G. J. Platz

Disease resistance genes in a doubled haploid population of two-rowed barley segregating for malting quality attributes

Malting barley (Hordeum vulgare L.) improvement involves selection for many quality traits, but the search for and deployment of resistance genes has continued to be an equally important endeavour. As an aid to phenotypic selection in breeding programs, gene mapping can serve to characterise genes known to exist in elite breeding lines. In the present study, 180 doubled haploid lines derived from the cross of VB9524/ND11231*12 were screened for disease resistance under field and greenhouse conditions. Quantitative trait locus (QTL) mapping and classical genetic linkage approaches were used to identify and map QTLs for resistance to powdery mildew (Blumeria graminis f.sp. hordei), net form of net blotch (Pyrenophora teres f. teres) and stem rust (Puccinia graminis f.sp. tritici). The analyses offered a comparison between QTL mapping and traditional genetic linkage analysis. Both approaches identified a QTL for powdery mildew resistance on chromosome 1H, which mapped to the approximate genomic location of the Mla6 gene. Similarly, both methods identified a major QTL for resistance to net form of net blotch on chromosome 6H and for stem rust resistance on chromosome 7H. The QTL for stem rust resistance on 7H mapped to the approximate location of the Rpg1 gene. Classical linkage analysis identified the 3 QTLs with major effects, but was unable to detect 3 other loci with minor effects.

Association mapping of resistance to Puccinia hordei in Australian barley breeding germplasm

Key message“To find stable resistance using association mapping tools, QTL with major and minor effects on leaf rust reactions were identified in barley breeding lines by assessing seedlings and adult plants.”AbstractThree hundred and sixty (360) elite barley (Hordeum vulgare L.) breeding lines from the Northern Region Barley Breeding Program in Australia were genotyped with 3,244 polymorphic diversity arrays technology markers and the results used to map quantitative trait loci (QTL) conferring a reaction to leaf rust (Puccinia hordei Otth). The F3:5 (Stage 2) lines were derived or sourced from different geographic origins or hubs of international barley breeding ventures representing two breeding cycles (2009 and 2011 trials) and were evaluated across eight environments for infection type at both seedling and adult plant stages. Association mapping was performed using mean scores for disease reaction, accounting for family effects using the eigenvalues from a matrix of genotype correlations. In this study, 15 QTL were detected; 5 QTL co-located with catalogued leaf rust resistance genes (Rph1, Rph3/19, Rph8/14/15, Rph20, Rph21), 6 QTL aligned with previously reported genomic regions and 4 QTL (3 on chromosome 1H and 1 on 7H) were novel. The adult plant resistance gene Rph20 was identified across the majority of environments and pathotypes. The QTL detected in this study offer opportunities for breeding for more durable resistance to leaf rust through pyramiding multiple genomic regions via marker-assisted selection.

An investigation of genetic variation among Australian isolates of Bipolaris sorokiniana from different cereal tissues and comparison of their abilities to cause spot blotch on barley

Bipolaris sorokiniana (teleomorph: Cochliobolus sativus), the causal agent of common rootrot (CRR) and foliar spot blotch (SB) diseases in barley and wheat, is an economically important fungal pathogen worldwide. However, the relationship between these two diseases is poorly understood. Differences within Australian B. sorokiniana populations were revealed by cluster analysis of amplified fragment length polymorphisms in genomic DNA of 48 B. sorokiniana isolates collected from the northern grain-growing region of Australia. Isolates collected from SB infections clustered apart from isolates collected from CRR infections. A subset of 31 B. sorokiniana isolates was assessed for their abilities to cause SB infections on barley leaves using a differential set of 15 barley genotypes and three other cereal species. The pathogen samples included 14 isolates from CRR infections of either wheat or barley and 14 isolates from SB infections of barley. Phenotypic experiments revealed that isolates of B. sorokiniana collected from barley SB infections showed a high level of pathogenic variability across the differential set. In contrast, isolates from CRR infections produced significantly less SB disease on inoculated barley leaves. Cluster analysis of the phenotypic infection response scores grouped isolates into three pathogenicity clusters demonstrating low, intermediate or high pathogenicity. The results of this study suggest divergence within Australian populations of B. sorokiniana in relation to host tissue specificity.

Pathotypes of Cochliobolus sativus on barley in Australia

Pathotypes of Cochliobolus sativus have been reported overseas but variation in the Australian population has not been investigated nor have the optimum conditions for the identification of variation been established. Preliminary experiments showed that maximum separation of infection responses of seedlings to infection by C. sativus conidia was obtained at 21°C using inocula containing 104 conidia/mL and a dew period of 40 h. Under these conditions, the existence of pathotypes in the Australian population of C. sativus was demonstrated. Using 20 lines, six pathotypes were identified among 34 isolates of the fungus. A differential set consisting of 12 barley cultivars is proposed.

Mapping of adult plant resistance to net form of net blotch in three Australian barley populations

Net form of net blotch (NFNB), caused by Pyrenophora teres Drechs. f. teres Smedeg., is a serious disease problem for the barley industry in Australia and other parts of the world. Three doubled haploid barley populations, Alexis/Sloop, WI2875-1/Alexis, and Arapiles/Franklin, were used to identify genes conferring adult plant resistance to NFNB in field trials. Quantitative trait loci (QTLs) identified were specific for adult plant resistance because seedlings of the parental lines were susceptible to the NFNB isolates used in this study. QTLs were identified on chromosomes 2H, 3H, 4H, and 7H in both the Alexis/Sloop and WI2875-1/Alexis populations and on chromosomes 1H, 2H, and 7H in the Arapiles/Franklin population. Using QTLNetwork, epistatic interactions were identified between loci on chromosomes 3H and 6H in the Alexis/Sloop population, between 2H and 4H in the WI2875-1/Alexis population, and between 5H and 7H in the Arapiles/Franklin population. Comparisons with earlier studies of NFNB resistance indicate the pathotype-dependent nature of many resistance QTLs and the importance of establishing an international system of pathotype nomenclature and differential testing.

Mating type locus-specific polymerase chain reaction markers for differentiation of Pyrenophora teres f. teres and P. teres f. maculata, the causal agents of barley net blotch.

Fourteen single nucleotide polymorphisms (SNPs) were identified at the mating type (MAT) loci of Pyrenophora teres f. teres (Ptt), which causes net form (NF) net blotch, and P. teres f. maculata (Ptm), which causes spot form (SF) net blotch of barley. MAT-specific SNP primers were developed for polymerase chain reaction (PCR) and the two forms were differentiated by distinct PCR products: PttMAT1-1 (1,143 bp) and PttMAT1-2 (1,421 bp) for NF MAT1-1 and MAT1-2 isolates; PtmMAT1-1 (194 bp) and PtmMAT1-2 (939 bp) for SF MAT1-1 and MAT1-2 isolates, respectively. Specificity was validated using 37 NF and 17 SF isolates collected from different geographic regions. Both MAT1-1 and MAT1-2 SNP primers retained respective specificity when used in duplex PCR. No cross-reactions were observed with DNA from P. graminea, P. tritici-repentis, or other ascomycetes, or barley. Single or mixed infections of the two different forms were also differentiated. This study provides the first evidence that the limited SNPs at the MAT locus are sufficient for distinguishing closely related heterothallic ascomycetes at subspecies levels, thus allowing pathogenicity and mating type characteristics of the fungus to be determined simultaneously. Methods presented will facilitate pathogen detection, disease management, and epidemiological studies.

Validation of a new spot form of net blotch differential set and evidence for hybridisation between the spot and net forms of net blotch in Australia

A recently developed spot form of blotch differential set of 16 barley lines was tested for reaction response to 60 Pyrenophora teres f. maculata isolates from geographically disperse barley crops of Australia. Twelve barley lines (Arimont, Barque, Chebec, CI5286, CI5791, CI9214, CII6150, Dairokkaku, Esperance Orge 289, Galleon, Keel, Skiff, Torrens and TR250) provided differential response between the isolates. The susceptible controls Gairdner and Kombar provided indication of isolate virulence or avirulence. Abundant pathogenic diversity was revealed with 33 designated pathotypes, some of which related to geographic region. AFLP analysis also revealed abundant diversity with each of the isolates representing a unique genotype and one isolate that contained both AFLP bands unique to P. teres f. maculata and P. teres f. teres, the cause of spot form and net form of net blotch respectively, suggesting that sexual recombination between the net form and spot form isolates may have occurred naturally in the field.

Fungicidal control of Pyrenophora tritici-repentis in wheat

The use of fungicides to control yellow spot in wheat, caused by Pyrenophora tritici-repentis, in the northern wheat region of Australia, was examined in three field experiments. Field conditions were conducive to disease development. This was aided by spreading stubble infested with P. tritici-repentis and by regular spray irrigations, and moderate yellow spot epidemics resulted in all 3 years. Within the three experiments, 12 fungicides were evaluated for their ability to reduce the impact of yellow spot on grain yield. Single spray applications of the fungicides, some at varying rates, were made at 90% flag leaf emergence. Treated controls had 3–4 applications of propiconazole at 62 mL a.i./ha or tebuconazole at 72.5 mL a.i./ha throughout the season. Grain yield losses of up to 59% were recorded in the untreated plots. A reduction in grain size rather than a decrease in the number of grains was responsible for the yield losses. Yield losses were significantly lower following application of flusilazol, prochloraz, propiconazole and tebuconazole in cw. Hartog and Banks, in the first experiment. Application of azoxystrobin at 125 and 250 g a.i./ha, difenoconazole at 125 mL a.i./ha, epoxyconizole at 72 mL a.i./ha, propiconazole at 125 mL a.i./ha, tebuconazole at 125 mL a.i./ha or trifloxystrolin at 150g a.i./ha significantly reduced losses in the third experiment, sown with cv. Sunstate. This work has demonstrated that fungicidal control of yellow spot in wheat in the northern region can be a viable control option in years of high yield potential and high disease pressure.

Spot form of net blotch resistance in barley is under complex genetic control

Key messageEvaluation of resistance toPyrenophora teresf.maculatain barley breeding populations via association mapping revealed a complex genetic architecture comprising a mixture of major and minor effect genes.AbstractIn the search for stable resistance to spot form of net blotch (Pyrenophora teres f. maculata, SFNB), association mapping was conducted on four independent barley (Hordeum vulgare L.) breeding populations comprising a total of 898 unique elite breeding lines from the Northern Region Barley Breeding Program in Australia for discovery of quantitative trait loci (QTL) influencing resistance at seedling and adult plant growth stages. A total of 29 significant QTL were validated across multiple breeding populations, with 22 conferring resistance at both seedling and adult plant growth stages. The remaining 7 QTL conferred resistance at either seedling (2 QTL) or adult plant (5 QTL) growth stages only. These 29 QTL represented 24 unique genomic regions, of which five were found to co-locate with previously identified QTL for SFNB. The results indicated that SFNB resistance is controlled by a large number of QTL varying in effect size with large effects QTL on chromosome 7H. A large proportion of the QTL acted in the same direction for both seedling and adult responses, suggesting that phenotypic selection for SFNB resistance performed at either growth stage could achieve adequate levels of resistance. However, the accumulation of specific resistance alleles on several chromosomes must be considered in molecular breeding selection strategies.

Characterisation of Rph24: A gene conferring adult plant resistance to Puccinia hordei in barley

We identified Rph24 as a locus in barley (Hordeum vulgare L.) controlling adult plant resistance (APR) to leaf rust, caused by Puccinia hordei. The locus was previously reported as a quantitative trait locus in barley line ND24260-1 and named qRphND. We crossed ND24260-1 to the leaf-rust-susceptible standard Gus and determined inheritance patterns in the progeny. For the comparative marker frequency analysis (MFA), resistant and susceptible tails of the F2 were genotyped with Diversity Arrays Technology genotyping-by-sequencing (DArT-Seq) markers. The Rph24 locus was positioned at 55.5 centimorgans on chromosome 6H on the DArT-Seq consensus map. Evaluation of F2:3 families confirmed that a single locus from ND24260-1 conferred partial resistance. The haploblock strongly associated with the Rph24 locus was used to estimate the allele frequency in a collection of 282 international barley cultivars. Rph24 was frequently paired with APR locus Rph20 in cultivars displaying high levels of APR to leaf rust. The markers identified in this study for Rph24 should be useful for marker-assisted selection.