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.

De Novo Genome Assembly of the Japanese Wheat Cultivar Norin 61 Highlights Functional Variation in Flowering Time and Fusarium-Resistant Genes in East Asian Genotypes.

Bread wheat is a major crop that has long been the focus of basic and breeding research. Assembly of its genome has been difficult because of its large size and allohexaploid nature (AABBDD genome). Following the first reported assembly of the genome of the experimental strain Chinese Spring (CS), the 10+ Wheat Genomes Project was launched to produce multiple assemblies of worldwide modern cultivars. The only Asian cultivar in the project is Norin 61, a representative Japanese cultivar adapted to grow across a broad latitudinal range, mostly characterized by a wet climate and a short growing season. Here, we characterize the key aspects of its chromosome-scale genome assembly spanning 15 Gb with a raw scaffold N50 of 22 Mb. Analysis of the repetitive elements identified chromosomal regions unique to Norin 61 that encompass a tandem array of the pathogenesis-related 13 family. We report novel copy-number variations in the B homeolog of the florigen gene FT1/VRN3, pseudogenization of its D homeolog and the association of its A homeologous alleles with the spring/winter growth habit. Furthermore, the Norin 61 genome carries typical East Asian functional variants different from CS, ranging from a single nucleotide to multi-Mb scale. Examples of such variation are the Fhb1 locus, which confers Fusarium head-blight resistance, Ppd-D1a, which confers early flowering, Glu-D1f for Asian noodle quality and Rht-D1b, which introduced semi-dwarfism during the green revolution. The adoption of Norin 61 as a reference assembly for functional and evolutionary studies will enable comprehensive characterization of the underexploited Asian bread wheat diversity.

Multiple wheat genomes reveal global variation in modern breeding.

Advances in genomics have expedited the improvement of several agriculturally important crops but similar efforts in wheat (Triticum spp.) have been more challenging. This is largely owing to the size and complexity of the wheat genome1, and the lack of genome-assembly data for multiple wheat lines2,3. Here we generated ten chromosome pseudomolecule and five scaffold assemblies of hexaploid wheat to explore the genomic diversity among wheat lines from global breeding programs. Comparative analysis revealed extensive structural rearrangements, introgressions from wild relatives and differences in gene content resulting from complex breeding histories aimed at improving adaptation to diverse environments, grain yield and quality, and resistance to stresses4,5. We provide examples outlining the utility of these genomes, including a detailed multi-genome-derived nucleotide-binding leucine-rich repeat protein repertoire involved in disease resistance and the characterization of Sm16, a gene associated with insect resistance. These genome assemblies will provide a basis for functional gene discovery and breeding to deliver the next generation of modern wheat cultivars.

NRLMFß: Beta-distribution-rescored neighborhood regularized logistic matrix factorization for improving the performance of drug-target interaction prediction.

Techniques for predicting interactions between a drug and a target (protein) are useful for strategic drug repositioning. Neighborhood regularized logistic matrix factorization (NRLMF) is one of the state-of-the-art drug-target interaction prediction methods; it is based on a statistical model using the Bernoulli distribution. However, the prediction is not accurate when drug-target interaction pairs have less interaction information (e.g., the sum of the number of ligands for a target and the number of target proteins for a drug). This study aimed to address this issue by proposing NRLMF with beta distribution rescoring (NRLMFß), which is an algorithm to improve the score of NRLMF. The score of NRLMFß is equivalent to the value of the original NRLMF score when the concentration of the beta distribution becomes infinity. The beta distribution is known as a conjugative prior distribution of the Bernoulli distribution and can reflect the amount of interaction information to its shape based on Bayesian inference. Therefore, in NRLMFß, the beta distribution was used for rescoring the NRLMF score. In the evaluation experiment, we measured the average values of area under the receiver operating characteristics and area under precision versus recall and the 95% confidence intervals. The performance of NRLMFß was found to be better than that of NRLMF in the four types of benchmark datasets. Thus, we concluded that NRLMFß improved the prediction accuracy of NRLMF. The source code is available at https://github.com/akiyamalab/NRLMFb.

Multiple grid arrangement improves ligand docking with unknown binding sites: Application to the inverse docking problem.

The identification of comprehensive drug-target interactions is important in drug discovery. Although numerous computational methods have been developed over the years, a gold standard technique has not been established. Computational ligand docking and structure-based drug design allow researchers to predict the binding affinity between a compound and a target protein, and thus, they are often used to virtually screen compound libraries. In addition, docking techniques have also been applied to the virtual screening of target proteins (inverse docking) to predict target proteins of a drug candidate. Nevertheless, a more accurate docking method is currently required. In this study, we proposed a method in which a predicted ligand-binding site is covered by multiple grids, termed multiple grid arrangement. Notably, multiple grid arrangement facilitates the conformational search for a grid-based ligand docking software and can be applied to the state-of-the-art commercial docking software Glide (Schrödinger, LLC). We validated the proposed method by re-docking with the Astex diverse benchmark dataset and blind binding site situations, which improved the correct prediction rate of the top scoring docking pose from 27.1% to 34.1%; however, only a slight improvement in target prediction accuracy was observed with inverse docking scenarios. These findings highlight the limitations and challenges of current scoring functions and the need for more accurate docking methods. The proposed multiple grid arrangement method was implemented in Glide by modifying a cross-docking script for Glide, xglide.py. The script of our method is freely available online at http://www.bi.cs.titech.ac.jp/mga_glide/.

Genetic Nature of Elemental Contents in Wheat Grains and Its Genomic Prediction: Toward the Effective Use of Wheat Landraces from Afghanistan.

Profiling elemental contents in wheat grains and clarifying the underlying genetic systems are important for the breeding of biofortified crops. Our objective was to evaluate the genetic potential of 269 Afghan wheat landraces for increasing elemental contents in wheat cultivars. The contents of three major (Mg, K, and P) and three minor (Mn, Fe, and Zn) elements in wheat grains were measured by energy dispersive X-ray fluorescence spectrometry. Large variations in elemental contents were observed among landraces. Marker-based heritability estimates were low to moderate, suggesting that the elemental contents are complex quantitative traits. Genetic correlations between two locations (Japan and Afghanistan) and among the six elements were estimated using a multi-response Bayesian linear mixed model. Low-to-moderate genetic correlations were observed among major elements and among minor elements respectively, but not between major and minor elements. A single-response genome-wide association study detected only one significant marker, which was associated with Zn, suggesting it will be difficult to increase the elemental contents of wheat by conventional marker-assisted selection. Genomic predictions for major elemental contents were moderately or highly accurate, whereas those for minor elements were mostly low or moderate. Our results indicate genomic selection may be useful for the genetic improvement of elemental contents in wheat.

Genome wide association mapping of stripe rust resistance in Afghan wheat landraces.

Mining of new genetic resources is of paramount importance to combat the alarming spread of stripe rust disease and breakdown of major resistance genes in wheat. We conducted a genome wide association study on 352 un-utilized Afghan wheat landraces against stripe rust resistance in eight locations. High level of disease variation was observed among locations and a core-set of germplasm showed consistence performance. Linkage disequilibrium (LD) decayed rapidly (R2≈0.16 at 0cM) due to germplasm peerless diversity. The mixed linear model resulted in ten marker-trait associations (MTAs) across all environments representing five QTL. The extensively short LD blocks required us to repeat the analysis with less diverse subset of 220 landraces in which R2 decayed below 0.2 at 0.3cM. The subset GWAS resulted in 36 MTAs clustered in nine QTL. The subset analysis validated three QTL previously detected in the full list analysis. Overall, the study revealed that stripe rust epidemics in the geographical origin of this germplasm through time have permitted for selecting novel resistance loci.

Analysis of grain elements and identification of best genotypes for Fe and P in Afghan wheat landraces.

This study was carried out with the aim of developing the methodology to determine elemental composition in wheat and identify the best germplasm for further research. Orphan and genetically diverse Afghan wheat landraces were chosen and EDXRF was used to measure the content of some of the elements to establish elemental composition in grains of 266 landraces using 10 reference lines. Four elements, K, Mg, P, and Fe, were measured by standardizing sample preparation. The results of hierarchical cluster analysis using elemental composition data sets indicated that the Fe content has an opposite pattern to the other elements, especially that of K. By systematic analysis the best wheat germplasms for P content and Fe content were identified. In order to compare the sensitivity of EDXRF, the ICP method was also used and the similar results obtained confirmed the EDXRF methodology. The sampling method for measurement using EDXRF was optimized resulting in high-throughput profiling of elemental composition in wheat grains at low cost. Using this method, we have characterized the Afghan wheat landraces and isolated the best genotypes that have high-elemental content and have the potential to be used in crop improvement.

Development of pre-breeding technology for root system study and selection of Kihara Afghan wheat landraces (KAWLR) to enhance wheat breeding in the rain-fed region.

To enhance a root trait-based selection program for rain-fed wheat breeding in Afghanistan, we simulated an efficient pre-breeding drought system. Plants were grown in 1 m pipes as control or 2 m pipes to simulate drought conditions soaking ground water up by capillary action supplemented by two different life supporting irrigations from top of the pipes (T1 and T2 droughts). T1 was used for studying genetic diversity in 360 Kihara Afghan wheat landraces (KAWLR). Both drought treatments were used to evaluate root traits in 30 selected genotypes. KAWLR showed large root length variations under T1, categorized as long root (>200 cm; LR), medium root (100-150 cm; MR) and short root (20-100 cm; SR) systems. LR genotypes were more drought resistant in terms of greater plant survivability under T1 and T2 compared with other groups and were capable of adjusting their root biomass partitioning at deepest part of the soil profile. Majority of the LR genotypes originated from predominantly rain-fed provinces, and most of their agronomic traits were strongly correlated with root biomass deep in the soil in response to drought. Three LR genotypes, including the longest root genotype LR-871 (KU7604), are recommended for rain-fed wheat breeding in Afghanistan.

Identification of potential inhibitors based on compound proposal contest: Tyrosine-protein kinase Yes as a target.

A search of broader range of chemical space is important for drug discovery. Different methods of computer-aided drug discovery (CADD) are known to propose compounds in different chemical spaces as hit molecules for the same target protein. This study aimed at using multiple CADD methods through open innovation to achieve a level of hit molecule diversity that is not achievable with any particular single method. We held a compound proposal contest, in which multiple research groups participated and predicted inhibitors of tyrosine-protein kinase Yes. This showed whether collective knowledge based on individual approaches helped to obtain hit compounds from a broad range of chemical space and whether the contest-based approach was effective.

Genome-wide transcriptional profiling of wheat infected with Fusarium graminearum.

Fusarium head blight (FHB) is a destructive disease in wheat caused by Fusarium graminearum (F. g). It infects during the flowering stage favored by warm and highly humid climates. In order to understand possible wheat defense mechanism, gene expression analysis in response to F. g was undertaken in three genotypes of wheat, Japanese landrace cultivar Nobeokabouzu (highly resistant), Chinese cv. Sumai 3 (resistant) and Australian cv. Gamenya (susceptible). For microarray analysis, 3 and 7 days after inoculation (dai) samples were used in Agilent wheat custom array 4x38k. At 3 dai, the highest number of genes was up-regulated in Nobeokabouzu followed by Sumai 3 and minimum expression in Gamenya. Whereas at 7 dai, Sumai 3 expressed more genes compared to others. Further narrowing down by excluding commonly expressed genes in three genotypes and grouping according to the gene function has identified differentially high expression of genes involved in detoxification process such as multidrug resistant protein, multidrug resistance-associated protein, UDP-glycosyltransferase and ABC transporters in Nobeokabouzu at 3 dai. However in Sumai 3 many defense-related genes such as peroxidase, proteases and genes involved in plant cell wall defense at 7 dai were identified. These findings showed the difference of molecular defense mechanism among the cultivars in response to the pathogen. The complete data was accessed in NCBI GEO database with accession number GSE59721.

Exploiting genetic diversity from landraces in wheat breeding for adaptation to climate change.

Climate change has generated unpredictability in the timing and amount of rain, as well as extreme heat and cold spells that have affected grain yields worldwide and threaten food security. Sources of specific adaptation related to drought and heat, as well as associated breeding of genetic traits, will contribute to maintaining grain yields in dry and warm years. Increased crop photosynthesis and biomass have been achieved particularly through disease resistance and healthy leaves. Similarly, sources of drought and heat adaptation through extended photosynthesis and increased biomass would also greatly benefit crop improvement. Wheat landraces have been cultivated for thousands of years under the most extreme environmental conditions. They have also been cultivated in lower input farming systems for which adaptation traits, particularly those that increase the duration of photosynthesis, have been conserved. Landraces are a valuable source of genetic diversity and specific adaptation to local environmental conditions according to their place of origin. Evidence supports the hypothesis that landraces can provide sources of increased biomass and thousand kernel weight, both important traits for adaptation to tolerate drought and heat. Evaluation of wheat landraces stored in gene banks with highly beneficial untapped diversity and sources of stress adaptation, once characterized, should also be used for wheat improvement. Unified development of databases and promotion of data sharing among physiologists, pathologists, wheat quality scientists, national programmes, and breeders will greatly benefit wheat improvement for adaptation to climate change worldwide.

Altered gene expression profiles of wheat genotypes against Fusarium head blight.

Fusarium graminearum is responsible for Fusarium head blight (FHB), which is a destructive disease of wheat that makes its quality unsuitable for end use. To understand the temporal molecular response against this pathogen, microarray gene expression analysis was carried out at two time points on three wheat genotypes, the spikes of which were infected by Fusarium graminearum. The greatest number of genes was upregulated in Nobeokabouzu-komugi followed by Sumai 3, whereas the minimum expression in Gamenya was at three days after inoculation (dai). In Nobeokabouzu-komugi, high expression of detoxification genes, such as multidrug-resistant protein, multidrug resistance-associated protein, UDP-glycosyltransferase and ABC transporters, in addition to systemic defense-related genes, were identified at the early stage of infection. This early response of the highly-resistant genotype implies a different resistance response from the other resistant genotype, Sumai 3, primarily containing local defense-related genes, such as cell wall defense genes. In Gamenya, the expression of all three functional groups was minimal. The differences in these molecular responses with respect to the time points confirmed the variation in the genotypes. For the first time, we report the nature of gene expression in the FHB-highly resistant cv. Nobeokabouzu-komugi during the disease establishment stage and the possible underlying molecular response.

Molecular evaluation of orphan Afghan common wheat (Triticum aestivum L.) landraces collected by Dr. Kihara using single nucleotide polymorphic markers.

BACKGROUND: Landraces are an important source of genetic diversity in common wheat, but archival collections of Afghan wheat landraces remain poorly characterised. The recent development of array based marker systems, particularly single nucleotide polymorphism (SNP) markers, provide an excellent tool for examining the genetic diversity of local populations. Here we used SNP analysis to demonstrate the importance of Afghan wheat landraces and found tremendous genetic diversity and province-specific characteristics unique to this geographic region. RESULTS: A total of 446 Afghan wheat landraces were analysed using genotype by sequencing (GBS) arrays containing ~10 K unique markers. Pair-wise genetic distance analyses revealed significant genetic distances between landraces, particularly among those collected from distanced provinces. From these analyses, we were able to divide the landraces into 14 major classes, with the greatest degree of diversity evident among landraces isolated from Badakhshan province. Population-based analyses revealed an additional 15 sub-populations within our germplasm, and significant correlations were evident in both the provincial and botanical varieties. Genetic distance analysis was used to identify differences among provinces, with the strongest correlations seen between landraces from Herat and Ghor province, followed closely by those between Badakhshan and Takhar provinces. This result closely resembles existing agro-climatic zones within Afghanistan, as well as the wheat varieties commonly cultivated within these regions. Molecular variance analysis showed a higher proportion of intra-province variation among landraces compared with variation among all landraces as a whole. CONCLUSION: The SNP analyses presented here highlight the importance and genetic diversity of Afghan wheat landraces. Furthermore, these data strongly refute a previous analysis that suggested low genetic diverse within this germplasm. Ongoing analyses include phenotypic characterisation of these landraces to identify functional traits associated with individual genotypes. Taken together, these analyses can be used to help improve wheat cultivation in Afghanistan, while providing insights into the evolution and selective pressures underlying these distinct landraces.

Variations for Fusarium head blight resistance associated with genomic diversity in different sources of the resistant wheat cultivar 'Sumai 3'.

Fusarium head blight (FHB), caused by Fusarium graminearum, is a serious disease of wheat (Triticum aestivum L.) associated with contamination by the mycotoxin deoxynivalenol (DON). The FHB-resistant wheat cultivar 'Sumai 3' has been used extensively around the world. The existence of variation in FHB resistance among 'Sumai 3' accessions has been discussed. In this study, genetic variation among 'Sumai 3' accessions collected from six countries were identified using SSR markers; our results demonstrate unique chromosome regions in Sumai 3-AUT and Sumai 3-JPN ('Sumai 3' accessions from Austria and Japan, respectively). Field evaluation indicated strong resistance to FHB in Sumai 3-AUT. The polymorphic rate (number of polymorphic markers/number of available markers × 100) based on a DArT array was 12.5% between the two 'Sumai 3' accessions. Genotyping for DNA markers flanking FHB-resistant quantitative trait loci (QTLs) revealed genetic variations for the QTL regions on 5AS and 2DS; however, no variation was observed for the QTL regions on 3BS and 6B. Thus, the variation in FHB resistance among 'Sumai 3' accessions in the field is due to genetic diversity.

Development of Capsicum EST-SSR markers for species identification and in silico mapping onto the tomato genome sequence.

Capsicum spp. are widely cultivated for use as vegetables and spices. The Kihara Institute for Biological Research, Yokohama City University, Japan, has stocks of approximately 800 lines of Capsicum spp. collected from various regions of Central and South America, the regions of origin for Capsicum spp. In this study, 5,751 primer pairs for simple sequence repeat markers, based on 118,060 publicly available sequences of expressed sequence tags of Capsicum annuum, were designed and subjected to a similarity search against the genomic sequence of tomato, a model Solanaceae species. Nucleotide sequences spanning 2,245 C. annuum markers were successfully mapped onto the tomato genome, and 96 of these, which spanned the entire tomato genome, were selected for further analysis. In genotyping analysis, 60 out of the 77 markers that produced specific DNA amplicons showed polymorphism among the Capsicum lines examined. On the basis of the resulting data, the 192 tested lines were grouped into five main clusters. The additional sequencing analysis of the plastid genes, matK and rbcL, divided the resources into three groups. As a result, 19 marker loci exhibited genotypes specific to species and cluster, suggesting that the DNA markers are useful for species identification. Information on the DNA markers will contribute to Capsicum genetics, genomics, and breeding. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11032-012-9774-z) contains supplementary material, which is available to authorized users.

Genetic relationships between resistances to Fusarium head blight and crown rot in bread wheat (Triticum aestivum L.).

Fusarium head blight (FHB) and crown rot (CR) are two wheat diseases caused by the same Fusarium pathogens. Progress towards CR resistance could benefit from FHB-resistant germplasm if the same genes are involved in resistance to these two different diseases. Two independent studies were conducted to investigate the relationship between host resistances to these two diseases. In the first study 32 genotypes were assessed and no significant correlation between their reactions to FHB and CR was detected. The second study was based on a QTL analysis of a doubled haploid population derived from a variety with resistance to both diseases. Results from this study showed that loci conferring resistance to FHB and CR are located on different chromosomes. Together, these results suggest that, despite a common aetiology, different host genes are involved in the resistance against FHB and CR in wheat. Thus, although it is possible that genes affecting both diseases may exist in other germplasm or under different conditions, separate screening seems to be needed in identifying sources of CR resistance.

Deoxynivalenol Distribution in Flour and Bran of Spring Wheat Lines with Different Levels of Fusarium Head Blight Resistance.

The level of the fungal toxin deoxynivalenol (DON) in milled flour and its partitioning between flour and bran of 15 spring wheat (Triticum aestivum) lines with different levels of Fusarium head blight (FHB; mainly caused by Fusarium graminearum) resistance was investigated in Hokkaido, Japan between 2004 and 2007. The DON levels in flour (DONf) and bran (DONb) showed positive linear relationships with DON levels in whole kernels (DONk) (R2 = 0.94, n = 60, P ≤ 0.01 and R2 = 0.93, n = 60, P ≤ 0.01, respectively). Results indicated that the DON partitioning ratio between flour and bran is primarily related to the level of DONk, and the DON level was approximately doubled in DONb and halved in DONf for any level of DONk. Only one pattern of DON partitioning supported the conventional strategy to select FHB-resistant lines.

QTL-specific microarray gene expression analysis of wheat resistance to Fusarium head blight in Sumai-3 and two susceptible NILs.

Fusarium head blight, predominantly caused by Fusarium graminearum (Schwabe) in North America, is a destructive disease that poses a serious threat to wheat (Triticum aestivum L.) production around the world. cDNA microarrays consisting of wheat ESTs derived from a wheat - F. graminearum interaction suppressive subtractive hybridization library were used to investigate QTL-specific differential gene expression between the resistant Chinese cultivar Sumai-3 and two susceptible near isogenic lines (NILs) following inoculation with F. graminearum. Stringent conditions were employed to reduce the false discovery rate. A total of 25 wheat unigenes were found to express differentially in response to F. graminearum infection. Genes encoding pathogenesis-related (PR) proteins such as beta-1,3-glucanase (PR-2), wheatwins (PR-4), and thaumatin-like proteins (PR-5) showed a significant upregulation in genotypes having the Sumai-3 3BS region. For these three genes, the gene activity was significantly less in the genotype (NIL-3) lacking the Sumai-3 3BS segment. Significant upregulation of phenylalanine ammonia-lyase was detected only in the resistant Sumai-3, indicating the importance of both the 2AL and 3BS regions in the activation of effective defense responses to infection by F. graminearum. Differences in gene expression between the resistant Sumai-3 and the susceptible NILs were found to be mainly quantitative in nature.

Marker-assisted characterization of Asian wheat lines for resistance to Fusarium head blight.

The major quantitative trait locus (QTL) on 3BS from Sumai 3 and its derivatives has been used as a major source of resistance to Fusarium head blight (FHB) worldwide, but resistance genes from other sources are necessary to avoid complete dependence on a single source of resistance. Fifty-nine Asian wheat landraces and cultivars differing in the levels of FHB resistance were evaluated for type II FHB resistance and for genetic diversity on the basis of amplified fragment length polymorphism (AFLP) and simple sequence repeats (SSRs). Genetic relationships among these wheat accessions estimated by cluster analysis of molecular marker data were consistent with their geographic distribution and pedigrees. Chinese resistant landraces had broader genetic diversity than that of accessions from southwestern Japan. The haplotype pattern of the SSR markers that linked to FHB resistance quantitative trait loci (QTLs) on chromosomes 3BS, 5AS and 6BS of Sumai 3 suggested that only a few lines derived from Sumai 3 may carry all the putative QTLs from Sumai 3. About half of the accessions might have one or two FHB resistance QTLs from Sumai 3. Some accessions with a high level of resistance, may carry different FHB resistance loci or alleles from those in Sumai 3, and are worth further investigation. SSR data also clearly suggested that FHB resistance QTLs on 3BS, 5AS, and 6BS of Sumai 3 were derived from Chinese landrace Taiwan Xiaomai.