Puccinia coronata var. coronata, a Crown Rust Pathogen of Two Highly Invasive Species, Is Detected Across the Midwest and Northeastern United States.

Puccinia coronata var. coronata (Pcc) causes crown rust disease of glossy buckthorn (Frangula alnus) and reed canarygrass (Phalaris arundinacea), two highly invasive plant species in North America. Pcc is closely related to major pathogens of cereals, turfgrasses, and forage grasses. It occurs throughout Europe but was first recorded in North America in 2013. Where its hosts co-occur, such as in wetlands in the Twin Cities metro area in Minnesota, we have observed Pcc causing significant infection that results in defoliation and fruit loss in glossy buckthorns and premature leaf senescence in reed canarygrass. In this research, we mapped the distribution of this likely recently introduced rust fungus and provided a description of disease signs and symptoms and pathogen morphology. Samples were acquired by two primary means: by surveys in Minnesota and by correspondence with users of iNaturalist.org, a social network for nature enthusiasts and community scientists. With an Oxford Nanopore MinION, we sequenced two to four loci from 22 samples across 13 states and identified samples by phylogenetic analysis and sequence similarity. Notably, four pure isolates appear to have intragenomic variation of the ITS region. We found that Pcc is present throughout the range of glossy buckthorn in the eastern United States. In Minnesota, Pcc is not common outside the range of glossy buckthorn despite the presence of susceptible grass hosts.

Aegilops sharonensis genome-assisted identification of stem rust resistance gene Sr62.

The wild relatives and progenitors of wheat have been widely used as sources of disease resistance (R) genes. Molecular identification and characterization of these R genes facilitates their manipulation and tracking in breeding programmes. Here, we develop a reference-quality genome assembly of the wild diploid wheat relative Aegilops sharonensis and use positional mapping, mutagenesis, RNA-Seq and transgenesis to identify the stem rust resistance gene Sr62, which has also been transferred to common wheat. This gene encodes a tandem kinase, homologues of which exist across multiple taxa in the plant kingdom. Stable Sr62 transgenic wheat lines show high levels of resistance against diverse isolates of the stem rust pathogen, highlighting the utility of Sr62 for deployment as part of a polygenic stack to maximize the durability of stem rust resistance.

Development of a Diagnostic Assay for Differentiation Between Genetic Groups in Clades I, II, III, and IV of Puccinia graminis f. sp. tritici.

Wheat stem rust has reemerged as a serious disease caused by new variants of Puccinia graminis f. sp. tritici. Variants with significant virulence and broad geographic distribution (Africa, Central Asia, and Europe) include the Ug99 race group, race TTRTF, and TKTTF race group. Genetic analysis has placed isolates representing these critical new virulent races into 12 genetic groups that make up clades I to IV. Development of molecular diagnostic assays for these 12 genetic groups will be an important component of global surveillance efforts. A single-nucleotide polymorphism database was mined for candidate markers that would differentiate between these 12 genetic groups. Thirty-five candidate markers were screened, and a core set of 17 markers was tested against a set of 94 isolates representing a broad range of genotypes and race phenotypes. These core markers were 100% accurate in identifying the 12 genetic groups for 52 isolates in clades I to IV, and no false positives were observed with nontarget isolates. The assay has built-in redundancy so that minor genetic changes or errors in genotyping calling will not affect the accuracy of the results. This assay is also effective in identifying the genetic groups in clade V from Germany and Georgia, the three main subgroups in North American clade VI, and clade VII consisting of race TTTTF found in North and South America. This assay provides a rapid diagnostic tool for both living and nonliving samples to detect these critical new races or race groups of P. graminis f. sp. tritici.[Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Field resistance to wheat stem rust in durum wheat accessions deposited at the USDA National Small Grains Collection.

Wheat stem rust, caused by Puccinia graminis f. sp. tritici, is a re-emerging disease, posing a significant threat to durum wheat production worldwide. The limited number of stem rust resistance genes in modern cultivars compels us to identify and incorporate new effective genes in durum wheat breeding programs. We evaluated 8,245 spring durum wheat accessions deposited at the USDA National Small Grains Collection (NSGC) for resistance in field stem rust nurseries in Debre Zeit, Ethiopia and St. Paul, MN (USA). A higher level of disease development was observed at the Debre Zeit nursery compared with St. Paul, and the effective alleles of Sr13 in this nursery did not display the level of resistance observed at the St. Paul nursery. Four hundred and ninety-one (∽6%) accessions exhibited resistant to moderately susceptible responses after three field evaluations at Debre Zeit and two at St. Paul. Nearly 70% of these accessions originated from Ethiopia, Mexico, Egypt, and USA. Eight additional countries, namely Portugal, Turkey, Italy, Canada, Chile, Australia, Syria, and Tunisia contributed to 19% of the resistant to moderately susceptible entries. Among the 491 resistant to moderately susceptible accessions, 53.8% (n = 265) were landraces, and 28.4% (n = 139) and 11.4% (n = 55) were breeding lines and cultivars, respectively. Breeding lines and cultivars displayed a higher level and frequency of resistance than the landraces. We concluded that a large number of durum wheat accessions from diverse origins deposited at the NSGC can be exploited for diversifying and improving stem rust resistance in wheat.

Function and evolution of allelic variations of Sr13 conferring resistance to stem rust in tetraploid wheat (Triticum turgidum L.).

The resistance gene Sr13 is one of the most important genes in durum wheat for controlling stem rust caused by Puccinia graminis f. sp. tritici (Pgt). The Sr13 functional gene CNL13 has haplotypes R1, R2 and R3. The R1/R3 and R2 haplotypes were originally designated as alleles Sr13a and Sr13b, respectively. To detect additional Sr13 alleles, we developed Kompetitive allele specific PCR (KASP™) marker KASPSr13 and four semi-thermal asymmetric reverse PCR markers, rwgsnp37-rwgsnp40, based on the CNL13 sequence. These markers were shown to detect R1, R2 and R3 haplotypes in a panel of diverse tetraploid wheat accessions. We also observed the presence of Sr13 in durum line CAT-A1, although it lacked any of the known haplotypes. Sequence analysis revealed that CNL13 of CAT-A1 differed from the susceptible haplotype S1 by a single nucleotide (C2200T) in the leucine-rich repeat region and differed from the other three R haplotypes by one or two additional nucleotides, confirming that CAT-A1 carries a new (R4) haplotype. Stem rust tests on the monogenic, transgenic and mutant lines showed that R1 differed from R3 in its susceptibility to races TCMJC and THTSC, whereas R4 differed from all other haplotypes for susceptibility to TTKSK, TPPKC and TCCJC. Based on these differences, we designate the R1, R3 and R4 haplotypes as alleles Sr13a, Sr13c and Sr13d, respectively. This study indicates that Sr13d may be the primitive functional allele originating from the S1 haplotype via a point mutation, with the other three R alleles probably being derived from Sr13d through one or two additional point mutations.

Mapping and characterization of two stem rust resistance genes derived from cultivated emmer wheat accession PI 193883.

KEY MESSAGE: Two stem rust resistance genes identified on chromosome arms 2BL and 6AL of the cultivated emmer wheat accession PI 193883 can be used for protecting modern varieties against Ug99 strains. The wheat research community consistently strives to identify new genes that confer resistance to stem rust caused by the fungal pathogen Puccinia graminis f. sp. tritici Eriks & E. Henn (Pgt). In the current study, our objective was to identify and genetically characterize the stem rust resistance derived from the cultivated emmer accession PI 193883. A recombinant inbred line population developed from a cross between the susceptible durum wheat line Rusty and PI 193883 was genotyped and evaluated for reaction to Pgt races TTKSK, TRTTF, and TMLKC. Two QTLs conferring resistance were identified on chromosome arms 2BL (QSr.fcu-2B) and 6AL (QSr.fcu-6A). The stem rust resistance gene (Sr883-2B) underlying QSr.fcu-2B was recessive, and based on its physical location it is located proximal to the Sr9 region. QSr.fcu-6A was located in the Sr13 region, but PI 193883 is known to carry the susceptible haplotype S4 for Sr13, indicating that the gene underlying QSr.fcu-6A (Sr883-6A) is likely a new allele of Sr13 or a gene residing close to Sr13. Three IWGSC scaffold-based simple sequence repeat (SSR) and two SNP-based semi-thermal asymmetric reverse PCR (STARP) markers were developed for the Sr883-2B region, and one STARP marker was developed for Sr883-6A. Sr883-2B was epistatic to Sr883-6A for reaction to TTKSK and TRTTF, and the two genes had additive effects for TMLKC. These two genes and the markers developed in this research provide additional resources and tools for the improvement in stem rust resistance in durum and common wheat breeding programs.

Presence of a Sexual Population of Puccinia graminis f. sp. tritici in Georgia Provides a Hotspot for Genotypic and Phenotypic Diversity.

Wheat stem rust, caused by Puccinia graminis f. sp. tritici, is a re-emerging disease exemplified by recent epidemics caused by new virulent races. Understanding the sources and origins of genetic variations in the pathogen populations globally can facilitate the development of better strategies in disease management. We analyzed 68 wheat stem rust samples collected between 2013 and 2015 from Georgia where stem rust incidences are frequent and the alternate host, common barberry, is present. A total of 116 single-pustule isolates were derived and evaluated on stem rust differential lines to determine the virulence phenotypes and 23 races were identified, many of which were detected for the first time. Unique virulence combinations including, Sr22+Sr24 and Sr13b+Sr35+Sr37 were detected. These virulence combinations pose new challenges to breeding programs because many of these genes are used in breeding for resistance to the Ug99 race group. Sixty-one isolates were genotyped using a custom single-nucleotide polymorphism chip and 17 genotypes were identified. The 2013 isolates contained 11 multilocus genotypes compared with isolates of 2014 and 2015, with five and three genotypes, respectively. The higher levels of virulence and genotypic diversity observed in the 2013 samples strongly indicated that sexual recombination occurs in the Georgian P. graminis f. sp. tritici population, and that the Caucasus region of Eurasia may be an important source of new races.[Formula: see text] Copyright © 2019 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Identification of New Sources of Resistance to Wheat Stem Rust in Aegilops spp. in the Tertiary Genepool of Wheat.

Recent stem rust epidemics in eastern Africa and elsewhere demonstrated that wheat stem rust is a re-emerging disease posing a threat to wheat production worldwide. The cultivated wheat gene pool has a narrow genetic base for resistance to virulent races, such as races in the Ug99 race group. Wild relatives of wheat are a tractable source of stem rust resistance genes. Aegilops species in the tertiary genepool have not been exploited to any great extent as a source of stem rust resistance. We evaluated 1,422 accessions of Aegilops spp. for resistance to three highly virulent races (TTKSK, TRTTF, and TTTTF) of Puccinia graminis f. sp. tritici. Species studied include Ae. biuncialis, Ae. caudata, Ae. comosa, Ae. cylindrica, Ae. geniculata, Ae. neglecta, Ae. peregrina, Ae. triuncialis, and Ae. umbellulata that do not share common genomes with cultivated wheat. High frequencies of resistance were observed as 977 (68.8%), 927 (65.2%), and 850 (59.8%) accessions exhibited low infection types to races TTKSK, TTTTF, and TRTTF, respectively. Contingency table analyses showed strong association for resistance to different races in several Aegilops spp., indicating that for a given species, the resistance genes effective against multiple races. Inheritance studies in selected accessions showed that resistance to race TTKSK is simply inherited.

Genotyping-by-Sequencing Facilitates a High-Density Consensus Linkage Map for Aegilops umbellulata, a Wild Relative of Cultivated Wheat.

High-density genetic maps are useful to precisely localize QTL or genes that might be used to improve traits of nutritional and/or economical importance in crops. However, high-density genetic maps are lacking for most wild relatives of crop species, including wheat. Aegilops umbellulata is a wild relative of wheat known for its potential as a source of biotic and abiotic stress resistance genes. In this work, we have developed a framework consensus genetic map using two biparental populations derived from accessions PI 298905, PI 542369, PI 5422375, and PI 554395. The framework map comprised 3009 genotype-by-sequence SNPs with a total map size of 948.72 cM. On average, there were three SNPs per centimorgan for each chromosome. Chromosome 1U was the shortest (66.5 cM), with only 81 SNPs, whereas the remaining chromosomes had between 391 and 591 SNP markers. A total of 2395 unmapped SNPs were added to the linkage maps through a recombination frequency approach, and increased the number of SNPs placed on the consensus map to a total of 5404 markers. Segregation distortion was disproportionally high for chromosome 1U for both populations used to construct component linkage maps, and thus segregation distortion could be one of the probable reasons for the exceptionally reduced linkage size for chromosome 1U. From comparative analysis, Aeumbellulata chromosomes except 4U showed moderate to strong collinearity with corresponding homeologous chromosomes of hexaploid wheat and barley. The present consensus map may serve as a reference map in QTL mapping and validation projects, and also in genome assembly to develop a reference genome sequence for Ae. umbellulata.

Discovery and characterization of two new stem rust resistance genes in Aegilops sharonensis.

KEY MESSAGE: We identified two novel wheat stem rust resistance genes, Sr-1644-1Sh and Sr-1644-5Sh in Aegilops sharonensis that are effective against widely virulent African races of the wheat stem rust pathogen. Stem rust is one of the most important diseases of wheat in the world. When single stem rust resistance (Sr) genes are deployed in wheat, they are often rapidly overcome by the pathogen. To this end, we initiated a search for novel sources of resistance in diverse wheat relatives and identified the wild goatgrass species Aegilops sharonesis (Sharon goatgrass) as a rich reservoir of resistance to wheat stem rust. The objectives of this study were to discover and map novel Sr genes in Ae. sharonensis and to explore the possibility of identifying new Sr genes by genome-wide association study (GWAS). We developed two biparental populations between resistant and susceptible accessions of Ae. sharonensis and performed QTL and linkage analysis. In an F6 recombinant inbred line and an F2 population, two genes were identified that mapped to the short arm of chromosome 1Ssh, designated as Sr-1644-1Sh, and the long arm of chromosome 5Ssh, designated as Sr-1644-5Sh. The gene Sr-1644-1Sh confers a high level of resistance to race TTKSK (a member of the Ug99 race group), while the gene Sr-1644-5Sh conditions strong resistance to TRTTF, another widely virulent race found in Yemen. Additionally, GWAS was conducted on 125 diverse Ae. sharonensis accessions for stem rust resistance. The gene Sr-1644-1Sh was detected by GWAS, while Sr-1644-5Sh was not detected, indicating that the effectiveness of GWAS might be affected by marker density, population structure, low allele frequency and other factors.

Genotype-by-sequencing facilitates genetic mapping of a stem rust resistance locus in Aegilops umbellulata, a wild relative of cultivated wheat.

BACKGROUND: Wild relatives of wheat play a significant role in wheat improvement as a source of genetic diversity. Stem rust disease of wheat causes significant yield losses at the global level and stem rust pathogen race TTKSK (Ug99) is virulent to most previously deployed resistance genes. Therefore, the objective of this study was to identify loci conferring resistance to stem rust pathogen races including Ug99 in an Aegilops umbelluata bi-parental mapping population using genotype-by-sequencing (GBS) SNP markers. RESULTS: A bi-parental F2:3 population derived from a cross made between stem rust resistant accession PI 298905 and stem rust susceptible accession PI 542369 was used for this study. F2 individuals were evaluated with stem rust race TTTTF followed by testing F2:3 families with races TTTTF and TTKSK. The segregation pattern of resistance to both stem rust races suggested the presence of one resistance gene. A genetic linkage map, comprised 1,933 SNP markers, was created for all seven chromosomes of Ae. umbellulata using GBS. A major stem rust resistance QTL that explained 80% and 52% of the phenotypic variations for TTTTF and TTKSK, respectively, was detected on chromosome 2U of Ae. umbellulata. CONCLUSION: The novel resistance gene for stem rust identified in this study can be transferred to commercial wheat varieties assisted by the tightly linked markers identified here. These markers identified through our mapping approach can be a useful strategy to identify and track the resistance gene in marker-assisted breeding in wheat.

Kenyan Isolates of Puccinia graminis f. sp. tritici from 2008 to 2014: Virulence to SrTmp in the Ug99 Race Group and Implications for Breeding Programs.

Frequent emergence of new variants in the Puccinia graminis f. sp. tritici Ug99 race group in Kenya has made pathogen survey a priority. We analyzed 140 isolates from 78 P. graminis f. sp. tritici samples collected in Kenya between 2008 and 2014 and identified six races, including three not detected prior to 2013. Genotypic analysis of 20 isolates from 2013 and 2014 collections showed that the new races TTHST, TTKTK, and TTKTT belong to the Ug99 race group. International advanced breeding lines were evaluated against an isolate of TTKTT (Sr31, Sr24, and SrTmp virulence) at the seedling stage. From 169 advanced lines from Kenya, 23% of lines with resistance to races TTKSK and TTKST were susceptible to TTKTT and, from two North American regional nurseries, 44 and 91% of resistant lines were susceptible. Three lines with combined resistance genes were developed to facilitate pathogen monitoring and race identification. These results indicate the increasing virulence and variability in the Kenyan P. graminis f. sp. tritici population and reveal vulnerabilities of elite germplasm to new races.