All BGRI Abstracts

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Genetic diversity among Puccinia graminis f. sp. secalis and winter rye populations

Flath Julius Kuehn-Institut, Institute of Plant Protection in Field Crops and Grassland, Germany

Rye stem rust (caused by Puccinia graminis f. sp. secalis, Pgs) causes considerable yield losses in rye crops grown in continental climates. In Germany, stem rust resistance in rye has attracted little attention until now. In order to implement resistance breeding, it is of utmost importance to (1) analyze Pgs populations in terms of diversity and pathotype distribution, and (2) identify resistance sources in winter rye populations. Within a three-year research project, we analyzed 389 single-pustule-isolates, collected mainly from German rye-growing areas, on 15 rye inbred differentials with different avirulence/virulence patterns; among them, 226 pathotypes were identified and only 56 occurred more than once. The majority of isolates infected 5-6 differentials. This high diversity was confirmed by a Simpson index of 0.99, a high Shannon index (5.27) and an evenness index of 0.97. In parallel, we investigated stem-rust resistance among and within 122 genetically heterogeneous rye populations originating from 19 countries across 3 to 15 environments (location-year combinations) in two replicates. While 7 German commercial rye populations were highly susceptible, 11 non-adapted populations, mainly from Russia, Austria and the USA, were highly resistant, harboring 32-70% resistant stems on plots averaged across 8 to 10 environments. Selections for low disease severity at the adult-plant stage in the field also displayed resistance in leaf-segment tests (r=0.86, P<0.01). In conclusion, rye stem rust pathogen populations are highly diverse and the majority of resistances in rye populations are race-specific. The new Pgs isolate set firstly developed within the project covers the current spectrum of virulences and can be used to assess the effectiveness of stem rust resistance genes or sources. New pathotypes can be detected using this differential set and farmers and industry can be alerted to circumvent economic damage. In the long term, resistances from non-adapted populations should be introgressed into commercial rye cultivars.

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Chromosomal locations of stem and leaf rust resistance genes from Ae. caudata, Ae. searsii and Ae. Mutica

Dundas School of Agriculture, Food and Wine, The University of Adelaide, Australia.
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The wild relatives of wheat represent a vast resource of potentially useful genes for agriculture. The genus Aegilops has provided several rust resistance genes used in commercial cultivars. Here we report progress on mapping of potentially new stem and leaf rust resistance from Ae. caudata, Ae. searsii and Ae. mutica (Amblyopyrum muticum). Addition lines derived from the amphiploids Alcedo/ Ae. caudata, TA3368, CS/ Ae. mutica, TA8024 (both from Wheat Genetics Resource Center, Kansas State University, USA) and CS/ Ae. searsii TE10 (kindly provided by Dr Moshe Feldman, Weizmann Institute, Rehovot, Israel) were produced after backcrossing the amphiploids with Australian cv. Angas or Westonia. Backcrossed generations were screened for stem rust and leaf rust responses and both resistant and susceptible plants were sampled for DNA marker analysis. Stem rust resistant plants derived from the Ae. caudata amphiploid and leaf rust resistant plants derived from the Ae. searsii amphiploid showed the presence of non-wheat marker bands after hybridizing restricted genomic DNA with the Triticeae group 5 RFLP probe PSR128, and after PCR using EST-based primers specific for Triticeae group 5. Susceptible plants did not show those non-wheat molecular markers. Hence, stem rust resistance from Ae. caudata was allocated to chromosome 5C, and the resistance gene is temporarily named SrAec1t. Leaf rust resistance from Ae. searsii was allocated in a similar manner to chromosome 5Ss, and is temporarily named LrAesr1t. Leaf rust resistance transferred from Ae. mutica was traced to a 6T chromosome after associating resistance with the presence of Triticeae group 6 RFLP probes (including BCD001, BCD269, BCD276, BCD1426, CDO772, CDO1380, WG933) and that gene is temporarily named LrAmm1t. The addition lines involving the 5C, 5Ss and 6T chromosomes were crossed with Sears’ ph1b mutant to induce homoeologous recombination with related wheat chromosomes.

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Determining the breeding value of CIMMYT’s International Bread Wheat Nursery (IBWSN) entries for leaf, stem and stripe rust resistance

Juliana Plant Breeding and Genetics Section, School of Integrative Plant Science, Cornell University, USA

The quest for durable rust resistance in wheat is burgeoning with the emergence of new virulent races. Breeders challenged with this unceasing plant-pathogen arms race have to devise strategies for effective evaluation and exploitation of the rust resistance genes. Considering the likely presence of useful variation for rust resistance in CIMMYT’s international bread wheat screening nurseries (IBWSN), we implemented genomic prediction in the 45th and 46th IBWSN entries to determine their genomic estimated breeding values (GEBV’s) for leaf, stem and stripe rust resistance. The 350 lines (45th IBWSN) and 329 lines (46th IBWSN) were phenotyped in replicated trials over two to three years in El Batan, Mexico (leaf rust); Njoro, Kenya (stem rust) and Toluca, Mexico (stripe rust). The filtered genotyping data for these two nurseries comprised of 6,786 and 11,218 genotyping by sequencing (GBS) markers. Our objective was to compare the GEBV’s estimated by four different models: multiple linear regression (MLR) with QTL-linked markers as fixed effects; Genomic-best linear unbiased prediction (G-BLUP); G-BLUP mixed model which includes QTL linked markers as fixed effects and Bayesian least absolute shrinkage and selection operator (LASSO). We observed that the prediction accuracies (calculated using 10-fold cross validation) were the highest for stripe rust (0.52 to 0.61), followed by stem rust (0.42 to 0.65) and leaf rust (0.15 to 0.45). Among the models, the MLR gave the lowest prediction accuracies (0.15,0.42 and 0.52), while G-BLUP (0.45,0.59 and 0.59), mixed G-BLUP (0.38,0.65 and 0.62) and the Bayesian LASSO (0.45,0.58 and 0.61) yielded relatively higher and almost similar accuracies. Overall, our results are promising and indicate that using genome-wide markers is advantageous than including only significant QTL-linked markers. We hope that implementing genomic prediction in breeding programs, would help to achieve rapid gains from selection and revolutionize our efforts in combating the rust pathogen.

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Supercharging the radars for pathogen surveillance

Saunders National Institute of Agricultural Botany, UK

Emerging and re-emerging diseases of humans, animals and plants pose a significant hazard to public health and food security. With recent advances in sequencing technology, bacteriologists and virologists are now integrating high-resolution genotypic data into pathogen surveillance activities. However, the application of genomics to emerging filamentous plant pathogens has lagged. To address this, we developed a robust and rapid “field pathogenomics” strategy. We applied this method in 2013 to the wheat yellow rust pathogen Puccinia striiformis f. sp. tritici (Pst), using gene sequencing of Pst-infected wheat leaves taken directly from the field to gain insight into the population structure of a re-emerging pathogen. Our analysis uncovered a dramatic shift in the Pst population in the UK and supports the hypothesis that recent introduction of a diverse set of exotic Pst lineages may have displaced the previous population. Gene sequencing of infected host tissue can also be leveraged to assess the genotype of the host, rapidly confirming whether previously resistant wheat varieties have indeed been overcome. We have now expanded this study to analyze Pst-infected plant samples from across Europe and beyond and will provide an update on the insights we have gained regarding Pst population dynamics. Working with cross-institutional and industrial partners we are now developing this technique further to reduce cost so it can be applied routinely within the U.K. cereal disease surveillance program.

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Genetic analysis and molecular mapping of resistance in barley to the heterologous rust pathogens Puccinia graminis f. sp. avenae and P. triticina

Haghdoust The University of Sydney, Plant Breeding Institute, Australia

Stem rust is considered one of the most important threats to world cereal production. The appearance and spread of the wheat stem rust pathogen [Puccinia graminis f. sp. tritici (Pgt)] race Ug99 has caused great concern for global wheat production. Barley is a host to different specialized pathogen species such as Pgt, but is characteristically a near nonhost to most non-adapted (heterologous) rust pathogens such as the wheat leaf rust pathogen [P. triticina] and oat stem rust pathogen [P. graminis f. sp. avenae (Pga)]. The barley research line SusPtrit, developed for susceptibility to heterologous rust pathogens, is a useful resource to study the genetics of nonhost resistance and to clone the genes involved, particularly due to the recent availability of the genome sequence. Studies in wheat suggest that resistance genes that are effective against multiple rust pathogens (pleiotropic) such as Lr34/Yr18/Sr55, confer durable disease control. We intercrossed the sequenced barley genotype Morex with SusPtrit to determine the inheritance of resistance to the wheat leaf rust and oat stem rust pathogens. The F2 population segregated for a single dominant resistance gene in response to both heterologous pathogens Pga and Pt. Subsequent progeny testing and genetic analysis of the segregating F3 population will be performed to map and determine the relationship between the resistance genes. Large F2 populations were developed to fine map and clone the genes, and ultimately to transfer them into related crop species as an alternative approach for crop protection.

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Cytogenetic analysis and molecular mapping of inheritance of resistance in barley to Puccinia striiformis

Elmansour The University of Sydney, Plant Breeding Institute, Australia

Plants are generally non-hosts to most diseases. Barley is a host to Puccinia striiformis f. sp. hordei, but is a near non-host to P. striiformis f. sp. tritici (Pst) and to P. striiformis f. sp. pseudohordei (Psp), which cause stripe rust on wheat and barley grass (Hordeum murinum, H. leporinum), respectively. This study was carried out to determine the inheritance of resistance in barley line 81882/BS1 using the mapping population: 81882/BS1/Biosaline-19. 81882/BS1 is a H. vulgare derivative of cv. Vada, carrying an introgression from H. bulbosum on chromosome 2HS, and Biosaline-19 is susceptible to both Pst and Psp. Phenotyping of F3 lines with Psp culture 981549 and Pst pathotype 134 E16 A+ showed that 81882/BS1 carried two genes for resistance to Psp, and three genes for resistance to Pst. Cytogenetic analysis and molecular mapping were performed to further characterize the resistance of 81882/BS1 to Psp. Joint phenotypic and cytogenetic analysis indicated that at least one of the genes for resistance to Psp was associated with the H. bulbosum introgression previously located on chromosome 2H (Zhang unpublished). Preliminary molecular mapping of 15 non-segregating resistant and 15 non-segregating susceptible lines using >10K DArTseq molecular markers located the second gene on chromosome 1H. This gene was probably contributed by Vada. Further studies are underway to confirm the locations of these two loci by fine mapping.

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Pathogenic variation in Puccinia striiformisf.sp.tritici(Pst) Following selfing on Berberis shensiana

Kang State Key Laboratory of Crop Stress Biology for Arid Areas and College of Plant Protection, Northwest A&F University, P.R. China

Pst is highly variable, and new races that overcome newly released resistant cultivars are regular events. The widely virulent race V26 (virulent to Yr26) has a significant potential to cause epidemics in China. In this study teliospores from a single urediniospore isolate of V26 (No. Pinglan 17-7) produced on the Nanjing wheat line 92R137 (Yr26) were induced to germinate and infect Berberis shensiana as a sexual host. One hundred and eighteen single aeciospore (SA) selfed progeny and the V26 parent were typed for pathogenicity on a set of differentials comprising 22 Yrnear-isogenic wheat lines (NILs). Virulence phenotyping was conducted twice for all isolates, and similar results were obtained each time. The V26 isolate (No. Pinglan 17-7) was avirulent on differentials with Yr5, Yr6, Yr8, Yr15, Yr43, YrSp, YrTr1 and virulent on those with Yr1, Yr2, Yr4, Yr7, Yr9, Yr10, Yr17, Yr25, Yr26, Yr27, Yr28, Yr32, Yr44, YrV23, and YrExp2. The progeny were all virulent to Yr1, Yr2 (Kalyansona), Yr7, Yr9, Yr10, Yr17, Yr25, Yr26, YrV23 (Vilmorin 23) and YrExp2, and all avirulent to Yr5, Yr8, Yr15, and YrTr1, suggesting that V26 is homozygous at the corresponding pathogenicity loci. Various segregation ratios were apparent for other Yrgenes (P values ranging from 0.6to 0.09).These included3 avirulent: 1 virulent with respect to Yr6 and Yr43, 1 avirulent : 3 virulent forYr27 and Yr28, 1 avirulent : 15 virulent forYr4, Yr32, and Yr44,and 13 avirulent : 3 virulent for YrSp. Among the 118 progeny,27 of new pathotypes were identified as compared with the avirulence/virulence loci of the parent isolate. A study of the population based on markers and development of a molecular map is in progress.

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Genetic gain in wheat from phenotypic and genomic selection for quantitative resistance to stem rust

Rutkoski 1International Programs in the College of Agriculture and Life Sciences, and Plant Breeding and Genetics Section in the School of Integrative Plant Science, Cornell University, USA, and CIMMYT, Mexico

Stem rust is a globally important wheat disease that can cause severe yield loss. Breeding for quantitative stem rust resistance (QSRR) is important for developing cultivars with durable resistance. Genomic selection (GS) could increase rates of genetic gain for quantitative traits, but few experiments comparing GS and phenotypic selection (PS) have been conducted. Our objectives were to compare realized gain from GS based on markers only with that of PS for QSRR in spring wheat using equal selection intensities; determine if gains agree with theoretical expectations; and compare the impact of GS and PS on inbreeding, genetic variance, and correlated response for pseudo-black chaff (PBC), a correlated and likely pleiotropic trait. Over two years, two cycles of GS were performed in parallel with one cycle of PS, with each method replicated twice. For GS, markers were generated using genotyping-by-sequencing, the prediction model was initially trained using historical data, and the model was updated before the second GS cycle. Overall, GS and PS led to a 31±11 and 42±12% increase in QSRR and a 138±22 and 180±70% increase in PBC, respectively. Genetic gains were not significantly different, but were in agreement with expectations. Per year, gains from GS and PS were equal, but GS led to significantly lower genetic variance. This shows that while GS and PS can lead to equal rates of short-term gains, GS can reduce genetic variance more rapidly. Further work to develop efficient GS implementation strategies in spring wheat is warranted.

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Genomic regions containing multiple disease resistance loci

Joukhadar Department of Animal, Plant and Soil Sciences, La Trobe University, Australia

Climatic changes permit the spread of plant diseases to new areas. To ensure grain production meets the needs of the growing world population, wheat breeding must combine multiple disease resistances in single cultivars in order to maintain current yield potential. Over the last three decades, classical mapping and association studies have identified disease resistance loci for individual diseases, but few studies have investigated loci that confer resistance to multiple diseases. To address this limitation, we extensively surveyed the literature to identify wheat genomic regions harboring resistance to multiple diseases. We identified 174 trait-linked markers distributed across all wheat chromosomes, except chromosome 4A, and the numbers of disease resistance loci in each region ranged from two to ten. Our survey suggests that some regions of the genome contain multiple disease resistance genes, or genes with pleiotropic effects. We are using the Chinese Spring flow sorted chromosome survey contigs to investigate the genic contents of genomic regions containing multiple disease trait loci to address this question.

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Identification of resistance factors against rust fungi: leveraging genetic and genomic tools in Brachypodium - rust fungi pathosystems

Figueroa Department of Plant Pathology, University of Minnesota, USA

During the past 15 years, significant efforts have been directed to develop the grass species Brachypodium distachyon as a genetically tractable model for monocot plants, especially economically valuable cereals such as wheat, barley and oat. Such efforts have led to an increasing availability of genomic, genetic and bioinformatics tools designed to bypass the experimental challenges faced when addressing important biological questions in complex systems. Moreover, such advances may translate in the use of other valuable species of Brachypodium (e.g., B. hybridum), which are not nearly as well characterized as B. distachyon. Given the 2050 global food demands and needs to increase grain production we seek to develop innovative and sustainable approaches to decrease crop yield losses due to rust fungi. One possible strategy is the use of transgenic plants harboring non-host resistance-related genes from closely related species. B. distachyon and B. hybridum can serve as potential sources to engineer plant resistance against highly destructive rust fungi, such as Puccinia graminis and P. coronata. Advancing our understanding of non-host resistance in monocot species has been a slow process. However, the amenability of Brachypodium as a model system offers a means to accelerate scientific discovery of factors controlling non-host pathogen interactions involving stem and crown rust fungi. In a multi-pronged approach, we are leveraging genetic and genomic tools, as well as generating new resources to provide foundational knowledge in order to support plant genetic engineering programs.

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