population. Simple interval mapping showed two peaks in Chr4D, named QPc.APR-4D.1 and QPc.APR-4D.2, which were detected in the OtanaA/CI4706-2 and OtanaA/CI9416-2 and in the Otana/PI189733, OtanaD/PI260616, and OtanaA/CI8000-4 populations, respectively. These results were validated by mapping two entire populations, Otana/PI189733 and OtanaA/CI9416, genotyped using Illumina HiSeq, in which polymorphisms were called against the OT3098 oat reference genome. Composite interval mapping results confirmed the presence of the two quantitative trait loci (QTL) located on oat chromosome 4D and an additional QTL with a smaller effect located on chromosome 6C. This mapping approach also narrowed down the physical intervals to between 5 and 19 Mb, and indicated that QPc.APR-4D.1, QPc.APR-4D.2, and QPc.APR-6C explained 43.4%, 38.5%, and 21.5% of the phenotypic variation, respectively. In a survey of the gene content of each QTL, several clusters of disease resistance genes that may contribute to APR were found. The allele specific PCR markers developed for these QTL regions would be beneficial for marker-assisted breeding, gene pyramiding, and future cloning of resistance genes from oat.

A chromosome-level, fully phased genome assembly of the oat crown rust fungus Puccinia coronata f. sp. avenae: a resource to enable comparative genomics in the cereal rusts.

Advances in sequencing technologies as well as development of algorithms and workflows have made it possible to generate fully phased genome references for organisms with nonhaploid genomes such as dikaryotic rust fungi. To enable discovery of pathogen effectors and further our understanding of virulence evolution, we generated a chromosome-scale assembly for each of the 2 nuclear genomes of the oat crown rust pathogen, Puccinia coronata f. sp. avenae (Pca). This resource complements 2 previously released partially phased genome references of Pca, which display virulence traits absent in the isolate of historic race 203 (isolate Pca203) which was selected for this genome project. A fully phased, chromosome-level reference for Pca203 was generated using PacBio reads and Hi-C data and a recently developed pipeline named NuclearPhaser for phase assignment of contigs and phase switch correction. With 18 chromosomes in each haplotype and a total size of 208.10 Mbp, Pca203 has the same number of chromosomes as other cereal rust fungi such as Puccinia graminis f. sp. tritici and Puccinia triticina, the causal agents of wheat stem rust and wheat leaf rust, respectively. The Pca203 reference marks the third fully phased chromosome-level assembly of a cereal rust to date. Here, we demonstrate that the chromosomes of these 3 Puccinia species are syntenous and that chromosomal size variations are primarily due to differences in repeat element content.

Genome-wide investigation of maize RAD51 binding affinity through phage display.

BACKGROUND: RAD51 proteins, which are conserved in all eukaryotes, repair DNA double-strand breaks. This is critical to homologous chromosome pairing and recombination enabling successful reproduction. Work in Arabidopsis suggests that RAD51 also plays a role in plant defense; the Arabidopsis rad51 mutant is more susceptible to Pseudomonas syringae. However, the defense functions of RAD51 and the proteins interacting with RAD51 have not been thoroughly investigated in maize. Uncovering ligands of RAD51 would help to understand meiotic recombination and possibly the role of RAD51 in defense. This study used phage display, a tool for discovery of protein-protein interactions, to search for proteins interacting with maize RAD51A1. RESULTS: Maize RAD51A1 was screened against a random phage library. Eleven short peptide sequences were recovered from 15 phages which bound ZmRAD51A1 in vitro; three sequences were found in multiple successfully binding phages. Nine of these phage interactions were verified in vitro through ELISA and/or dot blotting. BLAST searches did not reveal any maize proteins which contained the exact sequence of any of the selected phage peptides, although one of the selected phages had a strong alignment (E-value = 0.079) to a binding domain of maize BRCA2. Therefore, we designed 32 additional short peptides using amino acid sequences found in the predicted maize proteome. These peptides were not contained within phages. Of these synthesized peptides, 14 bound to ZmRAD51A1 in a dot blot experiment. These 14 sequences are found in known maize proteins including transcription factors putatively involved in defense. CONCLUSIONS: These results reveal several peptides which bind ZmRAD51A1 and support a potential role for ZmRAD51A1 in transcriptional regulation and plant defense. This study also demonstrates the applicability of phage display to basic science questions, such as the search for binding partners of a known protein, and raises the possibility of an iterated approach to test peptide sequences that closely but imperfectly align with the selected phages.

Study of Triticum aestivum Resistome in Response to Wheat dwarf India Virus Infection.

Susceptible and resistant germplasm respond differently to pathogenic attack, including virus infections. We compared the transcriptome changes between a resistant wheat cultivar, Sonalika, and a susceptible cultivar, WL711, to understand this process in wheat against wheat dwarf India virus (WDIV) infection. A total of 2760 and 1853 genes were differentially expressed in virus-infected and mock-inoculated Sonalika, respectively, compared to WL711. The overrepresentation of genes involved in signaling, hormone metabolism, enzymes, secondary metabolites, proteolysis, and transcription factors was documented, including the overexpression of multiple PR proteins. We hypothesize that the virus resistance in Sonalika is likely due to strong intracellular surveillance via the action of multiple PR proteins (PR1, RAR1, and RPM1) and ChiB. Other genes such as PIP1, LIP1, DnaJ, defensins, oxalate oxidase, ankyrin repeat protein, serine-threonine kinase, SR proteins, beta-1,3-glucanases, and O-methyltransferases had a significant differential expression and play roles in stress tolerance, may also be contributing towards the virus resistance in Sonalika. In addition, we identified putative genes with unknown functions, which are only expressed in response to WDIV infection in Sonalika. The role of these genes could be further validated and utilized in engineering resistance in wheat and other crops.

Identification of Main-Effect and Environmental Interaction QTL and Their Candidate Genes for Drought Tolerance in a Wheat RIL Population Between Two Elite Spring Cultivars.

Understanding the genetics of drought tolerance can expedite the development of drought-tolerant cultivars in wheat. In this study, we dissected the genetics of drought tolerance in spring wheat using a recombinant inbred line (RIL) population derived from a cross between a drought-tolerant cultivar, 'Reeder' (PI613586), and a high-yielding but drought-susceptible cultivar, 'Albany.' The RIL population was evaluated for grain yield (YLD), grain volume weight (GVW), thousand kernel weight (TKW), plant height (PH), and days to heading (DH) at nine different environments. The Infinium 90 k-based high-density genetic map was generated using 10,657 polymorphic SNP markers representing 2,057 unique loci. Quantitative trait loci (QTL) analysis detected a total of 11 consistent QTL for drought tolerance-related traits. Of these, six QTL were exclusively identified in drought-prone environments, and five were constitutive QTL (identified under both drought and normal conditions). One major QTL on chromosome 7B was identified exclusively under drought environments and explained 13.6% of the phenotypic variation (PV) for YLD. Two other major QTL were detected, one each on chromosomes 7B and 2B under drought-prone environments, and explained 14.86 and 13.94% of phenotypic variation for GVW and YLD, respectively. One novel QTL for drought tolerance was identified on chromosome 2D. In silico expression analysis of candidate genes underlaying the exclusive QTLs associated with drought stress identified the enrichment of ribosomal and chloroplast photosynthesis-associated proteins showing the most expression variability, thus possibly contributing to stress response by modulating the glycosyltransferase (TraesCS6A01G116400) and hexosyltransferase (TraesCS7B01G013300) unique genes present in QTL 21 and 24, respectively. While both parents contributed favorable alleles to these QTL, unexpectedly, the high-yielding and less drought-tolerant parent contributed desirable alleles for drought tolerance at four out of six loci. Regardless of the origin, all QTL with significant drought tolerance could assist significantly in the development of drought-tolerant wheat cultivars, using genomics-assisted breeding approaches.

Genome-Wide Association Mapping for Yield and Related Traits Under Drought Stressed and Non-stressed Environments in Wheat.

Understanding the genetics of drought tolerance in hard red spring wheat (HRSW) in northern USA is a prerequisite for developing drought-tolerant cultivars for this region. An association mapping (AM) study for drought tolerance in spring wheat in northern USA was undertaken using 361 wheat genotypes and Infinium 90K single-nucleotide polymorphism (SNP) assay. The genotypes were evaluated in nine different locations of North Dakota (ND) for plant height (PH), days to heading (DH), yield (YLD), test weight (TW), and thousand kernel weight (TKW) under rain-fed conditions. Rainfall data and soil type of the locations were used to assess drought conditions. A mixed linear model (MLM), which accounts for population structure and kinship (PC+K), was used for marker-trait association. A total of 69 consistent QTL involved with drought tolerance-related traits were identified, with p ≤ 0.001. Chromosomes 1A, 3A, 3B, 4B, 4D, 5B, 6A, and 6B were identified to harbor major QTL for drought tolerance. Six potential novel QTL were identified on chromosomes 3D, 4A, 5B, 7A, and 7B. The novel QTL were identified for DH, PH, and TKW. The findings of this study can be used in marker-assisted selection (MAS) for drought-tolerance breeding in spring wheat.

Identification of Candidate Susceptibility Genes to Puccinia graminis f. sp. tritici in Wheat.

Wheat stem rust disease caused by Puccinia graminis f. sp. tritici (Pgt) is a global threat to wheat production. Fast evolving populations of Pgt limit the efficacy of plant genetic resistance and constrain disease management strategies. Understanding molecular mechanisms that lead to rust infection and disease susceptibility could deliver novel strategies to deploy crop resistance through genetic loss of disease susceptibility. We used comparative transcriptome-based and orthology-guided approaches to characterize gene expression changes associated with Pgt infection in susceptible and resistant Triticum aestivum genotypes as well as the non-host Brachypodium distachyon. We targeted our analysis to genes with differential expression in T. aestivum and genes suppressed or not affected in B. distachyon and report several processes potentially linked to susceptibility to Pgt, such as cell death suppression and impairment of photosynthesis. We complemented our approach with a gene co-expression network analysis to identify wheat targets to deliver resistance to Pgt through removal or modification of putative susceptibility genes.

Increased virulence of Puccinia coronata f. sp.avenae populations through allele frequency changes at multiple putative Avr loci.

Pathogen populations are expected to evolve virulence traits in response to resistance deployed in agricultural settings. However, few temporal datasets have been available to characterize this process at the population level. Here, we examined two temporally separated populations of Puccinia coronata f. sp. avenae (Pca), which causes crown rust disease in oat (Avena sativa) sampled from 1990 to 2015. We show that a substantial increase in virulence occurred from 1990 to 2015 and this was associated with a genetic differentiation between populations detected by genome-wide sequencing. We found strong evidence for genetic recombination in these populations, showing the importance of the alternate host in generating genotypic variation through sexual reproduction. However, asexual expansion of some clonal lineages was also observed within years. Genome-wide association analysis identified seven Avr loci associated with virulence towards fifteen Pc resistance genes in oat and suggests that some groups of Pc genes recognize the same pathogen effectors. The temporal shift in virulence patterns in the Pca populations between 1990 and 2015 is associated with changes in allele frequency in these genomic regions. Nucleotide diversity patterns at a single Avr locus corresponding to Pc38, Pc39, Pc55, Pc63, Pc70, and Pc71 showed evidence of a selective sweep associated with the shift to virulence towards these resistance genes in all 2015 collected isolates.

Epigenetic regulation of gene expression improves Fusarium head blight resistance in durum wheat.

Eight advanced durum-breeding lines were treated with 5-methyl-azacytidine to test the feasibility of generating sources of Fusarium head blight (FHB) resistance. Of the 800 treated seeds, 415 germinated and were advanced up to four (M4) generations by selfing. Thirty-two of the resulting 415 M4 lines were selected following preliminary screening and were further tested for FHB resistance for three years at two field locations, and in the greenhouse. Five of the 32 M4 lines showed less than 30% disease severity, as compared to the parental lines and susceptible checks. Fusarium-damaged kernels and deoxynivalenol analyses supported the findings of the field and greenhouse disease assessments. Two of the most resistant M4 lines were crossed to a susceptible parent, advanced to third generation (BC1:F3) and were tested for stability and inheritance of the resistance. About, one third of the BC1:F3 lines showed FHB resistance similar to their M4 parents. The overall methylation levels (%) were compared using FASTmC method, which did not show a significant difference between M4 and parental lines. However, transcriptome analysis of one M4 line revealed significant number of differentially expressed genes related to biosynthesis of secondary metabolites, MAPK signaling, photosynthesis, starch and sucrose metabolism, plant hormone signal transduction and plant-pathogen interaction pathways, which may have helped in improved FHB resistance.