Department of Plant Pathology, University of Minnesota, USA
Plant pathogens elicit host transcriptome changes that facilitate hyphal infection and proliferation throughout host tissues. Host genes targeted by the pathogen to facilitate infection have been identified as susceptibility genes (S-genes). As biotrophic pathogens, rust fungi must infect the host to obtain nutrients while suppressing defense responses and likely target host-derived genes to aid in this process. To further understand the host-pathogen interaction RNA-seq analyses were performed on three compatible host-rust pathogen systems: barley cv. Morex was inoculated with Pgt race TTKSK (Ug99), soybean cv. Williams 82 was inoculated with a field collection of Phakopsora pachyrhizi, and maize cv. B73 was inoculated with an isolate of P. sorghi collected at St. Paul. Differential expression was characterized at 6, 12, 24, and 72 h post inoculation. Homologous and differentially expressed host genes were identified and compared for similarities in patterns of differential expression. In each system, a transmembrane nitrate transporter was identified as significantly upregulated early in the interaction compared to mock inoculated controls. Transporters with this molecular function were previously correlated with altering the host-pathogen interaction by knockdown and knockout mutants that were associated with resistance and heightened expression was associated with enhanced susceptibility. The transporter identified in barley was chosen for analysis to understand its effect on the host-pathogen interaction. Current objectives include site directed mutagenesis utilizing multiplexed CRISPR/Cas9 constructs to induce knockout host alleles.
National Institute of Agricultural Research, INRA, Morocco
Bread wheat is the most important staple crop in Morocco. However, the low national production represents less than 50% of national need. Yellow (stripe) rust (Pst) and Septoria tritici blotch (STB) are the most important foliar diseases limiting bread wheat production in Morocco. The objective of this study was to identify effective sources of resistance to Pst and STB and to select candidate lines displaying high yield and resistance traits. A total of 194 bread wheat accessions provided by ICARDA and CIMMYT were evaluated during 2014 for grain yield and reaction to STB and yellow rust. The field trials were carried out at Douyet station, a key Moroccan rainfed location, and were arranged in a replicated complete block design of 3 replicates. For rust, each entry was scored using the modified Cobb scale when the susceptible check had reached 60S. The Saari Prescott scale was used to score STB reactions. The crop season was characterized by early and late drought associated with high temperatures after flowering. About 50% of accessions (99) showed high yields ranging from 1% to 55% above the highest national check, Arrehane (1.86 t/ha). About 48% (93 accessions) were highly resistant to moderately resistant to yellow rust and about 84% (164 accessions) were resistant to moderately resistant to STB. Overall, about 30% (58 lines) of accessions were high yielding with final disease severities ranging from 0-10% for yellow rust and from resistant to moderately resistant for STB. This group incorporates potentially valuable traits beneficial for our wheat breeding program. QUAIU*2/KINDE was the most promising accession with the highest grain yield (2.9 t/ha) combined with foliar disease resistance. Further multi-environment screening and field evaluations are required before recommending accessions for potential release.
National Institute of Agricultural Research (INIA), La Estanzuela Experimental Station, Uruguay
Breeding for durable leaf rust resistance is a priority for our breeding programs; however, the availability of new resistance genes is a limiting factor. Two spring wheat populations totaling 186 lines derived from three resistant donors and two Uruguayan susceptible cultivars were used to detect genomic regions associated with seedling and field resistance to LR in Uruguay. AUDPC were recorded in three environments in the 2012 and 2013 cropping seasons, and seedling responses were determined using three Puccinia triticina races. The lines were also genotyped using GBS. A total of 5,222 SNP markers were used for genome-wide association analysis. Molecular markers were used to genotype APR genes Lr34 and Lr68. We identified 43 SNP markers significantly associated with seedling resistance and 19 for field resistance on chromosomes 1A, 1B, 1D, 2B, 2D, 3A, 4A, 5B, 6B, 7A, 7B and 7D. We confirmed the presence of Lr10 and Lr16 in seedling tests and Lr34 and Lr68 in field tests. Novel genomic regions were identified on chromosomes 4A associated with APR, and 5B associated with seedling resistance. These new resistance genes will be useful in breeding for durable LR resistance.
OR Melhoramento de Sementes Ltda., Brazil
Wheat is the best cropping option for the winter season in southern Brazil. According to current predictions the wheat area in 2015 will decline by 5.3% (146 thousand hectares) due to low prices for cereal grain and to high precipitation and disease severities (principally FHB and wheat blast) in 2014 that caused high losses in production. Due to high inoculum pressure leaf rust is controlled every year by resistance genes and fungicide application (an average three applications during the growing season to control all wheat diseases). Of 119 cultivars possibly cultivated in Brazil, 58 are moderately susceptible or susceptible to leaf rust. The objective of this study was to survey the Puccinia triticina population in order to identify the predominant races in Brazil. Infected leaves were collected from wheat crops in different regions in 2014. After isolating pustules and increasing the inoculum, the Thatcher differential lines [Lr1 2a 2c 3a (set 1), Lr9 16 24 26 (set 2), Lr3ka 11 17 30 (set 3), Lr10 18 21 23 (set 4), Lr14a 14b 10+26 20 (set 5), Lr3bg 27+31 and line ORL04002 - Toropi/Ônix) were inoculated. From 186 field samples, 74 isolates were evaluated. Race T(DF)T-MT, first identified in 2007, was predominant in 2013 and 2014. The frequency of race TDP-M(RT), first identified in 2008, increased to second position in 2014. Stripe rust and stem rust are not common diseases in Brazil; however, samples of both diseases were collected in 2013, but not in 2014. Most of the Brazilian OR Sementes lines evaluated in Argentina in 2014 stood out by resistance to stem rust, indicating the importance of incorporating resistance to all three rusts in Brazilian cultivars.
Indian Council of Agricultural Research
Leaf rust of wheat causes considerable losses worldwide. New pathotypes may cause previously resistant varieties to become susceptible. Identification of pathotypes and their relationships provide information for breeding efforts and designing management strategies. Traditional identification of pathotypes is based on responses of differential hosts. At present 50 pathotypes of P. triticina are maintained in the National collection. To determine variability and relationships at the molecular level we conducted analyses with 26 SSR primers, eight of which were polymorphic. Binary (0 or 1) molecular data generated by NTSYS-pc was used to construct a phylogenetic tree. Cluster analysis was done using the unweighted pair group arithmetic means (UPGMA) method in the SAHN program of NTSYS-pc. Pathotype groups and subgroups were determined based on the Jaccard similarity coefficients (JC). Manual observations indicated seven major groups. Among them, two groups each have one pathotype (pathotypes 16 and 17). Jaccard similarity coefficients supported groupings based on pathogenicity data. For example, pathotypes in the race 12 group (12, 12-1, 12-3, 12-4, 12-6, 12-7, 12-8, but excluding 12-2 and 12-5) had similarity coefficients greater than 0.7. Similar observations were recorded for the race 77 group. Maximum similarity was observed between 12-3 and 12-7 (JC value: 0.89) followed by 12-3, 12-7 and 12-6 (JC value: 0.82). Based on the phylogenetic tree and similarity coefficients data, there was substantial diversity among pathotypes. Thus SSR marker data can be used for effective characterization of pathotypes and for making evolutionary inferences.
Wheat rust is a group of deadly, constantly changing fungal pathogens that pose a serious threat worldwide and also equally most important in Pakistan. Three participatory wheat seed value chain workshops conducted in Pakistan in 2014 indicated the predominance of informal seed systems in Khyber Pakhtunkhwa and Pothwar region of Punjab. A relatively old and rust susceptible variety Sahar06 was covering nearly 50% area of irrigated wheat in Punjab while the coverage of TD-1 was nearly 75 % of wheat area in Sindh. Predominance of mega susceptible varieties with informal seed systems can be an important recipe for the outbreak of rust diseases threatening the stability of wheat production systems in Pakistan. It is challenging to continue to improve and sustain wheat productivity by reducing vulnerability of wheat varieties to rust diseases, both in time and space when more than 70% of wheat seed used comes from farmers' own farm saved seeds mostly of old and obsolete varieties. To address this, 17 recently released rust resistant wheat varieties with diversity in genetic background, adaptation and good yield potential have been deployed through a network of partners to nearly 10,000 smallholder farmers in parts of 62 districts of Pakistan. Varietal deployment was done by coordinating on farm participatory varietal selection with agronomic interventions and village level seed multiplication and marketing. Though, rusts pathogens can evolve into new strains that are more virulent and damaging to wheat crops, nevertheless, some of these varieties do carry known genes conferring resistance to yellow, leaf or stem rusts thereby help avert any sudden rust epidemics. This effort will be instrumental in improving the access to new seed varieties in the grassroots level and widening the genetic bases of wheat that will help in buffering the rust incidence and contribute to household food security of smallholder farmers in Pakistan.
Seed-Co Ltd., Rattray Arnold Research Station, Zimbabwe
Four Ug99 pathotypes occur in southern Africa. Although South African bread wheat cultivars and lines are regularly screened against representative isolates, the stem rust reactions of Zimbabwean germplasm to these variants were largely unknown. A collection of 49 wheat cultivars and lines, obtained from Seed-Co (Ltd.) and the Crop Breeding Institute in Zimbabwe, were tested as seedlings against pathotypes TTKSF, TTKSF+, TTKSP and PTKST. Twelve varieties and 21 experimental lines showed low infection types with all four pathotypes. Using molecular marker assays Sr31 was detected in 26 entries, Sr24 in five and Sr36 in one. The csSr2 marker suggested the presence of Sr2 in 20 entries. Screening of adult plants in the greenhouse using pathotype PTKST showed 34 entries with low infection types and 15 had high infection types. Stem rust field records in 2012 showed 5 susceptible entries with stem rust scores between 50S and 80S, whereas only 4 susceptible entries were identified in 2014 with scores ranging from 30S to 80S. Three lines were susceptible in both seasons. The study exposed the vulnerability of Zimbabwean wheat germplasm to Ug99 variants, but also identified suitable lines that can be used in breeding and possible commercialization.
Department of Molecular, Cellular and Biomedical Sciences, University of New Hampshire, USA
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In the northeastern United States, outside the boundaries of the 20th century federal barberry eradication zone, both common barberry (Berberis vulgaris) and Japanese barberry (B. thunbergii) are found in great abundance, to the extent that both are considered invasive species. Much less common and relatively less studied is their interspecific hybrid, B. ×ottawensis, which has been produced in the ornamental horticultural industry but which also occurs naturally. Since B. vulgaris is a competent host of Puccinia graminis and B. thunbergii is not, B. ×ottawensis presents a unique system for characterizing the genetic mechanism(s) underlying what appears to be non-host resistance to P. graminis in B. thunbergii. In this study, a natural population of about 1,000 individuals (mixed B. vulgaris, B. thunbergii, and B. ×ottawensis) in Sheffield, MA, was investigated. While wide morphological variation was observed among and within the populations of all three species at the site, the most pronounced variation was observed among B. ×ottawensis individuals. A subset of the population was selected for genotyping by sequencing (GBS) and evaluated for reaction to P. graminis via controlled inoculations. The response was found to segregate clearly among B. ×ottawensis individuals; and GBS was shown to be a viable means of generating molecular markers in these species, despite the lack of a reference genome. These results suggest that P. graminis resistance in B. thunbergii can be genetically mapped, and mapping populations are currently under development to accomplish this goal. The genomic resources developed in this work may facilitate both barberry surveillance efforts and ornamental barberry testing programs. Furthermore, knowledge of the genetics of response to P. graminis in the alternate host has the potential to inform efforts in breeding for stem rust resistance in wheat.
National Institute of Agricultural Technology (INTA), Argentina
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Argentina and Uruguay are neighboring countries located in the same rust epidemiological area. The last significant stem rust epidemic occurred in 1950. Since then, stem rust was frequently observed in experimental fields and off-season nurseries, but was mostly absent in commercial fields. During 2014, 4.6 million ha of wheat were grown, and there was a widespread incidence of stem rust, reaching levels of 80S on susceptible cultivars in both countries. Yield losses of 13 to 21% were estimated in experimental trials in Argentina. The epidemic was probably caused by the increasingly widespread cultivation of highly susceptible, but high yielding French cultivars during the last decade. In Argentina 42.3% of the commercial cultivars were susceptible to stem rust, and in Uruguay 23.0% were susceptible, 6.8% moderately susceptible and 20.3% were intermediate in reaction. However, the actual area sown to susceptible cultivars in Uruguay has continued to increase, from 22% of the wheat area in 2009 to 53.3% in 2014. Conductive weather conditions of high rainfall and warmer than average temperatures during the winter and spring, favored early infection. Cultivars with resistance genes Sr31 and Sr24 continue to be resistant in the region and are believed to be the most important genes currently providing resistance. Some Argentinean and Uruguayan cultivars that do not carry Sr31 and/or Sr24 were susceptible in 2011, but resistant in 2014, indicating a narrower range of virulence in 2014 compared to 2011. Both countries are working to improve resistance to local races and to the Ug99 race group. Disease modeling would be useful for understanding and predicting the occurrence and severity of this disease.
Department of Agronomy and Plant Genetics, University of Minnesota, USA
Leaf rust is common in wheat worldwide, consistently reducing yields by 5-15% or more. Fungicides are applied to U.S. spring wheat to mitigate crop losses, but combinations of resistance genes can provide less expensive, effective control. We use biparental and association mapping approaches to identify resistance genes in two Minnesota varieties and 3,000 diverse global lines. Six populations developed from selected National Small Grains Collection lines were evaluated for seedling resistance, which has been mapped using bulk segregant analysis. Two populations were developed to map adult plant resistance using recombinant inbred lines. Leveraging large association mapping panels can identify novel variants from the diverse NSGC lines. The results will identify successful gene combinations underlying durable resistance and associated genetic markers. We will explore the utility of association mapping to identify leaf rust resistance genes and evaluate the global distribution of leaf rust resistance to inform worldwide wheat improvement.