First reported sexual recombination between Pyrenophora teres isolates from barley and barley grass.

Barley grass (Hordeum leporinum), which often occurs in proximity to commercial barley (Hordeum vulgare) cultivars, is an alternative host to Pyrenophora teres, an economically important pathogen causing net blotch in barley. This study is the first to report the sexual recombination of P. teres isolates collected from barley with those collected from barley grass. The sexual recombination between P. teres isolates from barley and barley grass was confirmed using a neighbour-net network and haploblock plots based on whole genome sequencing of seven progeny isolates. Pathogenicity assays revealed that P. teres isolates from barley grass were not host specific and could infect both barley and barley grass and the progeny isolates were virulent on commercially grown barley cultivars. Our results contradict previous population and pathogenicity studies of P. teres isolates obtained from barley and barley grass which have reported that the two populations are genetically distinct and host specific, suggesting that isolates collected from barley or barley grass could be two different entities. Despite the genetic divergence of P. teres isolates from barley and barley grass revealed through our phylogenomic analysis, there seems to be no complete host or reproductive separation between these populations. Therefore, there is a potential for generation of novel pathotypes through sexual recombination between P. teres isolates associated with barley and barley grass, with a risk of increased impacts on commercial barley cultivars that do not carry resistance to these pathotypes.

Virulence Spectra of Hungarian Pyrenophora teres f. teres Isolates Collected from Experimental Fields Show Continuous Variation without Specific Isolate × Barley Differential Interactions.

Pyrenophora teres f. teres (Ptt), the causal agent of net form net blotch (NFNB) disease, is an important and widespread pathogen of barley. This study aimed to quantify and characterize the virulence of Ptt isolates collected from experimental fields of barley in Hungary. Infection responses across 20 barley differentials were obtained from seedling assays of 34 Ptt isolates collected from three Hungarian breeding stations between 2008 and 2018. Twenty-eight Ptt pathotypes were identified. Correspondence analysis followed by hierarchical clustering on the principal components and host-by-pathogen GGE biplots suggested a continuous range of virulence and an absence of specific isolate × barley differential interactions. The isolates were classified into four isolate groups (IG) using agglomerative hierarchical clustering. One IG could be distinguished from other IGs based on avirulence/virulence on one to five barley differentials. Several barley differentials expressed strong resistance against multiple Ptt isolates and may be useful in the development of NFNB-resistant barley cultivars in Hungary. Our results emphasize that the previously developed international barley differential set needs to be improved and adapted to the Hungarian Ptt population. This is the first report on the pathogenic variations of Ptt in Hungary.

Multi-parental fungal mapping population study to detect genomic regions associated with Pyrenophora teres f. teres virulence.

In recent years multi-parental mapping populations (MPPs) have been widely adopted in many crops to detect quantitative trait loci (QTLs) as this method can compensate for the limitations of QTL analyses using bi-parental mapping populations. Here we report the first multi-parental nested association mapping (MP-NAM) population study used to detect genomic regions associated with host-pathogenic interactions. MP-NAM QTL analyses were conducted on 399 Pyrenophora teres f. teres individuals using biallelic, cross-specific and parental QTL effect models. A bi-parental QTL mapping study was also conducted to compare the power of QTL detection between bi-parental and MP-NAM populations. Using MP-NAM with 399 individuals detected a maximum of eight QTLs with a single QTL effect model whilst only a maximum of five QTLs were detected with an individual bi-parental mapping population of 100 individuals. When reducing the number of isolates in the MP-NAM to 200 individuals the number of QTLs detected remained the same for the MP-NAM population. This study confirms that MPPs such as MP-NAM populations can be successfully used in detecting QTLs in haploid fungal pathogens and that the power of QTL detection with MPPs is greater than with bi-parental mapping populations.

Widespread genetic heterogeneity and genotypic grouping associated with fungicide resistance among barley spot form net blotch isolates in Australia.

Spot form net blotch, caused by Pyrenophora teres f. maculata, is a major foliar disease of barley worldwide. Knowledge of the pathogen's genetic diversity and population structure is critical for a better understanding of inherent evolutionary capacity and for the development of sustainable disease management strategies. Genome-wide, single nucleotide polymorphism data of 254 Australian isolates revealed genotypic diversity and an absence of population structure, either between states, or between fields and cultivars in different agro-ecological zones. This indicates there is little geographical isolation or cultivar directional selection and that the pathogen is highly mobile across the continent. However, two cryptic genotypic groups were found only in Western Australia, predominantly associated with genes involved in fungicide resistance. The findings in this study are discussed in the context of current cultivar resistance and the pathogen's adaptive potential.

Grain protein concentration at elevated [CO2] is determined by genotype dependent variations in nitrogen remobilisation and nitrogen utilisation efficiency in wheat.

Predictions for wheat grown under future climate conditions indicate a decline in grain protein concentration accompanied with an increase in yield due to increasing carbon dioxide concentrations. Currently, there is a lack of understanding as to the complete mechanism that governs the response of grain protein concentration (GPC) to elevated carbon dioxide (e[CO2]). We investigated the GPC of 18 wheat genotypes from a doubled haploid wheat population and the two parental genotypes, Kukri and RAC0875. In addition, other nitrogen and biomass related traits were analysed to further elucidate which traits are connected with the decline in GPC. Wheat was grown under ambient and elevated [CO2] in an environmentally controlled glasshouse. Plant nitrogen and biomass accumulation were measured at anthesis and maturity. We found that GPC declined under e[CO2] and that the response of GPC to e[CO2] was negatively correlated with nitrogen utilisation efficiency and harvest index. The extent that total biomass (anthesis), harvest index, photosynthesis, nitrogen utilisation and remobilisation efficiency, total nitrogen remobilisation and post-anthesis nitrogen uptake impacted GPC in response to e[CO2] varied across genotype, suggesting that multiple mechanisms are responsible for GPC decline at e[CO2] and that these mechanisms are effected differentially across genotypes.

Using a Hybrid Mapping Population to Identify Genomic Regions of Pyrenophora teres Associated With Virulence.

Net blotches caused by Pyrenophora teres are important foliar fungal diseases of barley and result in significant yield losses of up to 40%. The two types of net blotch, net-form net blotch and spot-form net blotch, are caused by P. teres f. teres (Ptt) and P. teres f. maculata (Ptm), respectively. This study is the first to use a cross between Ptt and Ptm to identify quantitative trait loci (QTL) associated with virulence and leaf symptoms. A genetic map consisting of 1,965 Diversity Arrays Technology (DArT) markers was constructed using 351 progenies of the Ptt/Ptm cross. Eight barley cultivars showing differential reactions to the parental isolates were used to phenotype the hybrid progeny isolates. Five QTL associated with virulence and four QTL associated with leaf symptoms were identified across five linkage groups. Phenotypic variation explained by these QTL ranged from 6 to 16%. Further phenotyping of selected progeny isolates on 12 more barley cultivars revealed that three progeny isolates are moderately to highly virulent across these cultivars. The results of this study suggest that accumulation of QTL in hybrid isolates can result in enhanced virulence.

Site-specific, genotypic and temporal variation in photosynthesis and its related biochemistry in wheat (Triticum aestivum).

Photosynthesis in wheat (Triticum aestivum L.) pericarps may contribute appreciably to wheat grain yield. Consequently, we investigated the temporal variation of traits related to photosynthesis and sucrose metabolism in the pericarps and flag leaves of three wheat genotypes, Huandoy, Amurskaja 75 and Greece 25, which are reported to differ in expression of genes related to the C4 pathway in wheat grain. Significant site-specific, genotypic and temporal variation in the maximum carboxylation rate (Vc max ) and maximum rates of electron transport (J max ) (biological capacity of carbon assimilation) were observed early in ontogeny that dissipated by late grain filling. Although the transcript abundance of rbcS and rbcL in flag leaves was significantly higher than in the pericarps, in line with their photosynthetic prominence, both organ types displayed similar expression patterns among growth stages. The higher N concentrations in the pericarps during grain enlargement suggest increased Rubisco; however, expression of rbcS and rbcL indicated the contrary. From heading to 14days post-anthesis, wheat pericarps exhibited a strong, positive correlation between biological capacity for carbon assimilation and expression of key genes related to sucrose metabolism (SPS1 , SUS1 and SPP1 ). The strong correlation between spike dry weight and the biological capacity for carbon assimilation along with other findings of this study suggest that metabolic processes in wheat spikes may play a major role in grain filling, total yield and quality.

Investigating In Vitro Mating Preference Between or Within the Two Forms of Pyrenophora teres and Its Hybrids.

Net blotch diseases result in significant yield losses to barley industries worldwide. They occur as net-form and spot-form net blotch caused by Pyrenophora teres f. teres and P. teres f. maculata, respectively. Hybridization between the forms was proposed to be rare, but recent identifications of field hybrids has renewed interest in the frequency and mechanisms underlying hybridization. This study investigates the mating preference of P. teres f. teres, P. teres f. maculata, and laboratory-produced hybrids in vitro, using 24 different isolates and four different experimental setups. Two crosses in our study produced ascospores during two intervals separated by a 32- to 35-day period of no ascospore production. For these crosses, P. teres f. teres isolates mated with isolates of the same form during the early ascospore production interval, and produced hybrids during the later interval. P. teres f. maculata isolates did not mate with isolates of the same form, but instead hybridized with P. teres f. teres isolates. Analyses based on DArTseq markers confirmed that laboratory-produced hybrids, when given the choice to mate with both P. teres f. teres and P. teres f. maculata, mated with P. teres f. teres isolates. These results unravel a novel concept that P. teres f. teres seems to have a greater reproduction vigor than P. teres f. maculata, which could lead to increased prevalence of hybrid incidences in vivo.

Population Structure of Pyrenophora teres f. teres Barley Pathogens from Different Continents.

Net form net blotch disease, caused by Pyrenophora teres f. teres, results in significant yield losses to barley industries. Up-to-date knowledge of the genetic diversity and structure of pathogen populations is critical for elucidating the disease epidemiology and unraveling pathogen survival and dispersal mechanisms. Thus, this study investigated long-distance dispersal and adaptation by analyzing the genetic structure of 250 P. teres f. teres isolates collected from Australia, Canada, Hungary, and Republic of South Africa (RSA), and historical isolates from Canada, Denmark, Japan, and Sweden. The population genetic structure detected by discriminant analysis of principal components, with the use of 5,890 Diversity Arrays Technology markers, revealed the presence of four clusters. Two of these contained isolates from all regions, and all isolates from RSA were grouped in these two. Australia and Hungary showed three clusters each. One of the Australian clusters contained only Australian isolates. One of the Hungarian clusters contained only Hungarian isolates and one Danish isolate. STRUCTURE analysis indicated that some isolates from Australia and Hungary shared recent ancestry with RSA, Canada, and historical isolates and were thus admixed. Subdivisions of the neighbor joining network indicated that isolates from distinct countries were closely related, suggesting that multiple introduction events conferred genetic heterogeneity in these countries. Through a neighbor joining analysis and amplification with form-specific DNA markers, we detected two hybrid isolates, CBS 281.31 from Japan and H-919 from Hungary, collected in 1931 and 2018, respectively. These results provide a foundation for exploring improved management of disease incursions and pathogen control through strategic deployment of resistance.

Genomic Regions Associated with Virulence in Pyrenophora teres f. teres Identified by Genome-Wide Association Analysis and Biparental Mapping.

Net form net blotch (NFNB), caused by the fungal pathogen Pyrenophora teres f. teres, is an important foliar disease present in all barley-producing regions of the world. This fungus is a hemibiotrophic and heterothallic ascomycete, where sexual recombination can lead to changes in disease expression in the host. Knowledge of the genetic architecture and genes involved in virulence is vital to increase the durability of NFNB resistance in barley cultivars. We used a genome-wide association mapping approach to characterize P. teres f. teres genomic regions associated with virulence in Australian barley cultivars. One hundred eighty-eight P. teres f. teres isolates collected across five Australian states were genotyped using Diversity Arrays Technology sequence markers and phenotyped across 20 different barley genotypes. Association mapping identified 14 different genomic regions associated with virulence, with the majority located on P. teres f. teres chromosomes 3 and 5 and one each present on chromosomes 1, 6, and 9. Four of the regions identified were confirmed by quantitative trait loci (QTL) mapping. The QTL regions are discussed in the context of their genomic architecture together with examination of their gene contents, which identified 20 predicted effectors. The number of QTL shown in this study at the population level clearly illustrates a complex genetic basis of P. teres f. teres virulence compared with pure necrotrophs, such as the wheat pathogens Parastagonospora nodorum and Parastagonospora tritici-repentis.

Exploring natural variation of photosynthesis in a site-specific manner: evolution, progress, and prospects.

MAIN CONCLUSION: Site-specific changes of photosynthesis, a relatively new concept, can be used to improve the productivity of critical food crops to mitigate the foreseen food crisis. Global food security is threatened by an increasing population and the effects of climate change. Large yield improvements were achieved in major cereal crops between the 1950s and 1980s through the Green Revolution. However, we are currently experiencing a significant decline in yield progress. Of the many approaches to improved cereal yields, exploitation of the mode of photosynthesis has been intensely studied. Even though the C4 pathway is considered the most efficient, mainly because of the carbon concentrating mechanisms around the enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase, which minimize photorespiration, much is still unknown about the specific gene regulation of this mode of photosynthesis. Most of the critical cereal crops, including wheat and rice, are categorized as C3 plants based on the photosynthesis of major photosynthetic organs. However, recent findings raise the possibility of different modes of photosynthesis occurring at different sites in the same plant and/or in plants grown in different habitats. That is, it seems possible that efficient photosynthetic traits may be expressed in specific organs, even though the major photosynthetic pathway is C3. Knowledge of site-specific differences in photosynthesis, coupled with site-specific regulation of gene expression, may therefore hold a potential to enhance the yields of economically important C3 crops.

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.

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].

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.

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.

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.

Changes of contrast-specific ultrasonic cerebral perfusion patterns in the course of stroke; reliability of region-wise and parametric imaging analysis.

Ultrasound perfusion imaging (UPI) reliably detects size and localization of acute stroke. It remains unclear which time window detects, most sensitively and specifically, early changes of cerebral perfusion patterns and whether region-wise analysis is superior to parametric imaging analysis. Bilateral phase inversion harmonic imaging examinations (bolus kinetic, fitted model function) were performed twice (acutely and 28 h later) in 10 patients with acute ischemic stroke (<12 h). Examinations were evaluated using a region-wise analysis of the time-intensity curve and by parametric images of the time-to-peak intensity maps. Results were correlated in-between the ultrasound examinations and to follow-up cranial computed tomography (CCT) scans. Correlation between the early region-wise UPI examination and follow-up CCT was the strongest (Spearman correlation coefficient 0.76, sensitivity 84%, specificity 96%). Spearman coefficient between the late UPI examination and CCT was 0.51; sensitivity and specificity were 71% and 82%. Values in between UPI examinations were 57% and 88%, with a Spearman coefficient of 0.47 (p for all < 0.001). Values of the analysis of the parametric images were less strong. Concordance between both of the UPI methods was 65% in the early examination and 72% in the late examination. Changes of perfusion patterns are most accurately detected in the early course of stroke, when core of infarction can be differentiated from penumbra and viable tissue. Reperfusion phenomena may impair the diagnostic impact in later examinations. Parametric imaging does not yet seem to be as accurate as region-wise analysis.