Wheat stripe (Puccinia striiformis f. sp. tritici,=Pst) and stem (Puccinia graminis f. sp. tritici =Pgt) rusts are the most important wheat disease in Egypt as well as present in all wheat growing areas. This study to evaluate a set of tester lines of wheat carrying stripe Yr's, stem Sr's rust genes and selected Egyptian varieties have been studied for their response to Pst and Pgt at adult plant stage under field conditions in Sakha Agriculture Research Station, during the 2011 to 2014 growing seasons. The results revealed that stripe rust, it has been observed that the new race Yr27-virulence to Pst. In addition pathotypes were virulent for Yr2, Yr6, Yr7, Yr8, Yr9, Yr27, while Yr18 showed moderate susceptibility. On the other hand, Yr1, Yr5, Yr10, Yr15, Yr17, Yr32 and YrSP exhibited high levels of resistance. Regarding, evaluation of resistance genes sources of stem rust on ICARDA, CIMMYT wheat germplasm, and Egyptian wheat varieties released i.e. Misr1 and Misr2 which having Ug99_resistance genes Sr2 and Sr25 were found susceptible to Pgt, also Sr31 recorded infection moderately susceptible to susceptible at adult stage. Genes Sr2 complex, Sr24, Sr26, Sr27, and Sr32 were resistant at adult plant stages. The combination of Sr26 with Sr2 and Sr25 provided stem rust resistance in some CIMMYT wheat germplasm. The objectives of this work are: race analysis of wheat stem and stripe rust disease, evaluation the level and distribution of wheat stripe and stem rust in Egypt, and identification the resistance genes in commercial varieties or new promising lines using standard and molecular genetic markers. Egyptian germplasm such as Misr1, and Misr2 and others tester lines of wheat carrying stem rust Sr's were evaluative under field condition at adult stage in Egypt during 2014 growing season, Egyptian cultivars Misr1 and Misr2 were susceptible rated 10S-20S and Sr31 rated MSS. that results clearly presence a new Sr31-virulence. On other hand, genes Sr2 complex, Sr24, Sr26, Sr27 and Sr32 were resistant and combination of Sr26 with (Sr2 and Sr25) produced stem rust resistance in some CIMMYT wheat germplasm. Shahin et al., 2015, in APS Annual Meeting, Aug. 1-5, Pasadena, CA, US, (In Press).
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Stem rust caused by Puccinia graminis f. sp. tritici (Pgt) is a destructive disease on bread and durum wheat. Following the identification and distribution of Ug99, major national and international efforts have been made to detect additional spread and emergence of new Pgt races. Since 2011, GRRC has accepted to receive live samples of stem rust year round, and up to 2014, a total of 428 dried samples of Pgt infected wheat tissue were received from 15 African and Asian countries, i.e., Azerbaijan, Egypt, Ethiopia, Iran, Iraq, Kenya, Lebanon, Nepal, Rwanda, Sudan, Tanzania, Turkey, Uganda, Yemen and Zimbabwe. Additional samples were received from Germany, Sweden and Denmark, where wheat stem rust re-emerged in 2013-2014. Recovery procedures using susceptible seedlings of cv. Morocco was done upon arrival and a total of 269 samples were successfully recovered, multiplied and stored in liquid nitrogen until further use. To date, 140 Pgt isolates have been pathotyped based on the method of Jin et al. (2008). Subsets of isolates were selected for molecular characterization including SNP genotyping and shipped to USDA-ARS, Cereal Disease Lab (CDL). The Pgt race TKTTF was widely distributed and found in ten countries including Egypt, Ethiopia, Iran, Iraq, Lebanon, Sudan, Turkey and the three European countries. Races of the Ug99 lineage were frequently observed in Africa. Clear indication of a new race in the Ug99 race group with additional virulence for SrTmp, TTKTK, was observed in samples from four African countries in 2014. PCR diagnostics developed by CDL confirmed the new race being member of the Ug99-lineage. The experimental work was supported by the DRRW project and new research facilities were funded by Aarhus University.
Use of large-scale computational resources has permitted the first quantitative study of airborne migration routes of fungal spores between numerous key epidemiological hot-spots of wheat stem rust in Africa, the Middle East and the Indian subcontinent. By coupling a state-of-the-art Lagrangian particle dispersion model (NAME) with mechanistic epidemiological models, we simulate turbulent atmospheric transport of large ensembles of fungal spores from source sites. The models use highly resolved global meteorological datasets from the UK Meteorological Office. We consider release of P. graminis uredinospores from numerous source locations over an 11 year period (2003-2014) and simulate atmospheric trajectories over a 10 km2 spatial sampling grid to elucidate spore deposition rates at national, regional, and continental spatial scales. Our systematic exploration permits the first quantitative perspective and ranking of likely airborne transmission routes of wheat stem rust. We identify migration trends within and between the “Rift valley epidemiological zone”, the Middle East, the Indian Subcontinent, as well as South Africa. Our results indicate (I) consistent seasonal dispersal patterns, (II) likely airborne transmission of stem rust from the Middle East to North-East Africa, and (III) suggest that there is considerable risk of spread of Ug99 or other virulent races from Eastern Yemen to the Indian subcontinent. Model results indicate that over the 11 year study period, viable spore deposition occurred between Eastern Yemen and Pakistan on average 22 days per year during overlapping wheat growing seasons. The validity of the modelling framework has been successfully tested by comparison with survey data from the 2013 epidemic outbreak in Ethiopia, and was recently used as a risk assessment tool to provide rapid response advice in different East-African countries. Known stem rust race distributions are also supportive of the model outputs. The research we have been doing allows a quantitative perspective on likely airborne transmission routes of Ug99 or other virulent races of wheat rust. By that we hope to provide new insights and recommendations for future risk assessment, survey and control strategies and also to contribute to fundamental understanding of epidemiological spread on regional and continental scales. The work we would like to present is the result of a joint effort of Dr Laura Burgin and Dr Matt Hort from the UK Meteorological office, Dr Dave Hodson from CIMMYT, and Dr James Cox, Matthew Hitchings and me from the Epidemiology and Modelling group of Prof Gilligan in Cambridge.
Wheat stem rust is one of the major wheat yield limiting factors in Ethiopia. A stem rust epidemic occurred in the wheat belts of Arsi and Bale zones in the 2013-2014 crop season caused by Pgt race TKTTF that is virulent to the widely grown Ug99-resistant variety Digelu. This epidemic highlighted the need for wheat varieties with resistance to multiple Pgt races. This study was therefore, carried out to evaluate the reaction of the major Ethiopian varieties and advanced breeding lines against the dominant Pgt races in Ethiopia. Races TKTTF, TTKSK, TRTTF and JRCQC were isolated from field samples and multiplied on the susceptible cultivar McNair starting in May 2014. Four wheat stem rust nurseries, each inoculated with a single Pgt race, were established at Kulumsa and monitored from July through October, 2014. Each nursery included 34 entries in two replicates and 137 entries in a single replicate, augmented with six sets of five repeating checks. An additional nursery established at Debre Zeit, containing 551 entries in an augmented design, was evaluated with the epidemic Pgt race TKTTF. These entries included the most relevant Ethiopian bread and durum wheat breeding lines and cultivars, and 34 seedling-susceptible lines to evaluate the race-specificity of adult plant resistance. Stem rust severities for the four races ranged from trace to 80 %. Out of all entries evaluated, 10 were resistant to all four Pgt races, while 11 entries were effective to three of the four races. At Debre Zeit, 31.4% of the entries were resistant to Pgt race TKTTF. This study showed that rapid isolation and increase of Pgt races in Ethiopia is possible to facilitate field screening of breeding lines to select for candidate cultivars with resistance to multiple virulent races of Pgt.
Stem rust is a potentially destructive fungal disease of wheat worldwide. In 1998 Pgt pathotype TTKSK virulent to Sr31 was detected in Uganda. The same pathotype was confirmed in Lorestan and Hamedan provinces of Iran in 2007. We used a derivative of race TTKSK to phenotype 62 Iranian wheat landraces (resistant to stripe rust in a previous study) at the seedling stage to this new pathotype (TTSSK). Twenty eight accessions were evaluated for the presence of resistance genes Sr2, Sr22, Sr24, Sr25, Sr26, Sr35, Sr36 and Srweb using SSR markers. None carried Sr2, Sr24 or Sr26, but the presence of Sr22, Sr25, Sr35 and Sr36 was indicated. Some susceptible landraces predicted to carry Sr2 by marker analysis require further investigation. To evaluate defense gene expression in compatible and incompatible stem rust interactions we sampled resistant and susceptible cultivars at 0, 12, 18, 24, 72 hours post-inoculation (hpi). ?-1,3 glucanase expression was studied using qGLU-S and qGLUU-AS primers and a real-time PCR step-one ABI machine, with ?-tubulin and EF1-? genes used as internal controls. In incompatible interactions defense gene expression was increased at 24 hpi, but in compatible interactions the highest level of expression occurred at 12 hpi and was significantly decreased at 18 hpi. The results revealed that expression of defense genes such as ?-1,3 glucanase was earlier in compatible than in incompatible interactions but the expression level was less in incompatible interactions. On the other hand, in susceptible genotypes the expression of defense genes increased immediately after inoculation and declined sharply after infection. In contrast defense gene expression in resistant genotypes began to increase after establishment of the pathogen.
Stem rust caused by Puccinia graminis tritici (Pgt) is one of the most serious diseases in wheat and is combated mainly through the use of resistant varieties. Because the fungus evolves virulence towards previously resistant varieties, continuous breeding and identification of new sources of resistance are necessary to combat the threat of rust epidemics. Our work on the flax rust model system has provided insights into how the plant immune system recognises and responds to rust pathogens. We have been extending this work to wheat stem rust by targeted cloning of resistance (R) genes in wheat and corresponding Avr genes in Pgt. Plant R genes encode immune receptors that recognise and respond to pathogen effector proteins delivered into host cells from haustoria. We recently isolated the Sr33 and Sr50 resistance genes from wheat and have begun functional analyses to determine how they trigger defense responses. We are also targeting effectors from Pgt that are recognised by wheat R genes. We used genome and transcriptome sequencing to predict ~400 candidate effector genes from Australian Pgt race 21- 0. To screen for recognition of these proteins by wheat R genes, we developed a bacterial Type III Secretion System delivery assay using Pseudomonas fluorescens to inject the effector candidates into wheat leaf cells. We are screening candidate effectors on a set of 18 wheat cultivars carrying 22 different R genes and have so far identified one effector that induces a cell death response specifically on a wheat genotype carrying Sr22. Understanding the nature of wheat R genes and the Avr proteins that they recognize will allow better prediction of R gene durability and enable the possibility of rational design of novel R genes. We are also developing techniques for stacking R genes in cassettes for deployment of multiple genes at a single locus in wheat.
Stem rust (SR) resistance is required for CIMMYT durum germplasm to keep relevance in Ethiopia, where Ug99 and other Pgt races are a major yield-limiting constraint, and in countries along the possible dissemination paths of these races. Resistance to Ug99 is widespread in most durum germplasm groups when tested in Kenya, but resistance is lost when exposed to Ethiopian races; hence selection at the Debre Zeit site in Ethiopia is essential for durum wheat. Due to difficulties with shuttling segregating populations between Mexico and Ethiopia, we have adopted a strategy involving the identification of resistant/moderately resistant lines at Debre- Zeit, and inter-crossing in Mexico followed by selection for resistance to leaf rust and agronomic type and finally screening for SR reaction in the resulting F6 lines at Debre-Zeit at the same time as they are tested for yield and quality in preliminary yield trials in Mexico. This has generated a significant increase in the proportion of resistant and moderately resistant genotypes within outgoing CIMMYT germplasm, from less than 3% at the onset of the initiative in 2008 to 16% in 2011, and 38% in 2013. SR-resistant germplasm was characterized by similar frequency distributions to other traits in the overall germplasm such as yield potential, drought tolerance and industrial quality parameters. Advances have also been realized using marker-assisted selection (MAS) to introgress Sr22 from bread wheat and to combine it with Sr25, producing advanced lines with 2-gene stacks with confirmed outstanding resistance and superior quality attributes. Since the two genes are closely linked but from different sources bringing them together required a very rare recombination event finally detected via MAS among thousands of plants. They are now essentially inherited together with a very low likelihood of generating recombinant individuals with either gene. The yield potential and stability of these lines are under evaluation in Ethiopia and the best lines are being used in a second round of breeding.
The Lr34/Yr18 gene has been used in agriculture for more than 100 years. In contrast to many other resistance sources against leaf rust and stripe rust, it has remained effective and no virulence has been reported. This makes Lr34 a unique and highly valuable resource for rust resistance breeding. The pleiotropic nature of the gene conferring partial resistance to different pathogen species, the associated leaf tip necrosis and its durability suggest a molecular mechanism that is different from major gene resistance. This is supported by the molecular nature of Lr34 which was recently found to encode an ABC transporter. Interestingly, all tested wheat lines contain an allele of the Lr34 gene on chromosome 7DS. In its susceptible form, the gene does not confer resistance. The difference between the encoded resistant and susceptible LR34 isoforms consists of only two amino acid changes, whereas the rest of the proteins are identical. These two changes must change the biochemical properties of the resistant LR34 transporter in such a way that the plant becomes resistant. We speculate that there is a slight conformational change in the resistant form of the protein, resulting either in modified specificity or kinetics of the transported molecule, or that the binding properties to an unknown second protein interacting with LR34 are changed, resulting in altered function. While the molecular nature of the molecule(s) transported by the LR34 protein remains unclear, it is likely that a physiological change related to Lr34 activity is at the basis of resistance. We are currently establishing transgenic approaches in heterologous grass species to further investigate the molecular activity of Lr34 and to better understand a physiological mechanisms resulting in disease resistance.
Stem rust resistance gene Sr43, derived from tall wheatgrass (Thinopyrum ponticum), is effective against Ug99 lineage Pgt races. Previous studies indicated that Sr43 was located on large Th. ponticum 7el2 chromosome segments in 7D/7el2 translocation stocks KS10-2 and KS24-1. In the present work, we applied a recently-established chromosome engineering procedure to reduce the size of the alien chromosome carrying Sr43. KS10-2 was crossed and backcrossed to the Chinese Spring (CS) ph1b mutant. BC1F1 plants were screened for stem rust response and Ph1- associated molecular markers. Resistant BC1F1 plants homozygous ph1bph1b were further backcrossed to CS. The resulting population of 706 BC2F1 plants was screened for stem rust response and with six co-dominant SSR markers. Wheat lines RWG33 and RWG34 carry Sr43 on shortened alien segments that are about 15% of that in KS10-2. Two molecular markers closely linked to Sr43 were identified; one was an SSR marker and the other a STS marker based on sequences of deletion bin-mapped expressed sequenced tags in wheat. The two new wheat lines with Sr43 and closely-linked markers may provide new resources for combating the threat of race Ug99 and derivatives.
Stem rust, caused by Puccinia graminis f. sp. tritici, is a devastating disease on wheat and barley. A single barley gene, Rpg1, has provided durable resistance since its commercial introduction in the 1940s. The cloned Rpg1 gene encodes a protein with two tandem protein kinase domains, one an active kinase (pK2) and one a pseudokinase (pK1). Function of both domains is required for resistance. The gene is constitutively expressed in all tissues with elevated levels in the epidermis. It is mostly cytoplasmic with small, but significant amount associated with the cell membrane. We have been studying this gene and protein to try to understand how it works and why it has been so durable. Here we report our most recent results showing that RPG1 is phosphorylated within 5 min after urediniospores from avirulent, but not virulent, races land on the leaf surface. Two effector proteins were isolated from the ungerminated spores and shown to work cooperatively to induce RPG1 phosphorylation and eventual degradation. The proteins were identified as a hypothetical protein (PGTG10537.2) with a fibronectin type III and BRCA1 C-terminal domains and vacuolar protein sortingassociated protein 9 (PGTG_16791). The rapidity of the effector function and the nature of the two protein effectors indicate that a unique mechanism for effector entry and signaling in the host cell is involved. This hypothetical mechanism may be similar to what is observed in animal cells where fibronectin proteins with an RGD-binding domain act to mediate communications between the extracellular matrix and plasma membrane.