Short Communication. Virulence of barley leaf rust in the South of Russia in 2017-2019

Aim of study: To analyze the structure of Puccinia hordei populations by virulence in southern Russia during 2017-2019. Area of study: South of Russia, the leading Russian region for barley production where barley leaf rust is an important foliar disease. Material and methods: Uredinial samples of P. hordei were collected at the production sites of winter barley in the south of Russia. Single uredinial isolates (total 95) were tested for virulence with 17 differentials with Rph resistance genes. Main results: No isolates found virulent to the host line with the Rph13 gene. There was a decrease in the number of fungal isolates virulent to the host lines with Rph5 and Rph7 genes. In 2017 and 2019, isolates containing a large number of virulence alleles (from 11 to 15) prevailed. In 2018, isolates with low (1-5) and medium (6-10) frequency of virulent alleles prevailed, as well as avirulent isolates. The values of the Nei index via diversity showed high similarity of the pathogen populations in 2017-2018 (N = 0.05) and minor differences in 2017-2019 and 2018-2019 (N = 0.13 and 0.16, respectively). The greatest frequency of virulence alleles in accordance with the Nei (Hs) index was noted for the 2018 population (Hs = 0.36). For the 2017 and 2019 populations, this indicator was average (Hs = 0.29 and 0.20, respectively). Research highlights: Analysis of genetics of the P. hopdei population is important for the strategy of varietal distribution in the region and development of rust-resistant cultivars.

Mining and predictive characterization of resistance to leaf rust (Puccinia hordei Otth) using two subsets of barley genetic resources

Sustainable barley (Hordeum vulgare L.) production will require access to diverse ex-situ conserved collections to develop varieties with high yields and capable of overcoming the challenges imposed by major abiotic and biotic stresses. This study aimed at searching efficient approaches for the identification of new sources of resistance to barley leaf rust (Puccinia hordei Otth). Two subsets, Generation Challenge Program Reference set (GCP) with 188 accessions and leaf rust subset constructed using the filtering approach of the Focused Identification of Germplasm Strategy (FIGS) with 86 accessions, were evaluated for the seedling as well as the adult plant stage resistance (APR) using two barley leaf rust (LR) isolates (ISO-SAT and ISO-MRC) and in four environments in Morocco, respectively. Both subsets yielded a high percent of accessions with a moderately resistant (MR) reaction to the two LR isolates at the seedling stage. For APR, more than 50% of the accessions showed resistant reactions in SAT2018 and GCH2018, while this rate was less than 20% in SAT2017 and SAT2019. Statistical analysis using chi-square test of independence revealed the dependency of LR reaction on subsets at the seedling (ISO-MRC), as well as at the APR (SAT2017 and SAT2018) stage. At seedling stage, the test of goodness of fit showed that GCP subset yielded higher percentages of resistant accessions than FIGS-LR in case of ISO-MRC isolate but the two subsets did not differ for ISO-SAT. At APR, FIGS approach performed better than GCP in yielding higher percentages of accessions in case of SAT2017 and SAT2018. Although some of the tested machine learning models had moderate to high accuracies, none of them was able to find a strong and significant relationship between the reaction to LR and the environmental conditions showing the needs for more fine tuning of approaches for efficient mining of genetic resources using machine learning.

Validating molecular markers for barley leaf rust resistance genes Rph20 and Rph24

Improving resistance to barley leaf rust (caused by Puccinia hordei) is an important breeding objective in most barley growing regions worldwide. The development and subsequent utilization of high-throughput polymerase chain reaction (PCR) based co-dominant molecular markers remains an effective approach to select genotypes with multiple effective resistance genes, permitting efficient gene deployment and stewardship. The genes Rph20 and Rph24, which confer widely effective adult plant resistance (APR) to leaf rust, are common in European and Australian barley germplasm (often in combination), and act interactively to confer high levels of resistance. Here we report on the development and validation of co-dominant insertion-deletion (indel) based PCR markers that are highly predictive for the resistance alleles Rph20.ai and Rph24.an (both referred to as Rph20 and Rph24).

Validating molecular markers for barley leaf rust resistance genes Rph20 and Rph24

Improving resistance to barley leaf rust (caused by Puccinia hordei) is an important breeding objective in most barley growing regions worldwide. The development and subsequent utilisation of high-throughput PCR-based co-dominant molecular markers remains an effective approach to select genotypes with multiple effective resistance genes, permitting efficient gene deployment and stewardship. The genes Rph20 and Rph24 confer widely effective adult plant resistance (APR) to leaf rust, are common in European and Australian barley germplasm (often in combination), and act interactively to confer high levels of resistance (Dracatos et al. 2015; Zeims et al. 2017; Singh et al. 2018). Here we report on the development and validation of co-dominant insertion-deletion (indel) based PCR markers that are highly predictive for the Rph20 and Rph24 resistances.

De Novo Genome Assembly and Comparative Genomics of the Barley Leaf Rust Pathogen Puccinia hordei Identifies Candidates for Three Avirulence Genes

Puccinia hordei (Ph) is a damaging pathogen of barley throughout the world. Despite its importance, almost nothing is known about the genomics of this pathogen, and a reference genome is lacking. In this study, the first reference genome was assembled for an Australian isolate of Ph (“Ph560”) using long-read SMRT sequencing. A total of 838 contigs were assembled, with a total size of 207 Mbp. This included both haplotype collapsed and separated regions, consistent with an estimated haploid genome size of about 150Mbp. An annotation pipeline that combined RNA-Seq of Ph-infected host tissues and homology to proteins from four other Puccinia species predicted 25,543 gene models of which 1,450 genes were classified as encoding secreted proteins based on the prediction of a signal peptide and no transmembrane domain. Genome resequencing using short-read technology was conducted for four additional Australian strains, Ph612, Ph626, Ph608 and Ph584, which are considered to be simple mutational derivatives of Ph560 with added virulence to one or two of three barley leaf rust resistance genes (viz. Rph3, Rph13 and Rph19). To identify candidate genes for the corresponding avirulence genes AvrRph3, AvrRph13 and AvrRph19, genetic variation in predicted secreted protein genes between the strains was correlated to the virulence profiles of each, identifying 35, 29 and 46 candidates for AvrRph13, AvrRph3 and AvrRph19, respectively. The identification of these candidate genes provides a strong foundation for future efforts to isolate these three avirulence genes, investigate their biological properties, and develop new diagnostic tests for monitoring pathogen virulence.