Mapping of crown rust (Puccinia coronata f. sp. avenae) resistance gene Pc54 and a novel quantitative trait locus effective against powdery mildew (Blumeria graminis f. sp. avenae) in the oat (Avena sativa) line Pc54

The Pc54 oat line carries the crown rust resistance gene ‘Pc54’ and an unknown gene effective against powdery mildew. In this study two recombinant inbred line populations were developed to identify the genomic locations of the two genes and producing lists of molecular markers with a potential for marker assisted selection. The RILs and parents were phenotyped for crown rust and powdery mildew in a controlled environment. They were also genotyped using the 6K Illumina Infinium iSelect oat SNP chip. Multiple interval mapping placed Pc54 on the linkage group Mrg02 (chromosome 7D) and the novel powdery mildew QTL ‘QPm.18’ on Mrg18 (chromosome 1A) both in the mapping and validating population. A total of nine and 31 significant molecular markers were identified linked with the Pc54 gene and QPm.18, respectively. Reactions to crown rust inoculations have justified separate identity of Pc54 from other genes and QTL that have previously been reported on Mrg02 except for ’qPCRFd’. Pm3 is the only powdery mildew resistance gene previously mapped on Mrg18. However, the pm3 differential line, Mostyn was susceptible to the powdery mildew race used in this study suggesting that Pm3 and QPm.18 are different genes. Determining the chromosomal locations of Pc54 and QPm.18 is helpful for better understanding the molecular mechanism of resistance to crown rust and powdery mildew in oats. Furthermore, SNPs and SSRs that are closely linked with the genes could be valuable for developing PCR based molecular markers and facilitating the utilization of these genes in oat breeding programs.

Discovery and Chromosomal Location a Highly Effective Oat Crown Rust Resistance Gene Pc50-5

Crown rust, caused by Puccinia coronata f. sp. avenae, is one of the most destructive fungal diseases of oat worldwide. Growing disease-resistant oat cultivars is the preferred method of preventing the spread of rust and potential epidemics. The object of the study was Pc50-5, a race-specific seedling crown rust resistant gene, highly effective at all growth stages, selected from the differential line Pc50 (Avena sterilis L. CW 486-1 × Pendek). A comparison of crown rust reaction as well as an allelism test showed the distinctiveness of Pc50-5, whereas the proportions of phenotypes in segregating populations derived from a cross with two crown rust-susceptible Polish oat cultivars, Kasztan × Pc50-5 and Bingo × Pc50-5, confirmed monogenic inheritance of the gene, indicating its usefulness in oat breeding programs. Effective gene introgression depends on reliable gene identification in the early stages of plant development; thus, the aim of the study was to develop molecular markers that are tightly linked to Pc50-5. Segregating populations of Kasztan × Pc50-5 were genotyped using DArTseq technology based on next-generation Illumina short-read sequencing. Markers associated with Pc50-5 were located on chromosome 6A of the current version of the oat reference genome (Avena sativa OT3098 v2, PepsiCo) in the region between 434,234,214 and 440,149,046 bp and subsequently converted to PCR-based SCAR (sequence-characterized amplified region) markers. Furthermore, 5426978_SCAR and 24031809_SCAR co-segregated with the Pc50-5 resistance allele and were mapped to the partial linkage group at 0.6 and 4.0 cM, respectively. The co-dominant 58163643_SCAR marker was the best diagnostic and it was located closest to Pc50-5 at 0.1 cM. The newly discovered, very strong monogenic crown rust resistance may be useful for oat improvement. DArTseq sequences converted into specific PCR markers will be a valuable tool for marker-assisted selection in breeding programs.

Oat crown rust disease severity estimated at many time points using multispectral aerial photos

All plant breeding programs are dependent on plant phenotypic and genotypic data, but the development of phenotyping technology has been slow relative to that of genotyping. Crown rust (Puccinia coronata f. sp. avenae Erikss.) is the most important disease of cultivated oat (Avena sativa L.) making the development of disease resistant oat cultivars an important breeding objective. Visual observation is the most common scoring method, but it can be laborious and subjective. We visually scored a diverse collection of 256 oat lines at a total of twenty-seven time points in three disease nursery environments. Multispectral aerial photos were collected using an unmanned aerial vehicle at the same time points as the visual observations. The photos were analyzed and a subset of spectral properties of each plot were measured. Random forest modeling was used to model the relationship between the spectral properties of the plots and visually observed disease severity. The ability of the photo data and the random forest model to estimate visually observed disease severity was evaluated using three different cross-validation analyses. We specifically address the issue of assessing phenotyping accuracy across and within time points. The accuracy of the photo estimates was greatest for adult plants shortly before they began to senesce. Accuracy outside of that time frame is generally low, but statistically significant. Unmanned aerial vehicle mounted sensors could increase disease scoring efficiency, but additional investigation into the spectral signature of disease severity at all plant growth stages may be necessary to automate accurate full-season measurements.

First report of stem rust, caused by Puccinia graminis, and crown rust, caused by Puccinia coronata var. avenae f. sp. avenae, on tall fescue (Festuca arundinacea) in Georgia, USA

Stem rust, caused by Puccinia graminis, and crown rust, caused by P. coronata, are common rust diseases on cool-season grasses (Karakkat et al. 2018), for which long-distance spore dispersal was recorded in northern US (Harder and Haber 1992). During the summers of 2019 and 2020, severe infection of stem rust and crown rust was observed on > 60% of tall fescue (Festuca arundinacea) germplasm plants in a breeding nursery located at the University of Georgia, Griffin GA. Rust-infected leaves first presented uredinia pustules, then black telia towards the end of the season. The uredinia pustules of stem rust and crown rust were brick-red and, yellow and arranged along the host veins, respectively. The urediniospores were one-celled, round to ovoid and measured from 20.75±2.44 μm (crown rust) to 27±3.60 μm long (stem rust). The teliospores were two-celled and measured from 45.75±10.14 μm (stem rust) to 51.60±4.0 μm long (crown rust) (Leonard et al. 2005; Cummins 1971). Urediniospores of both rusts were collected from infected plants in the field in April of 2020 using a Piston vacuum pump (Welch by Gardner Denver Ltd.) and stored at -80 °C in 1.5 ml Eppendorf tubes. Genomic DNA was extracted by grinding the urediniospores in liquid nitrogen using mortar and pestle, followed by the cetyltrimethylammonium bromide method (Doyle and Doyle 1987). The internal transcribed spacer (ITS) region of the ribosomal DNA was amplified using the ITS5-ITS4 primers (White et al. 1990). BLASTn and phylogenetic analyses revealed that the sequence of stem rust (GenBank acc. no. MW430963) and crown rust (GenBank acc. no. MW431324) pathogens had >99% similarity with P. graminis (GenBank acc. no. HQ317538) and P. coronata var. avenae f. sp. avenae (clade V; Liu and Hambleton 2013) (GenBank acc. no. EU014044), respectively. Pathogenicity tests were conducted on the tall fescue cultivar ‘Bandit’. For each rust, 12 pots (10 cm × 10 cm) were planted, each containing 13 seeds in a Sungro professional growing mix soil (Sun Gro Horticulture Distribution Inc.). The plant materials were kept in the greenhouse at 20°C/ 25°C (night/day),15-hrs of light, and watered twice a week for 4-weeks. Urediniospores were recovered from -80°C and allowed to acclimate at room temperature for 1 h. For each rust, 20 ml of suspension containing 1×105 urediniospores ml−1 and 5 μl of Tween-twenty (Agdia Inc. Elkhart, IN) were used to inoculate 6 pots; while 6 control pots were sprayed with sterile water. After inoculation, plants were allowed to dry for 1 h and then transferred to a dark chamber at 20°C and 90% of humidity for 12-15 h. At 10-days post inoculation, all inoculated plants developed rust symptoms identical to those observed in the field, whereas control plants had no symptoms. Stem and crown rust pathogens were re-isolated from the artificially inoculated tall fescue plants. Based on form, size, color and numbers of cells forming the spores, a 1947 Festuca elatior specimen from Georgia mentioning Puccinia coronata (Hanlin 1966), held at the Julian H. Miller Mycological Herbarium (Catalog No. GAM00013162), was discarded as an earlier record of P. coronata var. avenae and could have been misdiagnosed. Due to the fragile integrity of the original infected plant sample as well as the incipient infection, DNA identification was unsuccessful. To our knowledge, this is the first morphological, genetic and taxonomic report of P. graminis and P. coronata var. avenae f. sp. avenae on tall fescue in Georgia, USA

Bipolaris victoriae (Victoria blight of oats).

A disease almost unknown prior to 1944 (Romanko, 1957), Bipolaris victoriae is a potentially destructive disease of oats (it reduced the Iowa oat crop by 32% in 1947) (Anon, 1965) that currently has a limited host range and geographical spread. It has been reported from several continents and associated with several crops, however its status as a pathogen on crops could be questioned in all cases with the exception of oats, timothy grass and switch grass. It is seedborne and thus any trade in seed both for cultivation or for livestock feed is a potential means of spread. The virulence of the fungus is due to the production of a peptide often called a host-specific toxin 'victorin'. The disease has generally been controlled through the use of genetically resistant lines but the resistance gene which confers resistance to victorin actually causes plants to become susceptible to crown rust, caused by Puccinia coronata (Lorang et al., 2007).

Bipolaris victoriae (Victoria blight of oats).

A disease almost unknown prior to 1944 (Romanko, 1957), Bipolaris victoriae is a potentially destructive disease of oats (it reduced the Iowa oat crop by 32% in 1947) (Anon, 1965) that currently has a limited host range and geographical spread. It has been reported from several continents and associated with several crops, however its status as a pathogen on crops could be questioned in all cases with the exception of oats, timothy grass and switch grass. It is seedborne and thus any trade in seed both for cultivation or for livestock feed is a potential means of spread. The virulence of the fungus is due to the production of a peptide often called a host-specific toxin 'victorin'. The disease has generally been controlled through the use of genetically resistant lines but the resistance gene which confers resistance to victorin actually causes plants to become susceptible to crown rust, caused by Puccinia coronata (Lorang et al., 2007).

Evidence of occurrence of Crown Rust of Barley Caused by Puccinia coronata var. hordei and sexual reproduction of the pathogen under field conditions in China

Crown rust of barley, caused by Puccinia coronata var. hordei (Pch), was first reported by Jin and Steffenson in 1992, and the fungus has been reported only in the United States and Hungary. In China, stripe, stem, and leaf rusts have been reported on barley, but not for crown rust. Recently, a sample (HZJ0004) of rust collected from barley in Qilian county, Qinghai, China, appeared different from the three rusts based on color, size, arrangements of uredinia and/or telia. Teliospores had crown-shaped appendages on the top. Based on the disease symptoms and morphology of urediniospores and teliospores, the fungus was identified as Pch. Using the internal transcribed spacer sequences, the isolates HZJ0004 from barley and POR3 from buckthorn (Rhamnus sp.) were clustered in one clade with Pch isolates from barley and Elymus repens but in a different clade from the isolate POC8 from wild oat and the varieties of P. coronata from oats and grasses. At the seedling stage, most of the tested cultivars of barley and rye were susceptible to Pch isolates HZJ0004 and POR3, but the cultivars of oats, triticale, wheat, and the most grasses of Aegilops, Brachypodium, Bromus, Calamagrostis, Deschampsia, Elymus, Festuca, and Phleum were resistant, indicating their host specialization on barley. This is the first report of crown rust on barley in China.

Hidden Diversity of Crown Rust Resistance within Genebank Resources of Avena sterilis L.

The most widespread and damaging fungal disease of the oat plant is crown rust. Resistance to the crown rust pathogen, Puccinia coronata Cda. f. sp. avenae (Pca), at the seedling stage of Avena sterilis accessions from the Polish national genebank was characterised by five North American and Polish pathotypes of Pca of diverse pathogenicity. Pca pathogenicity was determined on a series of 34 differential lines carrying known seedling resistance genes. Seventy-five percent of studied accessions showed a heterogeneous infection pattern, 17% behaved as homogenous susceptibles, and 7% of tested genotypes could be unambiguously described as resistant. This study proved that A. sterilis accessions preserved in a genebank as complex populations could be a very valuable source of resistance to crown rust. The complexity of analysed populations was ascertained by a detailed variance analysis of transformed resistance/susceptibility data. We demonstrate here that hidden sources of resistance may be discovered in accessions with general susceptibility.

Crown rust on oat genotypes with different levels of resistance: damages and losses

ABSTRACT: The evolution in virulence of Puccinia coronata f. sp. avenae has challenged the genetic resistance to crown rust in oat genotypes. New resistance sources are constantly required for yield stabilization and costs reduction. This study aimed to characterize the crown rust resistance of oat genotypes and the disease´s productive and economic impact. Experiments were conducted in two environments, with and without chemical control. Resistance was measured by disease progress, apparent rate of infection, final severity and pustule size. Damages were based on reduction of yield, 1000 grain mass and hectoliter mass. UFRGS16Q6030-2 was immune. In both environments, UFRGS166091-2 and URS Brava exhibited the greatest level of resistance and the smallest grain yield reduction, while URS 22 was highly susceptible, reducing more than 70% of its grain yield. These results surpass the negative impacts of the disease previously reported in oats, particularly related to grain yield. The data presented in this paper highlighted the importance of genetic resistance, particularly partial resistance, for maintaining oat genetic yield potential, reducing environmental contamination with less fungicides, and increasing economic gains with oat cultivation.

Selection of Pyramided Barley Advanced Lines for Stripe Rust, Leaf Rust and Crown Rust Diseases Using Molecular Markers

Barley diseases are the major yield limiting factors for barley cultivation in Nepal. Stripe/Yellow rust (P. striformis f.sp. hordei and P. striformis f.sp. tritici), leaf rust (Puccinia hordei), and crown rust (P. coronata) are the major rust diseases in Nepal. Pyramiding resistance genes against all these rust diseases are possible through molecular marker assisted breeding. Sweden originated barley variety ‘Bonus’ is found resistant to stripe rust and having linked microsatellite markers for stripe rust and crown rust resistance. Similarly, Nepalese hull-less barley variety ‘Solu Uwa’ and Nepalese awn-less barley landrace NPGR Acc# 2478 have linked microsatellite markers for leaf rust resistance. Therefore, one polymorphic sequence tagged sites (STS) marker (ABG054) for stripe rust resistance, two polymorphic simple sequence repeats (SSR) markers (Bmac0144h and HVM049) for leaf rust and one polymorphic SSR marker (Bmag0006) for crown rust resistance were used to select the advanced barley lines (at F8 stage) from above parents. Field screening of stripe rust resistance was also conducted. Among 51 advanced and field disease resistance lines from Bonus/Solu Uwa cross, we have selected 10 pyramided lines for all three types of barley rust resistance. Similarly, among 39 advanced and field disease resistance lines from Bonus/NPGR Acc#2478 cross we have selected three pyramided lines and advanced for further yield testing for general cultivation purpose. The chances of losing the desired gene are high in late generation selection using molecular marker assisted selection (MAS); but the chances of getting agronomically superior varietal output will also increase.