Manisha Shankar

Quantitative trait loci for seedling and adult plant resistance to Stagonospora nodorum in wheat

Stagonospora nodorum blotch (SNB) caused by Stagonospora nodorum is a severe disease of wheat (Triticum aestivum) in many areas of the world. S. nodorum affects both seedling and adult plants causing necrosis of leaf and glume tissue, inhibiting photosynthetic capabilities, and reducing grain yield. The aims of this study were to evaluate disease response of 280 doubled haploid (DH) individuals derived from a cross between resistant (6HRWSN125) and susceptible (WAWHT2074) genotypes, compare quantitative trait loci (QTL) for seedling and adult plant resistance in two consecutive years, and assess the contribution of QTL on grain weight. Flag leaves and glumes of individuals from the DH population were inoculated with mixed isolates of S. nodorum at similar maturity time to provide accurate disease evaluation independent of morphological traits and identify true resistance for QTL analysis. Fungicide protected and inoculated plots were used to measure relative grain weight (RGW) as a yield-related trait under pathogen infection. The lack of similar QTL and little or no correlation in disease scores indicate different genes control seedling and adult plant disease and independent genes control flag leaf and glume resistance. This study consistently identified a QTL on chromosome 2DL for flag leaf resistance (QSnl.daw-2D) and 4BL for glume resistance (QSng.daw-4B) from the resistant parent, 6HRWSN125, explaining 4 to 19% of the phenotypic variation at each locus. A total of 5 QTL for RGW were consistently detected, where two were in the same marker interval for QSnl.daw-2D and QSng.daw-4B indicating the contribution of these QTL to yield related traits. Therefore, RGW measurement in QTL analysis could be used as a reliable indicator of grain yield affected by S. nodorum infection.

Ubiquity of ToxA and absence of ToxB in Australian populations of Pyrenophora tritici-repentis.

Pyrenophora tritici-repentis, the causal organism of the necrotrophic foliar wheat disease tan spot [also known as yellow (leaf) spot in Australia] is an important disease in Australia and in many parts of the world. North American isolates of the pathogen have been shown to produce combinations of three host-specific toxins, ToxA, ToxB and ToxC. Each toxin interacts with a host sensitivity locus, respectively Tsn1, Tsc2 and Tsc1. The virulence of an isolate is partially correlated with the presence of these toxins and resistance in the host is associated with absence of the sensitivity loci. Breeding for resistance to tan spot can, therefore, be aided by knowledge of the prevalence of the toxin-encoding genes in local pathogen populations. Two of the toxins, A and B, are encoded by known genes and molecular tests for the genes have been developed. We screened a diverse collection of 119 tan spot isolates collected between 1984 and 2008 and from all affected regions of Australia (Queensland, New South Wales, Victoria and Western Australia). In all cases, the gene for ToxA was present and the gene for ToxB was absent. The implications for resistance breeding and epidemiology of the disease are discussed. We also define a diagnostic molecular marker for P. tritici-repentis.

Mining synthetic hexaploids for multiple disease resistance to improve bread wheat

A collection of 253 synthetic hexaploid wheats (SHWs) produced from 192 Aegilops tauschii accessions and 39 elite durum varieties were studied to identify, characterise, and evaluate potentially untapped diversity of disease resistance in wheat. The diseases for which resistance was sought included cereal cyst nematode (CCN), root lesion nematode (RLN), Stagonospora nodorum blotch (SNB), Septoria tritici blotch (STB), and the 3 rusts, leaf rust, stem rust, and stripe rust, all important diseases of bread wheat worldwide, which can severely reduce wheat yield and quality. The SHWs exhibited a wide spectrum of resistance to the 8 pathogens. The frequency of disease-resistant SHWs ranged from 1% for one species of RLN (Pratylenchus neglectus), 3% and 10% for Septoria nodorum leaf and glume blotch, 10% for seedling resistance to yellow leaf spot, 16% for CCN, 21% for the second species of RLN (Pratylenchus thornei), 73% for Septoria tritici blotch, and 15%, 40%, and 24% for leaf rust, stem rust, and stripe rust, respectively. Five SHWs, Aus26860, Aus30258, Aus30294, Aus30301, and Aus30304, exhibited high levels of resistance to CCN, YLP, STB, LR, and SR, while 56 SHWs showed resistance to either 3 or 4 diseases. The genetics of resistance to CCN in some of the SHWs revealed that some of the accessions carry the same CCN gene(s) against pathotype Ha13, while others may carry different resistance gene(s). Additional studies were carried out to understand the relationship between the resistances identified in SHWs and the ones already present in common wheat, in particular the resistance genes Cre1 and Cre3 against CCN. The use of perfect markers associated with Cre1 and Cre3 suggested that some SHWs may carry a new CCN resistance gene(s), which could be deployed in breeding programs to increase the diversity of available resistance. The identification of SHWs with resistance to a range of diseases provides an opportunity to generate genetic knowledge and resistant germplasm to be used in future variety development.

Identification of alleles at two loci controlling resistance to Phomopsis stem blight in narrow-leafed lupin (Lupinus angustifolius L.)

The genetics of resistance to Phomopsis stem blight caused by Diaporthe toxica Will., Highet, Gams & Sivasith. in narrow-leafed lupin (Lupinus angustifolius L.) was studied in crosses between resistant cv. Merrit, very resistant breeding line 75A:258 and susceptible cv. Unicrop. A non-destructive glasshouse infection test was developed to assess resistance in the F1, F2, selected F2-derived F3 (F2:3) families, and in selfed parent plants. The F1 of Unicrop × 75A:258 (and reciprocal cross) was very resistant, and the F2 segregated in a ratio of 3:1 (resistant: susceptible), which suggested the presence of a single dominant allele for resistance in 75A:258. In Merrit × Unicrop (and reciprocal), the F1 was moderately resistant, and the F2 segregated in a ratio of 3:1 (resistant: susceptible). Thus Merrit appeared to carry an incompletely dominant resistance allele for resistance. The F1 of Merrit × 75A:258 (and reciprocal) was very resistant and the F2 segregated in a ratio of 15:1 (resistant: susceptible), which supported the existence of independently segregating resistance alleles for resistance in 75A:258 and Merrit. Alleles at loci for early flowering (Ku) and speckled seeds (for which we propose the symbol Spk) segregated normally and independently of the resistance alleles. Resistant F2 plants gave rise to uniformly resistant or segregating F2:3 families, whereas susceptible F2 plants gave rise only to susceptible F2:3 families. However, the variation in resistance in the F2 and some F2:3 families of crosses involving 75A:258, from moderately to extremely resistant, was greater than that expected by chance or environmental variation. We propose the symbols Phr1 to describe the dominant resistance allele in 75A:258, and Phr2 for the incompletely dominant resistance allele in Merrit. Phr1 appears to be epistatic to Phr2, and expression of Phr1 may be altered by independently segregating modifier allele(s).

Loci on chromosomes 1A and 2A affect resistance to tan (yellow) spot in wheat populations not segregating for tsn1

Key messageQTL for tan spot resistance were mapped on wheat chromosomes 1A and 2A. Lines were developed with resistance alleles at these loci and at the tsn1 locus on chromosome 5B. These lines expressed significantly higher resistance than the parent withtsn1only.AbstractTan spot (syn. yellow spot and yellow leaf spot) caused by Pyrenophora tritici-repentis is an important foliar disease of wheat in Australia. Few resistance genes have been mapped in Australian germplasm and only one, known as tsn1 located on chromosome 5B, is known in Australian breeding programs. This gene confers insensitivity to the fungal effector ToxA. The main aim of this study was to map novel resistance loci in two populations: Calingiri/Wyalkatchem, which is fixed for the ToxA-insensitivity allele tsn1, and IGW2574/Annuello, which is fixed for the ToxA-sensitivity allele Tsn1. A second aim was to combine new loci with tsn1 to develop lines with improved resistance. Tan spot severity was evaluated at various growth stages and in multiple environments. Symptom severity traits exhibited quantitative variation. The most significant quantitative trait loci (QTL) were detected on chromosomes 2A and 1A. The QTL on 2A explained up to 29.2% of the genotypic variation in the Calingiri/Wyalkatchem population with the resistance allele contributed by Wyalkatchem. The QTL on 1A explained up to 28.1% of the genotypic variation in the IGW2574/Annuello population with the resistance allele contributed by Annuello. The resistance alleles at both QTL were successfully combined with tsn1 to develop lines that express significantly better resistance at both seedling and adult plant stages than Calingiri which has tsn1 only.

Discovering new alleles for yellow spot resistance in the Vavilov wheat collection

Key messageGWAS detected 11 yellow spot resistance QTL in the Vavilov wheat collection. Promising adult-plant resistance loci could provide a sustainable genetic solution to yellow spot in modern wheat varieties.AbstractYellow spot, caused by the fungal pathogen Pyrenophora tritici-repentis (Ptr), is the most economically damaging foliar disease of wheat in Australia. Genetic resistance is considered to be the most sustainable means for disease management, yet the genomic regions underpinning resistance to Ptr, particularly adult-plant resistance (APR), remain vastly unknown. In this study, we report results of a genome-wide association study using 295 accessions from the Vavilov wheat collection which were extensively tested for response to Ptr infections in glasshouse and field trials at both seedling an adult growth stages. Combining phenotypic datasets from multiple experiments in Australia and Russia with 25,286 genome-wide, high-quality DArTseq markers, we detected a total of 11 QTL, of which 5 were associated with seedling resistance, 3 with all-stage resistance, and 3 with APR. Interestingly, the novel APR QTL were effective even in the presence of host sensitivity gene Tsn1. These genomic regions could offer broad-spectrum yellow spot protection, not just to ToxA but also other pathogenicity or virulence factors. Vavilov wheat accessions carrying APR QTL combinations displayed enhanced levels of resistance highlighting the potential for QTL stacking through breeding. We propose that the APR genetic factors discovered in our study could be used to improve resistance levels in modern wheat varieties and contribute to the sustainable control of yellow spot.

Genetic Relationship of Stripe Rust Resistance Genes Yr34 and Yr48 in Wheat and Identification of Linked KASP Markers

The Australian continent was free from wheat stripe rust caused by Puccinia striiformis f. sp. tritici until exotic incursions occurred in 1979 and 2002. The 2002 incursion enabled the identification of a new stripe rust resistance gene (Yr34) in the advanced breeding line WAWHT2046. In this study, we developed and validated markers closely linked with Yr34, which is located in the distal region in the long arm of chromosome 5A. Four kompetitive allele-specific polymerase chain reaction (KASP) and three sequence-tagged site (STS) markers derived from the International Wheat Genome Sequencing Consortium RefSeq v1.0 scaffold-77836 cosegregated with Yr34. Markers sun711, sun712, sun725, sunKASP_109, and sunKASP_112 were shown to be suitable for marker-assisted selection in a validation panel of 71 Australian spring wheat genotypes, with the exception of cultivar Orion that carried the Yr34-linked alleles for sunKASP_109 and sunKASP_112. Markers previously reported to be linked with adult plant stripe rust resistance gene Yr48 also cosegregated with Yr34. Wheat genotypes carrying Yr34 and Yr48 produced identical haplotypes for the Yr34-linked markers identified in this study and those previously reported to be linked with Yr48. Phenotypic testing of genotypes carrying Yr34 and Yr48 showed that both genes conferred similar seedling responses to pre-2002 and post-2002 P. striiformis f. sp. tritici pathotypes. Further testing of 600 F2 plants from a cross between WAWHT2046 and RIL143 (Yr48) with P. striiformis f. sp. tritici pathotype 134 E16A+Yr17+Yr27+ failed to reveal any susceptible segregants. Our results strongly suggest that Yr34 and Yr48 are the same gene, and that Yr48 should be considered a synonym of Yr34.

Responses of commercial wheat varieties and differential lines to western Australian powdery mildew (Blumeria graminis f. Sp. tritici) populations

Powdery mildew, caused by Blumeria graminis f. sp. tritici, is an important disease of bread wheat (Triticum aestivum L.). Little is known about the powdery mildew population structure and the economic loss that occurs on commercial wheat cultivars in Western Australia. Eighteen wheat lines (including sixteen commercial wheat varieties, line 7HRWSN58 and a susceptible control) and a set of differential lines with known powdery mildew (Pm) resistant genes were evaluated using a cumulative pool of powdery mildew (Pm) inoculum each year to examine the changes in their responses over the years. Trials with a randomised block design were conducted under controlled environment over the years 2011 to 2014. Resistance genes Pm2, Pm3a, Pm3e, Pm4a, Pm13 and Pm27 were postulated to be effective in WA while genes Pm1a, Pm3c, Pm4b, Pm5a, Pm7, Pm17, Pm24 and Pm28 were ineffective. Differential lines carrying Pm6 and Pm8 genes were moderately resistant at adult plant stage but susceptible at seedling stage. Twelve out of sixteen commercial wheat varieties were susceptible or highly susceptible at seedling and adult plant stages lacking gene/s for resistance to powdery mildew. Most of these varieties were stable but some of the differential lines showed a breakdown in resistance over the four years. The cultivars Fortune, Magenta and Yitpi were moderately susceptible (MS) at adult plant stage but susceptible (S) at the seedling stage. The variety Arrino and line 7HRWSN58 were moderately resistant (MR) and resistant (R) to powdery mildew respectively.

Germplasm Enhancement for Resistance to Pyrenophora tritici-repentis in Wheat

Yellow spot (syn. tan spot), caused by Pyrenophora tritici-repentis, is an important foliar disease of wheat in Australia that causes losses exceeding 50 % when conditions are favourable for disease development. Although good progress has been made internationally to understand yellow spot resistance, relatively few resistance genes have been identified and mapped in Australian germplasm and only one (tsn1 on chromosome 5BL) is in general and known use in Australian breeding programs. Although tsn1 is an important yellow spot resistance gene, it doesn’t explain the full spectrum of resistance and there is a significant opportunity to enhance expression of yellow spot resistance through identification of resistance factors other than tsn1. Six doubled haploid (DH) mapping populations (five of which were fixed for tsn1) were screened for yellow spot resistance at the seedling/tillering and adult plant stages at the Department of Agriculture and Food, Western Australia (DAFWA) and the Department of Environment and Primary Industries Victoria (DEPIVic) from 2009 to 2012. Four of the above populations were screened at the Department of Agriculture, Fisheries and Forestry Queensland (DAFFQ). Frequency distribution of individuals within each population for various levels of yellow spot resistance was continuous indicating that resistance is conditioned by several genes with partial effects. A few lines within each population consistently showed high levels of resistance probably resulting from a combination of several genes with additive effects. Nine new loci for yellow spot resistance were mapped by the Australian Wheat and Barley Molecular Marker Program (AWBMMP) at the University of Adelaide on chromosomes 1AS, 2AS, 5AS, 5AL, 4B, 6BS, 7BL, 2D and 7D in five of the six populations phenotyped so far. High LOD scores of 9–11 have been obtained for some of the QTL with a percentage disease reduction of 24–42 %. Efforts are now focused on identifying additional yellow spot resistance genes using newly developed populations and rapid phenotyping methods and developing a series of fixed lines, each carrying yellow spot resistance genes from various sources, in elite Australian backgrounds. These materials will provide proof-of-concept for achieving better resistance by pyramiding resistance genes, and they should be directly useful as parents for wheat breeding.