Anke Martin

Assessment of Infection by Fusarium pseudograminearum in Wheat Seedling Tissues Using Quantitative PCR and a Visual Discoloration Scale

Assessment among cereal genotypes of relative seedling resistance to the crown rot pathogen Fusarium pseudograminearum has been primarily based on visual discoloration of the leaf sheaths. This study is the first to investigate the relationship between the widely used visual rating of seedling leaf sheath discoloration and the degree of colonization of these tissues by the pathogen, based on quantitative polymerase chain reaction (qPCR) of fungal DNA using primers specific for the translation elongation factor α sequence. Fourteen-day-old seedlings of four hard white spring wheat genotypes which differ in their degree of resistance to the pathogen, based on the expression of visible symptoms, were inoculated using a droplet method and assessed weekly from 7 to 35 days after inoculation (dai) for both discoloration and fungal DNA content per unit of tissue weight. Both visual assessment of disease symptoms and qPCR of fungal biomass indicated significant differences between the partially resistant and susceptible wheat genotypes from 14 dai. Visual discoloration of leaf sheath tissues was strongly correlated with fungal biomass estimated by qPCR in all four genotypes; however, this correlation became weaker with increasing time after inoculation. Significant correlations between these parameters were indicated at 14, 21, and 28 dai whereas, by 35 dai, the correlation was not significant. Evaluation of plants at 14 dai provided a rapid test which gave clear discrimination between lines for both parameters and was the time point of closest correlation between fungal colonization and disease symptoms. Symptom expression at all times following inoculation was accompanied by tissue infection, and at no time was symptomless infection observed under this screening environment. These qPCR results confirm that visual assessments of disease symptoms reflect the extent of tissue colonization by the pathogen in recently colonized tissues and confirm the validity of visual assessments for disease rating in high-throughput screening of breeding materials.

Pentaploid Wheat Hybrids: Applications, Characterisation, and Challenges

Interspecific hybridisation between hexaploid and tetraploid wheat species leads to the development of F1 pentaploid hybrids with unique chromosomal constitutions. Pentaploid hybrids derived from bread wheat (Triticum aestivum L.) and durum wheat (Triticum turgidum spp. durum Desf.) crosses can improve the genetic background of either parent by transferring traits of interest. The genetic variability derived from bread and durum wheat and transferred into pentaploid hybrids has the potential to improve disease resistance, abiotic tolerance, and grain quality, and to enhance agronomic characters. Nonetheless, pentaploid wheat hybrids have not been fully exploited in breeding programs aimed at improving crops. There are several potential barriers for efficient pentaploid wheat production, such as low pollen compatibility, poor seed set, failed seedling establishment, and frequent sterility in F1 hybrids. However, most of the barriers can be overcome by careful selection of the parental genotypes and by employing the higher ploidy level genotype as the maternal parent. In this review, we summarize the current research on pentaploid wheat hybrids and analyze the advantages and pitfalls of current methods used to assess pentaploid-derived lines. Furthermore, we discuss current and potential applications in commercial breeding programs and future directions for research into pentaploid wheat.

QTL associated with barley (Hordeum vulgare) feed quality traits measured through in situ digestion

Barley (Hordeum vulgare) is a major feed source for the intensive livestock industry. Competitiveness against other cereal grains depends largely on the price per unit of expressed feed quality. The traits which contribute to feed quality in barley are largely quantitative in nature but little is known about their genetic control. A study to identify the quantitative trait loci (QTLs) associated with feed quality was performed using a F6-derived recombinant inbred barley population. Samples from each line were incubated in the rumen of fistulated cattle, recovered, washed and dried for determination of in situ dry matter digestibility. Additionally, both pre- and post-digestion samples were analysed to quantify the content of key quality components relating to acid detergent fibre, total starch and protein. The data was used to identify trait-associated QTLs. Genetic analysis identified significant QTLs on chromosomes 2H, 5H and 7H. Genetic markers linked to these QTL should provide an effective tool for the selection and improvement of feed barley in the future.

Effects of Elevated Carbon Dioxide on Photosynthesis and Carbon Partitioning: A Perspective on Root Sugar Sensing and Hormonal Crosstalk

Plant responses to atmospheric carbon dioxide will be of great concern in the future, as carbon dioxide concentrations ([CO2]) are predicted to continue to rise. Elevated [CO2] causes increased photosynthesis in plants, which leads to greater production of carbohydrates and biomass. Which organ the extra carbohydrates are allocated to varies between species, but also within species. These carbohydrates are a major energy source for plant growth, but they also act as signaling molecules and have a range of uses beyond being a source of carbon and energy. Currently, there is a lack of information on how the sugar sensing and signaling pathways of plants are affected by the higher content of carbohydrates produced under elevated [CO2]. Particularly, the sugar signaling pathways of roots are not well understood, along with how they are affected by elevated [CO2]. At elevated [CO2], some plants allocate greater amounts of sugars to roots where they are likely to act on gene regulation and therefore modify nutrient uptake and transport. Glucose and sucrose also promote root growth, an effect similar to what occurs under elevated [CO2]. Sugars also crosstalk with hormones to regulate root growth, but also affect hormone biosynthesis. This review provides an update on the role of sugars as signaling molecules in plant roots and thus explores the currently known functions that may be affected by elevated [CO2].

Rare pyrenophora teres hybridization events revealed by development of sequence-specific PCR markers

Pyrenophora teres f. teres and P. teres f. maculata cause net form and spot form, respectively, of net blotch on barley (Hordeum vulgare). The two forms reproduce sexually, producing hybrids with genetic and pathogenic variability. Phenotypic identification of hybrids is challenging because lesions induced by hybrids on host plants resemble lesions induced by either P. teres f. teres or P. teres f. maculata. In this study, 12 sequence-specific polymerase chain reaction markers were developed based on expressed regions spread across the genome. The primers were validated using 210 P. teres isolates, 2 putative field hybrids (WAC10721 and SNB172), 50 laboratory-produced hybrids, and 7 isolates collected from barley grass (H. leporinum). The sequence-specific markers confirmed isolate WAC10721 as a hybrid. Only four P. teres f. teres markers amplified on DNA of barley grass isolates. Amplified fragment length polymorphism markers suggested that P. teres barley grass isolates are genetically different from P. teres barley isolates and that the second putative hybrid (SNB172) is a barley grass isolate. We developed a suite of markers which clearly distinguish the two forms of P. teres and enable unambiguous identification of hybrids.

Genome inheritance in populations derived from hexaploid/tetraploid and tetraploid/hexaploid wheat crosses

Hexaploid/tetraploid and tetraploid/hexaploid wheat hybrids were established using the hexaploid (Triticum aestivum L.) bread wheat LRC2010-150 and the tetraploid durum wheat (T. turgidum spp. durum) WID802. Thirty F2 progeny from each cross were characterised using Diversity Arrays Technology (DArTseq™) markers to determine whether there are differences between the crosses in the proportion of A, B and D genomic material inherited from each parent. Inheritance of the A and B genome from the tetraploid durum parent varied from 32 to 63% among the 60 lines assessed, and results indicated significant differences between the two F2 populations in the mean overall proportion of chromosomes A and B inherited from each parent. Significant differences were also observed between the crosses in the proportion of chromosomal segments on 2B, 3A, 3B and 4A inherited from the tetraploid parent. The F2 populations also showed significant differences in the average retention of D chromosomes per line with the tetraploid/hexaploid cross retaining a mean of 2.83 chromosomes while the reciprocal cross retained a mean of 1.8 chromosomes per line. A strong negative correlation was observed in individual lines from both populations between the proportion of the A and B genome inherited from the tetraploid durum parent and the retention of the D genome. The implication of these results for the design of efficient crossing strategies between hexaploid and tetraploid wheats is discussed.

A cytological and molecular analysis of D-genome chromosome retention following F2–F6 generations of hexaploid × tetraploid wheat crosses

Abstract. Both hexaploid bread wheat (AABBDD) (Triticum aestivum L.) and tetraploid durum wheat (AABB) (T. turgidum spp. durum) are highly significant global food crops. Crossing these two wheats with different ploidy levels results in pentaploid (AABBD) F1 lines. This study investigated the differences in the retention of D chromosomes between different hexaploid × tetraploid crosses in subsequent generations by using molecular and cytological techniques. Significant differences (P < 0.05) were observed in the retention of D chromosomes in the F2 generation depending on the parents of the original cross. One of the crosses, 2WE25 × 950329, retained at least one copy of each D chromosome in 48% of its F2 lines. For this cross, the retention or elimination of D chromosomes was determined through several subsequent self-fertilised generations. Cytological analysis indicated that D chromosomes were still being eliminated at the F5 generation, suggesting that in some hexaploid × tetraploid crosses, D chromosomes are unstable for many generations. This study provides information on the variation in D chromosome retention in different hexaploid × tetraploid wheat crosses and suggests efficient strategies for utilising D genome retention or elimination to improve bread and durum wheat, respectively.

Transposable Element Genomic Fissuring in Pyrenophora teres Is Associated With Genome Expansion and Dynamics of Host–Pathogen Genetic Interactions

Pyrenophora teres, P. teres f. teres (PTT) and P. teres f. maculata (PTM) cause significant diseases in barley, but little is known about the large-scale genomic differences that may distinguish the two forms. Comprehensive genome assemblies were constructed from long DNA reads, optical and genetic maps. As repeat masking in fungal genomes influences the final gene annotations, an accurate and reproducible pipeline was developed to ensure comparability between isolates. The genomes of the two forms are highly collinear, each composed of 12 chromosomes. Genome evolution in P. teres is characterized by genome fissuring through the insertion and expansion of transposable elements (TEs), a process that isolates blocks of genic sequence. The phenomenon is particularly pronounced in PTT, which has a larger, more repetitive genome than PTM and more recent transposon activity measured by the frequency and size of genome fissures. PTT has a longer cultivated host association and, notably, a greater range of host–pathogen genetic interactions compared to other Pyrenophora spp., a property which associates better with genome size than pathogen lifestyle. The two forms possess similar complements of TE families with Tc1/Mariner and LINE-like Tad-1 elements more abundant in PTT. Tad-1 was only detectable as vestigial fragments in PTM and, within the forms, differences in genome sizes and the presence and absence of several TE families indicated recent lineage invasions. Gene differences between P. teres forms are mainly associated with gene-sparse regions near or within TE-rich regions, with many genes possessing characteristics of fungal effectors. Instances of gene interruption by transposons resulting in pseudogenization were detected in PTT. In addition, both forms have a large complement of secondary metabolite gene clusters indicating significant capacity to produce an array of different molecules. This study provides genomic resources for functional genetics to help dissect factors underlying the host–pathogen interactions.

Association between presence of Triticum timopheevii introgression and D-genome retention in hexaploid/tetraploid wheat crosses

The 2G Triticum timopheevii introgression harbours genes for multiple disease resistance and quality traits in bread wheat. In order to transfer this segment from bread wheat into durum, the bread wheat line Sunguard, which carries this introgressed 2G segment was crossed with three tetraploid durum parents. A significant difference was observed in the segregation ratio of the 2G segment in the different crosses at the F2 generation with two of the three populations indicating segregation distortion against the hexaploid 2G segment. In these populations, the presence of the 2G segment was strongly correlated with the presence of D-genome chromosomes. These results were confirmed in the F4 generation of these populations. Six plants were identified in the F4 generation, which had retained the introgressed 2G segment in a homozygous condition and did not have a complete D-genome set. Two of these lines only had two non-homologous D-genome chromosomes in the F5 generation. Thus, the 2G segment and possibly other translocations can be transferred into durum wheat through hexaploid/tetraploid hybridisation.

Identification and mapping of net form of net blotch resistance in South African barley

Net form of net blotch (NFNB) caused by the fungus Pyrenophora teres f. teres is an economically important foliar disease of barley (Hordeum vulgare) in southern and eastern Africa. Little attention has been given to disease resistance breeding, and knowledge about the presence of NFNB resistance in breeding lines is limited. Deploying resistance into varieties used in this region is important for future control of the disease. We have identified NFNB disease resistance in existing South African breeders’ lines and have mapped the resistance in line UVC8. Six different trials, three conducted in South Africa and another three in Australia, were used to identify resistance QTL. A major QTL was identified on chromosome 6H having a LOD score of 40.5 and 55% of the phenotypic variance explained. Kompetitive Allele Specific PCR (KASP™) markers were designed for this QTL region. These and microsatellite markers can now be used to routinely select for NFNB resistance.