A number of stem rust resistance genes derived from wild relatives of wheat appeared to be more effective against race TTKSK (Ug99) of Puccinia graminis f. sp. tritici than Sr genes of wheat origin. In an attempt to identify sources of stem rust resistance genes effective against TTKSK, we evaluated several cultivated and wild relatives of wheat for resistance to TTKSK and other stem rust races with broad virulence in seedling tests. Preliminary results indicated that TTKSK resistance could readily be found, but frequencies of resistance varied among the species. Aegilops speltoides had the highest frequency of resistance (nearly 100%). Other species having high frequencies of TTKSK resistance included triticale (77.7% of 567 accessions), Triticum urartu (96.8% of 205 accessions), and T. monococcum (61% of 1020 accessions). Frequencies of TTKSK resistance in other species were: 14.7% in Ae. tauschii (456 accessions), 15% in T. timopheevii (298 accessions), and 17% in T. turgidum ssp. dicoccoides (157 accessions). Based on specific infection types to several races, we postulated that novel genes for resistance to TTKSK are present in some of these species. Accessions with putatively new resistance genes were selected to develop crosses for introgressing resistance into wheat and for developing mapping populations.
Primary Author: Yue Jin, USDA-ARS, Cereal Disease Laboratory
An assessment was made of stem rust race analysis on a global scale. Responses were obtained from 23 rust workers representing 21 countries. Five laboratories have an institutional history in stem rust race analysis of more than 60 years, whereas personal experience in this field ranged from 0 to 35 years. The number of stem rust samples processed from 2006 to 2008 varied greatly between countries. For the three year period most collections were characterized in Canada, followed by Georgia, USA, South Africa and Australia. Most laboratories use the North American differential set and nomenclature system. However, these entries are often supplemented by additional tester lines from the Stakman set, other single gene lines or local cultivars. Differential sets varied between eight and 50 entries. More than half of the respondents indicated that they often encounter seed mixtures amongst their differentiating lines. In recent surveys most races were detected in Ethiopia, followed by Georgia and China. One race dominated the USA and Canadian stem rust population. In South America and Australia stem rust has been rare in commercial wheat for many years. Races within the Ug99 cluster were frequently identified in stem rust collections from Kenya and Ethiopia. Two races related to Ug99, but avirulent on Sr31, occur in South Africa. Several laboratories are in the process of purifying and bulking differential seed, which appears to be one of the major limiting factors in reliable stem rust race analysis. Improvement of infrastructure and training of individuals inexperienced with stem rust should improve global surveillance efforts. In addition, countries doing race analysis should keep viable culture collections in long-term storage.
Primary Author: Zac Pretorius, Department of Plant Sciences, University of the Free State, South Africa
Durum wheat (Triticum durum Desf.) is a major stable crop and it represents a base of the Mediterranean diet. This region is subject to a Mediterranean climate, which is extremely unpredictable with severe changes in moisture and temperature occurring each crop season. This unpredictability is summarized by breeders as GxE and the identification of traits controlling this interaction is quintessential to ensure stability in production season after season. To study the genetics of yield stability, four RILs populations derived from elite x elite crosses were assessed for yield and 1,000-kernel weights across five diverging environments in Morocco and Lebanon. These 550 RILs were characterized with 4,909 polymorphic SNPs via genotyping by sequencing. A consensus map was derived by merging the individual genetic maps of each population. Finally, imputation was used to fill all the missing haplotypes and reach a reduction of missing data to below 8%. Several significant QTLs were identified to be linked to TKW, grain yield and a stability index, namely AMMI wide adaptation index (AWAI). A second approach to identify loci controlling stability was the use of a global panel of 288 elites, accessions and landraces tested in 15 diverging environment. Multi-locations data were compiled via GxE models to derive the AWAI stability index. In addition, this panel was characterized with 8,173 polymorphic SNPs via Axiom 35K array. Significant associations were identified for all traits, including QTLs unique to AWAI. The sum of the identified QTLs can now be pyramid via marker assisted selection and molecular designed crosses in order to obtain very stable cultivars.
Primary Author: ZAIM, University of Mohammed V/ICARDA
Wheat cultivation in many regions faces threats by devastating fungal infections. However, wheat cultivar 92R137 shows resistance to Puccinia striiformis infection. To isolate the stripe rust resistance gene Yr26, an integrated transcriptomic and comparative genomics approach was undertaken. Near-isogenic lines of wheat (carrying Yr26 or not) infected with two Puccinia striiformis f. sp. tritici (Pst) (Virulence or avirulence to Yr26) were analysed in a dual detailed time series RNA-seq study. The emerging IWGSC refseq v1.0 genome assembly sequence serves as a valuable template and was also used for comparative genomics studies of the gene candidate region with the genome sequences of close relatives and wheat progenitors. Using bulked segregant analysis (BSA) to identify polymorphic SNPs between parent and resistant DNA (R-bulk) and susceptible DNA (S-bulk), flanking markers for Yr26 were identified. These two markers were mapped to the Chinese spring reference genome sequence, spanning a region of about 250 kb. The synteny analysis of this candidate region in CS chr1B with chr1A, chr1D, Wild Emmer Wheat (Td_chr1A and Td_chr1B) and Barley (chr1H) identified three candidate Yr26 genes. To detect gene candidates a dual time series RNA-seq analysis was performed. Genes differently expressed between rust susceptible (NIL-S) host lines and rust resistant (NIL-R) lines, carrying the Yr26 candidate gene were analysed. Both lines were inoculated with Pst carrying different avirulence factors (Pst-CYR32 and Pst-V26), compatible or incompatible with the corresponding wheat lines. Differential gene expression analysis (DEG) between compatible and incompatible interaction revealed DEGs in the wheat genome and in the Pst genome. From a network analysis of both wheat and Pst genes, we inferred connected co-expressed modules. Resulting modules showed particular enrichments for disease resistance, defense response to fungus and cell wall components.
Primary Author: Zeng, Northwest A&F University
A Pst pathotype group named V26, virulent to wheat lines possessing Yr26 (=Yr24) has become the third most frequent group in China after races CYR32 and CYR33. Twenty four near-isogenic lines (NILs) and 19 Chinese differentials were used to identify the avirulence/virulence spectra of 36 Yr26-virulent isolates from four provinces (Qinghai, Gansu, Sichuan and Ningxia). Eight races were identified when tested on the NIL set, and 7 races were identified on the Chinese set. There was no relationship with province of origin. Three races identified on the NILs occurred at relatively high frequencies (23, 3, and 3 isolates). Virulence differences existed for Yr1, Yr4, Yr6, Yr9, Yr17, Yr25, Yr32, YrSp, and YrTr1. Among the 7 races identified on the Chinese differentials, one (CYR32 + Yr26 virulence) was represented by 13 isolates and another (CYR33 + Yr26 virulence) included 15 isolates. Among the entire group there were virulence differences on Trigo-Eureka (Yr6+), Lovrin 13 (Yr9+), Kangyin 655, Fengchan 3 (Yr1+), Lovrin 10 (Yr9+), and Hybrid 46 (Yr4+). All isolates were avirulent on Zhong 4 and T. spelta. Using 18 polymorphic simple sequence repeat (SSR) markers, we identified 35 genotypes clustered into two molecular groups (MGs) at a similarity coefficient level of 0.70. SSR analysis also indicated a high level of recombination within the V26 group. The considerable diversity indicates a threat not only to cultivars carrying Yr26, but also to other currently resistant materials.
Primary Author: Zhan, State Key Laboratory of Crop Stress Biology for Arid Areas and College of Plant Protection, Northwest A&F University, P.R. China
Stripe rust, caused by Puccinia striiformis Westend f.sp. tritici, is currently one of the most prevalent and damaging disease on wheat. Up to now, some genes in wheat which are resistant to wheat stripe rust have been cloned, but little is known about the corresponding avirulence gene according to the gene-for-gene hypothesis. A population containing 118 progeny isolates population acquired by selfing an isolate, PL17-7, with virulence to Yr26 was derived. Seventy-two progeny isolates were different in genotype depending on 92 simple sequence repeat (SSR) markers. The progeny population segregated for avirulence to Yr6 at one locus (3 avirulent :1 virulent ratio). The parental isolate and 72 of 118 progeny isolates were resequenced to find candidate avirulence genes corresponding to Yr6. Overall, 7.6 million reads per sample were obtained and mapped to the draft genome of a Chinese Pst isolate CY32. The median depth of coverage was 63.6 fold. For each isolate, between 97.6% and 98.1% of the sequence reads were mapped to the race CY32 genome, which covered between 87.3% and 95.4% of the reference genome bases. An average of 97357 single nucleotide polymorphisms (SNP) per isolate was found, which covered 8.1% of the reference genome. Different SNPs and Indels were found when isolates virulent and avirulent to wheat cultivar containing Yr6 were grouped into two groups. Though screening discrepant SNP and indel in these two groups, candidate avirulence genes corresponding to Yr6 may be found.
Primary Author: Zhan, State Key Laboratory of Crop Stress Biology for Arid Areas and College of Plant Protection, Northwest A&F University
Multiple rust resistance gene combinations are considered as a practical solution for providing durable rust resistance and preventing resistance breakdown arising from single gene deployment. The stem rust resistance locus Sr26, originally derived from Thinopyrum ponticum and introgressed into wheat as a chromosome translocation, is one of the very few genes conferring durable resistance for almost 40 years to all known races of stem rust, including the highly virulent stem rust race Ug99 (TTKSK) and its derivatives (Dundas et al. 2015). To understand the underlying mechanisms of its unusual long-term effectiveness and to explore allelic diversity in different Th. ponticum accessions for other functional alleles that may offer new sources of resistance, we used comparative genomics and gene capture techniques (Resistance gene enrichment sequencing, RenSeq) as complementary strategies for isolating the target gene (Steuernage et al. 2016). Sr26 region was first mapped using NB-LRR (Nucleotide-binding site and leucine-rich repeat) sequences from the orthologous gene members located on the long arm of chromosome 6D from Aegilops tauschii (the D-genome donor of wheat) reference genome. Subsequently, we revealed a cluster of NB-LRR sequences located at the distal end of the Th. ponticum introgression segment that were absent in the smallest interstitial Sr26 deletion mutant. Therefore, we substantially narrowed down the genetic interval for Sr26. In addition to this approach, we subjected the mutant population to RenSeq pipeline. A candidate gene of Sr26 has been successfully identified to be a NBS-LRR type resistance gene. Validation of the gene candidate by complementation studies is currently in progress. In order to enhance durable resistance, genetic stocks of Sr26 from different backgrounds as well as a panel of Sr26-APR (Adult Plant Resistance) gene combinations have been generated to further investigate the resistance response of Sr26 in combination with different multi-pathogen APR genes.
Primary Author: Zhang, CSIRO Agriculture and Food, Australia
Most rust resistant genes in wheat are race-specific (R), with relatively few genes conferring resistance only at the adult stage that have been described as slow rusting genes (APR). Pyramiding multiple R, APR or APR+R genes has been used successfully over many years to achieve durable rust resistance. To further enhance this strategy, a genetic genomics approach was exploited to identify genes with different resistant mechanisms and the most effective gene pyramids.
Several new combinations of rust genes were created and tested in the Thatcher background, revealing synergistic ("booster") effects involving Lr21 with Lr16. With QTL mapping approach, we found that genes combined from 7D, 1B and 7B conferred an almost immune response to leaf rust, while genes from 7D, 1B and 3B provided an almost immune response to stripe rust. With a genomics approach, a large scale transcriptome analysis was conducted on key rust resistant genes including six R genes, three APR genes and one gene pyramid with Lr34+Lr16 over a time series during the infection process of both seedlings and adult plants. Detailed transcriptome analysis of gene expression associated with different major and minor leaf rust genes, alone or in combination, identified common and unique aspects of defense responses. For example, Lr9 is different from the other three leaf rust genes tested, with resistance triggered at a very early stage, consistent with pre-haustorial resistance. R genes Lr21 and Lr16 were also significantly different compared to other R and APR genes. With gene co-expression network analysis, a shared unique gene module mediated by Lr34 and Lr67 was also identified. This large transcriptome dataset also allowed the development of a rust-wheat interactome atlas for rust functional genomics research in wheat.
Primary Author: Zhang, National Research Council of Canada (NRC)-Saskatoon
Genetic analysis of YrA resistance in Avocet R confirmed two complementary resistance genes. Marker-trait association analysis on a doubled haploid (DH) population derived from Teal/Avocet R mapped one of the genes to chromosome 5BL. A DArT-Seq genetic map for the population indicated the presence of a T5B-7B reciprocal translocation. Fluorescence in situ hybridization (FISH) confirmed the translocation in the susceptible parent Teal relative to Avocet R. Additional FISH examinations on Cappelle Desprez (CD), Chinese Spring (CS) and Avocet S as controls indicated that the translocation in Teal was similar to that in CD. FISH studies also revealed additional polymorphisms in both chromosome 7B arms of Avocet R and Avocet S relative to CS, and that chromosome 5B in Avocet S lacked about 32% of the long arm relative to Avocet R and CS. It was postulated that complementary gene Yr74 was deleted with the missing segment. Australian cultivars Banks, Condor and Egret are also polymorphic for stripe rust response, and intercrosses in earlier studies between the S selections failed to confirm complementary genes. FISH analyses are currently underway to test the hypothesis that the S selections carry the deletion. Results will be reported in the poster.
Primary Author: Zhang, The University of Sydney, Plant Breeding Institute, Australia
Leaf rust is the most widely occurring disease of wheat worldwide. Resistance is the most practical and effective way to control the disease. Most leaf rust resistance genes are race-specific (“R”, qualitative resistance) and a relatively few are adult plant resistance genes, some of which have been described as slow rusting (“APR”, quantitative resistance). Due to limited knowledge, most resistance genes have been deployed in cultivars by an inefficient “blind” approach. This results in the well known “boom and bust cycle” (resistance followed by susceptibility) because the pathogen evolves rapidly and migrates over long distances. Therefore, a breeding-by-design approach is needed to achieve durable resistance. Pyramiding multiple R, APR or APR+R genes has been used successfully over many years to achieve durable resistance to leaf rust in Canada and some other countries. To further enhance this strategy we seek to understand the molecular mechanisms underlying key resistance genes. To identify the molecular mechanisms underlying rust resistance conferred by major R and APR genes, we performed an integrated systemic transcriptome analysis via RNA-seq on the Thatcher NILs with Lr16, Lr22a, Lr21, Lr34, Lr34+Lr16, and Lr67 challenged with Pt race BBBD. Sampling was conducted over a time series during the infection process of both seedlings and adult plants. Through RNA-seq we were able to capture the dynamic interactome of host-pathogen interactions conferred by these R and APR genes. Preliminary results revealed that resistance reactions conferred by R gene Lr21 and APR gene Lr67 were significantly different compared to other R and APR genes. Significantly, the Thatcher NIL line with Lr34+Lr16 showed the combines defense reactions of Lr16 and Lr34.
Primary Author: Zhang, National Research Council of Canada, Canada