Origin and genetic analysis of stem rust resistance in wheat line Tr129.

Wheat line Tr129 is resistant to stem rust, caused by Puccinia graminis f. sp. tritici (Pgt). The resistance in Tr129 was reportedly derived from Aegilops triuncialis, but the origin and genetics of resistance have not been confirmed. Here, genomic in situ hybridization (GISH) showed that no Ae. triuncialis chromatin was present in Tr129. Genetic and phenotypic analysis was conducted on F2 and DH populations from the cross RL6071/Tr129. Seedlings were tested with six Pgt races and were genotyped using an Illumina iSelect 90 K SNP array and kompetitive allele specific PCR (KASP) markers. Mapping and phenotyping showed that Tr129 carried four stem rust resistance (Sr) genes on chromosome arms 2BL (Sr9b), 4AL (Sr7b), 6AS (Sr8a), and 6DS (SrTr129). SrTr129 co-segregated with markers for SrCad, however Tr129 has a unique haplotype suggesting the resistance could be new. Analysis of a RL6071/Peace population revealed that like SrTr129, SrCad is ineffective against three North American races. This new understanding of SrCad will guide its use in breeding. Tr129 and the DNA markers reported here are useful resources for improving stem rust resistance in cultivars.

Localization of the Stem Rust Resistance Gene Pg2 to Linkage Group Mrg20 in Cultivated Oat (Avena sativa).

Stem rust is an important disease of cultivated oat (Avena sativa) caused by Puccinia graminis f. sp. avenae. In North America, host resistance is the primary strategy to control this disease and is conferred by a relatively small number of resistance genes. Pg2 is a widely deployed stem rust resistance gene that originates from cultivated oat. Oat breeders wish to develop cultivars with multiple Pg genes to slow the breakdown of single gene resistance, and often require DNA markers suited for marker-assisted selection. Our objectives were to (i) construct high density linkage maps for a major oat stem rust resistance gene using three biparental mapping populations, (ii) develop Kompetitive allele-specific PCR (KASP) assays for Pg2-linked single-nucleotide polymorphisms (SNPs), and (iii) test the prediction accuracy of those markers with a diverse panel of spring oat lines and cultivars. Genotyping-by-sequencing SNP markers linked to Pg2 were identified in an AC Morgan/CDC Morrison recombinant inbred line (RIL) population. Pg2-linked SNPs were then analyzed in an AC Morgan/RL815 F2 population and an AC Morgan/CDC Dancer RIL population. Linkage analysis identified a common location for Pg2 in all three populations on linkage group Mrg20 of the oat consensus genetic map. The most predictive markers were identified and converted to KASP assays for use in oat breeding programs. When used in combination, the KASP assays for the SNP loci avgbs2_126549.1.46 and avgbs_cluster_23819.1.27 were highly predictive of Pg2 status in panel of 54 oat breeding lines and cultivars.

Mapping of the stem rust resistance gene Pg13 in cultivated oat.

KEY MESSAGE: The widely deployed, oat stem rust resistance gene Pg13 was mapped by linkage analysis and association mapping, and KASP markers were developed for marker-assisted selection in breeding programs. Pg13 is one of the most extensively deployed stem rust resistance genes in North American oat cultivars. Identification of markers tightly linked to this gene will be useful for routine marker-assisted selection, identification of gene pyramids, and retention of the gene in backcrosses and three-way crosses. To this end, high-density linkage maps were constructed in four bi-parental mapping populations using SNP markers identified from 6K oat Infinium iSelect and genotyping-by-sequencing platforms. Additionally, genome-wide associations were identified using two sets of association panels consisting of diverse elite oat lines in one set and landrace accessions in the other. The results showed that Pg13 was located at approximately 67.7 cM on linkage group Mrg18 of the consensus genetic map. The gene co-segregated with the 7C-17A translocation breakpoint and with crown rust resistance gene Pc91. Co-segregating markers with the best prediction accuracy were identified at 67.7-68.5 cM on Mrg18. KASP assays were developed for linked SNP loci for use in oat breeding.

Characterization of genes required for both Rpg1 and rpg4-mediated wheat stem rust resistance in barley.

BACKGROUND: Puccinia graminis f. sp. tritici (Pgt) race TTKSK and its lineage pose a threat to barley production world-wide justifying the extensive efforts to identify, clone, and characterize the rpg4-mediated resistance locus (RMRL), the only effective resistance to virulent Pgt races in the TTKSK lineage. The RMRL contains two nucleotide-binding domain and leucine-rich repeat (NLR) resistance genes, Rpg5 and HvRga1, which are required for resistance. The two NLRs have head-to-head genome architecture with one NLR, Rpg5, containing an integrated C-terminal protein kinase domain, characteristic of an "integrated sensory domain" resistance mechanism. Fast neutron mutagenesis of line Q21861 was utilized in a forward genetics approach to identify genetic components that function in the RMRL or Rpg1 resistance mechanisms, as Q21861 contains both genes. A mutant was identified that compromises both RMRL and Rpg1-mediated resistances and had stunted seedling roots, designated required for P. graminis resistance 9 (rpr9). RESULTS: The rpr9 mutant generated in the Q21861 background was crossed with the Swiss landrace Hv584, which carries RMRL but contains polymorphism across the genome compared to Q21861. To map Rpr9, a Hv584 x rpr9 F6:7 recombinant inbred line (RIL) population was developed. The RIL population was phenotyped with Pgt race QCCJB. The Hv584 x rpr9 RIL population was genotyped with the 9 k Illumina Infinium iSelect marker panel, producing 2701 polymorphic markers. A robust genetic map consisting of 563 noncosegregating markers was generated and used to map Rpr9 to an ~ 3.4 cM region on barley chromosome 3H. The NimbleGen barley exome capture array was utilized to capture rpr9 and wild type Q21861 exons, followed by Illumina sequencing. Comparative analysis, resulting in the identification of a 1.05 Mbp deletion at the chromosome 3H rpr9 locus. The identified deletion contains ten high confidence annotated genes with the best rpr9 candidates encoding a SKP1-like 9 protein and a F-box family protein. CONCLUSION: Genetic mapping and exome capture rapidly identified candidate gene/s that function in RMRL and Rpg1 mediated resistance pathway/s. One or more of the identified candidate rpr9 genes are essential in the only two known effective stem rust resistance mechanisms, present in domesticated barley.

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.

Characterization of Sr9h, a wheat stem rust resistance allele effective to Ug99.

KEY MESSAGE: Wheat stem rust resistance gene SrWeb is an allele at the Sr9 locus that confers resistance to Ug99. Race TTKSK (Ug99) of Puccinia graminis f. sp. tritici, the causal fungus of stem rust, threatens global wheat production because of its broad virulence to current wheat cultivars. A recently identified Ug99 resistance gene from cultivar Webster, temporarily designated as SrWeb, mapped near the stem rust resistance gene locus Sr9. We determined that SrWeb is also present in Ug99 resistant cultivar Gabo 56 by comparative mapping and an allelism test. Analysis of resistance in a population segregating for both Sr9e and SrWeb demonstrated that SrWeb is an allele at the Sr9 locus, which subsequently was designated as Sr9h. Webster and Gabo 56 were susceptible to the Ug99-related race TTKSF+ from South Africa. Race TTKSF+ possesses unique virulence to uncharacterized Ug99 resistance in cultivar Matlabas. This result validated that resistance to Ug99 in Webster and Gabo 56 is conferred by the same gene: Sr9h. The emergence of pathogen virulence to several resistance genes that are effective to the original Ug99 race TTKSK, including Sr9h, suggests that resistance genes should be used in combinations in order to increase resistance durability.

Genetics and mapping of stem rust resistance to Ug99 in the wheat cultivar Webster.

New races of wheat stem rust, namely TTKSK (Ug99) and its variants, pose a threat to wheat production in the regions where they are found. The accession of the wheat cultivar Webster (RL6201) maintained at the Cereal Research Centre in Winnipeg, Canada, shows resistance to TTKSK and other races of stem rust. The purpose of this study was to study the inheritance of seedling resistance to stem rust in RL6201 and genetically map the resistance genes using microsatellite (SSR) markers. A population was produced by crossing the stem rust susceptible line RL6071 with Webster. The F(2) and F(3) were tested with TPMK, a stem rust race native to North America. The F(3) was also tested with TTKSK. Two independently assorting genes were identified in RL6201. Resistance to TPMK was conferred by Sr30, which was mapped with microsatellites on chromosome 5DL. The second gene, temporarily designated SrWeb, conferred resistance to TTKSK. SrWeb was mapped to chromosome 2BL using SSR markers. Comparison with previous genetic maps showed that SrWeb occupies a locus near Sr9. Further analysis will be required to determine if SrWeb is a new gene or an allele of a previously identified gene.

Effect of Barley yellow dwarf virus Infection on Yield and Malting Quality of Barley.

Barley yellow dwarf virus (BYDV) infection occurs frequently in barley in the Upper Midwest region of the United States; however, the impact of this disease on the yield and quality of malting cultivars has not been adequately addressed. Studies were conducted at Fargo, North Dakota (from 1989 to 1990) to determine the effect of BYDV infection on yield and malt quality parameters in barley. Three malting cultivars varying in yield potential and malting characteristics were artificially inoculated at the seedling stage with a North Dakota BYDV isolate of the PAV serotype. Overall yields were reduced 32.5 to 38% in 1989 and 8.5 to 19.8% in 1990 by BYDV infection. Thousand-kernel weight (3.2 to 14.9%) and kernel plumpness (11.9 to 38.9%) also were reduced. Kernel color and three malt quality parameters (α-amylase, malt recovery, and wort viscosity) were not affected by BYDV infection. Increases in wort protein (2.5 to 14.5%) and diastatic power (3.8 to 12.6%), and decreases in malt extract (1.1 to 5.6%) were found. Most notably, total protein increased 4.6 to 17.5% with BYDV infection. Higher proportions of thin seed, as a result of BYDV infection, contributed to the effects on several quality parameters. In summary, the negative effect of BYDV infection on overall quality is significant and should be considered when assessing the impact of this disease. Although annual BYD disease-related losses are typically not extensive in the Midwestern malting barley region, the incorporation of host resistance into recommended cultivars would ameliorate the negative effects of BYD disease on crop quality as well as on yield.

Rating Scales for Assessing Infection Responses of Barley Infected with Cochliobolus sativus.

Spot blotch, caused by Cochliobolus sativus, is a common foliar disease of barley that is controlled primarily through the deployment of resistant cultivars. Resistance is often assessed at the seedling and adult plant stages, but currently no comprehensive visual scale exists that describes the full spectrum of infection responses (IRs) occurring on barley. From the evaluation of a diverse collection of barley germ plasm and C. sativus isolates, a 1 to 9 IR scale was developed based on the type (presence of necrosis and chlorosis) and relative size of spot blotch lesions observed on the second leaves of barley seedlings. The nine IRs were classified into three general categories of low (IRs 1 to 3), intermediate (IRs 4 and 5), and high (IRs 6 to 9) host-parasite compatibility. Low IRs consisted of minute to small necrotic lesions with no or very slight diffuse marginal chlorosis. Intermediate IRs consisted of medium-sized necrotic lesions with a distinct but restricted chlorotic margin, while high IRs consisted of large necrotic lesions with distinct chlorotic margins and varying degrees of expanding diffuse chlorosis. In addition to the seedling IR scale, a four-class adult plant IR scale (R = resistant, MR = moderately resistant, MS = moderately susceptible, and S = susceptible) was developed based again on the type and relative size of lesions present on the leaves. These rating scales should be useful for many types of studies on spot blotch of barley.