A region of barley chromosome 6H harbors multiple major genes associated with net type net blotch resistance

Net type net blotch (NTNB), caused by Pyrenophora teres f. teres Drechs., is prevalent in barley growing regions worldwide. A population of 118 doubled haploid (DH) lines developed from a cross between barley cultivars ‘Rika’ and ‘Kombar’ were used to evaluate resistance to NTNB due to their differential reaction to various isolates of P. teres f. teres. Rika was resistant to P. teres f. teres isolate 15A and susceptible to isolate 6A. Conversely, Kombar was resistant to 6A, but susceptible to 15A. A progeny isolate of a 15A × 6A cross identified as 15A × 6A#4 was virulent on both parental lines. The Rika/Kombar (RK) DH population was evaluated for disease reactions to the three isolates. Isolate 15A induced a resistant:susceptible ratio of 78:40 (R:S) whereas isolate 6A induced a resistant:susceptible ratio of 40:78. All but two lines had opposite disease reactions indicating two major resistance genes linked in repulsion. Progeny isolate 15A × 6A#4 showed a resistant:susceptible ratio of 1:117 with the one resistant line also being the single line that was resistant to both 15A and 6A. An RK F₂ population segregated in a 1:3 (R:S) ratio for both 15A and 6A indicating that resistance is recessive. Molecular markers were used to identify a region on chromosome 6H that harbors the two NTNB resistance genes. This work shows that multiple NTNB resistance genes exist at the locus on chromosome 6H, and the recombinant DH line harboring the resistance alleles from both parents will be useful for the development of NTNB-resistant barley germplasm.     

Genetics of seedling and adult plant resistance to net blotch (Pyrenophora teres f. teres) and spot blotch (Cochliobolus sativus) in barley

Net blotch (caused by Pyrenophora teres f. teres) and spot blotch (Cochliobolus sativus) are important foliar diseases of barley in the midwestern region of the USA. To determine the number and chromosomal location of Mendelian and quantitative trait loci (QTL) controlling resistance to these diseases, a doubled haploid population (Steptoe/Morex) was evaluated to the pathogens at the seedling stage in the greenhouse and at the adult plant stage in the field. Alleles at two or three unlinked loci were found to confer resistance to the net blotch pathogen at the seedling stage depending on how progeny exhibiting an intermediate infection response were classified. This result was corroborated in the quantitative analysis of the raw infection response data as 2 major QTL were identified on chromosomes 4 and 6M. A third QTL was also identified on chromosome 6P. Seven QTL were identified for net blotch resistance at the adult plant stage and mapped to chromosomes 1P, 2P, 3P, 3M, 4, 6P, and 7P. The 7 QTL collectively accounted for 67.6% of the phenotypic variance under a multiple QTL model. Resistance to the spot blotch pathogen was conferred by a single gene at the seedling stage. This gene was mapped to the distal region of chromosome 1P on the basis of both qualitative and quantitative data analyses. Two QTL were identified for spot blotch resistance at the adult plant stage: the largest QTL effect mapped to chromosome 5P and the other mapped to chromosome 1P near the seedling resistance locus. Together, the 2 QTL explained 70.1% of the phenotypic variance under a multiple QTL model. On the basis of the chromosomal locations of resistance alleles detected in this study, it should be feasible to combine high levels of resistance to both P. teres f. teres and C. sativus in barley cultivars.     

Different Assessment Techniques of Leaf Blotch (Rhynchosporium secalis (Oud.) J. Davis) in Winter Barley after Artificial Inoculation

Seven winter barley cultivars expressing different field resistance against leaf blotch (Rhynchosporium secalis) and 60 doubled haploid (DH) lines descending from and F₁ hybrid resistant x susceptible were used to develop a test for the quantitative determination of disease severity. Differences in the level of resistance could be measured very precisely after artificial infection of seedlings at the 4-leaf-stage and subseqent incubation at 7 C. The protein of six isolates was usedf, or antiserum production which was subsequently supplied for serological testing (Enzyme-Linked-Immunosorbent-Assay) of infected leaves. The results of visible scoring of the seedlings was significantly correlated with measuring with the ELISA reader. The, results of the seedling test also corresponded to field scores after natural infection.     

Monosomic and double monosomic substitutions of Hordeum bulbosum L. chromosomes into H. vulgare L.

Summary One of the aims of the interspecific crossing programme between Hordeum vulgare L. and H. bulbosum L. has been to introgress desirable genes into barley from the wild species. However, despite their close taxonomic relationship there have been few reports of achieving this objective using amphidiploid hybrids. In order to broaden the range of available hybrids, partially fertile triploids between H. vulgare (2n = 2x = 14) and H. bulbosum (2n = 4x = 28) were developed and crossed with H. vulgare as female parent. From 580 progeny which were screened, eight putative single monosomic chromosome substitution lines and one double monosomic substitution were identified by cytological analysis. These involved the substitution of H. vulgare chromosome 1 (two lines), 6 (four lines), 6L (one line), 7 (one line) and 1 + 4 (one line) by their respective H. bulbosum homoeologues. The H. bulbosum chromosome was frequently eliminated during plant development, but it was observed regularly in pollen mother cells of two lines. However, pairing between the H. bulbosum chromosome and its H. vulgare homoeologue was low. Several of the lines were more resistant than their H. vulgare parents to powdery mildew (Erysiphe graminis DC. f.sp. hordei Em. Marchai), net blotch (Drechslera teres Sacc.) and scald (Rhynchosporium secalis (Oudem.) Davis). Apart from their use in studying genome relationships, their value to plant breeders will depend on the ease of inducing translocations between the parental chromosomes.     

AB-QTL analysis in spring barley. I. Detection of resistance genes against powdery mildew, leaf rust and scald introgressed from wild barley

The objective of this study was to map new resistance genes against powdery mildew (Blumeria graminis f. sp. hordei L.), leaf rust (Puccinia hordei L.) and scald [Rhynchosporium secalis (Oud.) J. Davis] in the advanced backcross doubled haploid (BC₂DH) population S42 derived from a cross between the spring barley cultivar Scarlett and the wild barley accession ISR42-8 (Hordeum vulgare ssp. spontaneum). Using field data of disease severity recorded in eight environments under natural infestation and genotype data of 98 SSR loci, we detected nine QTL for powdery mildew, six QTL for leaf rust resistance and three QTL for scald resistance. The presence of the exotic QTL alleles reduced disease symptoms by a maximum of 51.5, 37.6 and 16.5% for powdery mildew, leaf rust and scald, respectively. Some of the detected QTL may correspond to previously identified qualitative (i.e. Mla) and to quantitative resistance genes. Others may be newly identified resistance genes. For the majority of resistance QTL (61.0%) the wild barley contributed the favourable allele demonstrating the usefulness of wild barley in the quest for resistant cultivars.     

Role of reactive oxygen species in the response of barley to necrotrophic pathogens

Summary.  The interactions between Hordeum vulgare (barley) and two fungal necrotrophs, Rhynchosporium secalis and Pyrenophora teres (causal agents of barley leaf scald and net blotch), were investigated in a detached-leaf system. An early oxidative burst specific to epidermal cells was observed in both the susceptible and resistant responses to R. secalis, and later on, a second susceptible-specific burst was observed. Time points of the first and the second burst correlated closely with pathogen contact to the plasma membrane and subsequent cell death, respectively. HO₂ ^•/O₂ ⁻ levels in resistant and susceptible responses to P. teres were limited in comparison. During later stages, HO₂ ^•/O₂ ⁻ was only detected in 2 to 3 epidermal cells immediately adjacent to phenolic browning and cell death observed during the susceptible response. However, H₂O₂ was detected in the majority of mesophyll cells adjacent to the observed lesion caused by P. teres. In contrast to observations during challenge with R. secalis, no direct contact between P. teres and the plasma membrane at sites of reactive oxygen species production was evident. Preinfiltration of leaves with antioxidants prior to challenge with either pathogen had no effect on resistance responses but did limit the growth of the pathogens and inhibit the extent of cell death during susceptible responses. These results suggest a possible role for reactive oxygen species in the induction of cell death during the challenge of a susceptible plant cell with a necrotrophic fungal leaf pathogen. Received May 2, 2002; accepted July 26, 2002; published online May 21, 2003 RID="*" ID="*" Correspondence and reprints: Department of Plant Science, Waite Campus PMB1, University of Adelaide, Glen Osmond, South Australia 5064, Australia.     

Research advances in the Pyrenophora teres–barley interaction

Pyrenophora teres f. teres and P. teres f. maculata are significant pathogens that cause net blotch of barley. An increased number of loci involved in P. teres resistance or susceptibility responses of barley as well as interacting P. teres virulence effector loci have recently been identified through biparental and association mapping studies of both the pathogen and host. Characterization of the resistance/susceptibility loci in the host and the interacting effector loci in the pathogen will provide a path for targeted gene validation for better-informed release of resistant barley cultivars. This review assembles concise consensus maps for all loci published for both the host and pathogen, providing a useful resource for the community to be used in pathogen characterization and barley breeding for resistance to both forms of P. teres.     

A first genome assembly of the barley fungal pathogen <Emphasis Type="Italic">Pyrenophora teres</Emphasis> f. <Emphasis Type="Italic">teres</Emphasis>

Background  

Pyrenophora teres f. teres is a necrotrophic fungal pathogen and the cause of one of barley's most important diseases, net form of net blotch. Here we report the first genome assembly for this species based solely on short Solexa sequencing reads of isolate 0-1. The assembly was validated by comparison to BAC sequences, ESTs, orthologous genes and by PCR, and complemented by cytogenetic karyotyping and the first genome-wide genetic map for P. teres f. teres.     

Identification of quantitative trait loci associated with resistance to net form net blotch in a collection of Nordic barley germplasm

Key message

Association mapping of resistance to Pyrenophora teres f. teres in a collection of Nordic barley germplasm at different developmental stages revealed 13 quantitative loci with mostly small effects.

Abstract

Net blotch, caused by the necrotrophic fungus Pyrenophora teres, is one of the major diseases in barley in Norway causing quantitative and qualitative yield losses. Resistance in Norwegian cultivars and germplasm is generally insufficient and resistance sources have not been extensively explored yet. In this study, we mapped quantitative trait loci (QTL) associated with resistance to net blotch in Nordic germplasm. We evaluated a collection of 209 mostly Nordic spring barley lines for reactions to net form net blotch (NFNB; Pyrenophora teres f. teres) in inoculations with three single conidia isolates at the seedling stage and in inoculated field trials at the adult stage in 4 years. Using 5669 SNP markers genotyped with the Illumina iSelect 9k Barley SNP Chip and a mixed linear model accounting for population structure and kinship, we found a total of 35 significant marker-trait associations for net blotch resistance, corresponding to 13 QTL, on all chromosomes. Out of these QTL, seven conferred resistance only in adult plants and four were only detectable in seedlings. Two QTL on chromosomes 3H and 6H were significant during both seedling inoculations and adult stage field trials. These are promising candidates for breeding programs using marker-assisted selection strategies. The results elucidate the genetic background of NFNB resistance in Nordic germplasm and suggest that NB resistance is conferred by a number of genes each with small-to-moderate effects, making it necessary to pyramid these genes to achieve sufficient levels of resistance.

     

Comparative virulence of Pyrenophora teres f. teres from Syria and Tunisia and screening for resistance sources in barley: implications for breeding

Aims: The aim of this study is to investigate the pathogenic diversity and virulence groups among Pyrenophora teres f. teres isolates, sampled from Syria and Tunisia, and to identify the most effective source of resistance in barley that could be used in breeding programmes to control net blotch in both countries. Methods and Results: One hundred and four isolates of P. teres f. teres were collected from barley in different agroecological zones of Tunisia and Syria. Their virulence was evaluated using 14 barley genotypes as differential hosts. The upgma clustering identified high pathogenic variability; the isolates were clustered onto 20 pathotypes that were sheltered under three virulence groups, with high, intermediate and low disease scores. According to susceptibility/resistance frequencies and mean disease ratings, CI05401 cultivar ranked as the best differential when inoculated with the Syrian isolates. However, CI09214 cultivar was classified as the best effective source of resistance in Tunisia. Conclusions: All P. teres f. teres isolates were differentially pathogenic. CI09214 and CI05401 cultivars were released as the most effective sources of resistance in Syria and Tunisia. Significance and Impact of the Study: National and international barley breeding programmes that seek to develop resistance against P. teres f. teres in barley should strongly benefit from this study. This resistance cannot be achieved without the proper knowledge of the pathogen virulence spectrum and the sources of host resistance.     

Identification and mapping of a gene conferring resistance to the spot form of net blotch (Pyrenophora teres f maculata) in barley

 Spot form of net blotch (SFNB) (Pyrenophora teres f maculata) is an economically damaging foliar disease of barley in many of the world’s cereal growing areas. The development of SFNB-resistant cultivars may be accelerated through the use of molecular markers. A screen for SFNB resistance in 96 lines identified four new sources of resistance, including a feed variety, ‘Galleon’, for which a fully mapped doubled haploid population was available. Segregation data indicated SFNB resistance was conferred by a single gene in the ‘Galleon’בHaruna Nijo’ cross, positioned on the long arm of chromosome 7H. This gene is designated Rpt4 and is flanked by the RFLP loci Xpsr117(D) and Xcdo673 at distances of 6.9 cM and 25.9 cM, respectively. The marker Xpsr117(D) was validated using another population segregating for Rpt4, correctly predicting SFNB resistance with more than 90% accuracy. Received: 24 September 1998 / Accepted: 19 December 1998     

Mapping quantitative trait loci associated with spot blotch and net blotch resistance in a doubled-haploid barley population

Spot blotch and net blotch are important foliar barley (Hordeum vulgare L.) diseases in Canada and elsewhere. These diseases result in significant yield reduction and, more importantly, loss of grain quality, downgrading barley from malt to feed. Combining resistance to these diseases is a breeding priority but is a significant challenge using conventional breeding methodology. In the present investigation, an evaluation of the inheritance of resistance to spot and net blotch was conducted in a doubled-haploid barley population from the cross CDC Bold (susceptible)?×?TR251 (resistant). The population was screened at the seedling stage in the Phytotron and at the adult-plant stage in the field for several years. Chi-squared analysis indicated one- to four-gene segregation depending on disease, isolate, plant development stage, location and year. A major seedling and adult-plant resistance quantitative trait locus (QTL), designated QRpt6, was re-confirmed for net-form net blotch resistance, explaining 32?C61% of phenotypic variation in different experiments. Additional QTL for seedling and adult-plant resistance to net blotch were identified. For spot blotch resistance, a major seedling resistance QTL (QRcss1) was detected on chromosome 1H for isolate WRS1909, explaining 79% of the phenotypic variation. A highly significant QTL on 3H (QRcs3) was identified for seedling resistance to isolate WRS1908 and adult-plant resistance at Brandon, MB, Canada in 2008. The identification of QTL at only one location or from 1?year suggests spot blotch resistance is complex and highly influenced by the environment. Efforts are being made to combine spot and net blotch resistance in elite barley lines using molecular marker-assisted selection.     

Effect of Four Fungicides on Barley Net Blotch Caused by Drechslera teres

The effect of four fungicides (benomyl, carbendazim, triadimefon and sulp hur), used in intensive barley cropping on the epidemiology of Drechslera teres f. maculata and f. teres, was investigated under a controlled environment. Two fungicides (benomyl and carbendazim) increased disease development, especially at the recommended field rate, sulphur had the same effect at lower concentrations and triadimefon promoted net blotch caused by D. teres f. teres. At the recommended field rate, carbendazim enhanced sporulation in the two pathogen forms, whereas benomyl and sulphur enhanced 14 sporulation only on the maculata form. The results showed that, under controlled conditions, most of the fungicides promotednet blotch development and that sporulation increased either as a result of the pathogen form or of the concentration of the active ingredient.     

Development of sequence‐tagged microsatellites for the barley net blotch pathogen,Pyrenophora teres

A modified sequenced‐tagged microsatellite (STM) profiling procedure was used to develop 80 STMs for the barley net blotch pathogen, Pyrenophora teres. Of these, 60 STMs amplified 67 loci in one or both of the spot (P. teres f. maculata) and net (P. teres f. teres) forms of the pathogen. When screened on six field‐sampled isolates of each pathogen form, 25 STMs revealed 26 polymorphic loci, with an average of 3.2 ± 1.0 alleles and mean gene diversity of 0.59 ± 0.12.     

Expression of resistance to Leptosphaeria maculans in Brassica napus double haploid lines in France and Australia is influenced by location

Blackleg, caused by Leptosphaeria maculans, is a major disease of oilseed rape (Brassica napus), worldwide, including Australia and France. The aims of these studies were first, to determine if higher levels of resistance to L. maculans could be generated in double haploid (DH) lines derived from spring‐type B. napus cv. Grouse, which has a good level of field resistance to blackleg; and second, to determine whether the resistance to blackleg disease of individual DH lines responds differentially to different L. maculans field populations within and between the two countries. DH lines were extracted from cv. Grouse and tested in field experiments carried out in both France and Australia against natural L. maculans populations. Extracting and screening DH lines were an effective means to select individual lines with greatly improved expression of resistance to blackleg crown canker disease in comparison with the original parental population. However, relative disease resistance rankings for DH lines were not always consistent between sites. The higher level of resistance in France was shown to be because of a high expression level of quantitative resistance in the French growing conditions. Big differences were observed for some DH lines between the 2004 and the 2005 field sites in Australia where the L. maculans populations differed by their virulence on single dominant gene‐based resistant lines derived from Brassica rapa ssp. sylvestris. This differential behaviour could not be clearly explained by the specific resistance genes until now identified in these DH lines. This investigation highlights the potential to derive DH lines with superior levels of resistance to L. maculans compared with parental populations. However, in locations with particularly high pathogen diversity, such as in southern Australia, multiyear and multisite evaluations should be performed to screen for the most efficient material in different situations.     

QTL mapping of multiple foliar disease and root-lesion nematode resistances in wheat

A genetic linkage map, based on a cross between the synthetic hexaploid CPI133872 and the bread wheat cultivar Janz, was established using 111 F₁-derived doubled haploid lines. The population was phenotyped in multiple years and/or locations for seven disease resistance traits, namely, Septoria tritici blotch (Mycosphaeralla graminicola), yellow leaf spot also known as tan spot (Pyrenophora tritici-repentis), stripe rust (Puccinia striiformis f. sp. tritici), leaf rust (Puccinia triticina), stem rust (Puccinia graminis f. sp. tritici) and two species of root-lesion nematode (Pratylenchyus thornei and P. neglectus). The DH population was also scored for coleoptile colour and the presence of the seedling leaf rust resistance gene Lr24. Implementation of a multiple-QTL model identified a tightly linked cluster of foliar disease resistance QTL in chromosome 3DL. Major QTL each for resistance to Septoria tritici blotch and yellow leaf spot were contributed by the synthetic hexaploid parent CPI133872 and linked in repulsion with the coincident Lr24/Sr24 locus carried by parent Janz. This is the first report of linked QTL for Septoria tritici blotch and yellow leaf spot contributed by the same parent. Additional QTL for yellow leaf spot were detected in 5AS and 5BL. Consistent QTL for stripe rust resistance were identified in chromosomes 1BL, 4BL and 7DS, with the QTL in 7DS corresponding to the Yr18/Lr34 region. Three major QTL for P. thornei resistance (2BS, 6DS, 6DL) and two for P. neglectus resistance (2BS, 6DS) were detected. The recombinants combining resistance to Septoria tritici blotch, yellow leaf spot, rust diseases and root-lesion nematodes from parents CPI133872 and Janz constitute valuable germplasm for the transfer of multiple disease resistance into new wheat cultivars.     

Barley 4H QTL confers NFNB resistance to a global set of <Emphasis Type="Italic">P. teres</Emphasis> f. <Emphasis Type="Italic">teres</Emphasis> isolates

Net form net blotch (NFNB), caused by Pyrenophora teres f. teres Drechs., is prevalent in barley-growing regions worldwide. A population of 132 recombinant inbred lines (RILs) developed from a cross of the barley varieties ‘Falcon’ and ‘Azhul’ were used to evaluate resistance to NFNB due to their differential reactions to isolates of P. teres f. teres from Australia, Canada, Japan, and the USA. Falcon is a six-rowed, hulless feed barley harboring resistance to NFNB, while Azhul is a six-rowed, hulless food barley with high levels of susceptibility to many P. teres f. teres isolates. Seedling disease resistance data were collected on seedlings of parents, RILs, and checks in a growth chamber. The population was genotyped using Illumina’s GoldenGate assay, and quantitative trait loci (QTL) were detected on chromosomes 2H, 3H, 4H, and 6H. We identified a single genetic region on barley chromosome 4H that provided varying levels of resistance to all P. teres f. teres isolates evaluated.     

Marker-trait associations in Virginia Tech winter barley identified using genome-wide mapping

Genome-wide association studies (GWAS) provide an opportunity to examine the genetic architecture of quantitatively inherited traits in breeding populations. The objectives of this study were to use GWAS to identify chromosome regions governing traits of importance in six-rowed winter barley (Hordeum vulgare L.) germplasm and to identify single-nucleotide polymorphisms (SNPs) markers that can be implemented in a marker-assisted breeding program. Advanced hulled and hulless lines (329 total) were screened using 3,072 SNPs as a part of the US. Barley Coordinated Agricultural Project (CAP). Phenotypic data collected over 4 years for agronomic and food quality traits and resistance to leaf rust (caused by Puccinia hordei G. Otth), powdery mildew [caused by Blumeria graminis (DC.) E.O. Speer f. sp. hordei Em. Marchal], net blotch (caused by Pyrenophora teres), and spot blotch [caused by Cochliobolus sativus (Ito and Kuribayashi) Drechsler ex Dastur] were analyzed with SNP genotypic data in a GWAS to determine marker-trait associations. Significant SNPs associated with previously described quantitative trait loci (QTL) or genes were identified for heading date on chromosome 3H, test weight on 2H, yield on 7H, grain protein on 5H, polyphenol oxidase activity on 2H and resistance to leaf rust on 2H and 3H, powdery mildew on 1H, 2H and 4H, net blotch on 5H, and spot blotch on 7H. Novel QTL also were identified for agronomic, quality, and disease resistance traits. These SNP-trait associations provide the opportunity to directly select for QTL contributing to multiple traits in breeding programs.     

Necrotrophic effector‐triggered susceptibility (NETS) underlies the barley–Pyrenophora teres f. teres interaction specific to chromosome 6H

Barley net form net blotch (NFNB), caused by the necrotrophic fungus Pyrenophora teres f. teres, is a destructive foliar disease in barley‐growing regions worldwide. Little is known about the genetic and molecular basis of this pathosystem. Here, we identified a small secreted proteinaceous necrotrophic effector (NE), designated PttNE1, from intercellular wash fluids of the susceptible barley line Hector after inoculation with P. teres f. teres isolate 0–1. Using a barley recombinant inbred line (RIL) population developed from a cross between the sensitive/susceptible line Hector and the insensitive/resistant line NDB 112 (HN population), sensitivity to PttNE1, which we have named SPN1, mapped to a common resistance/susceptibility region on barley chromosome 6H. PttNE1–SPN1 interaction accounted for 31% of the disease variation when the HN population was inoculated with the 0–1 isolate. Strong accumulation of hydrogen peroxide and increased levels of electrolyte leakage were associated with the susceptible reaction, but not the resistant reaction. In addition, the HN RIL population was evaluated for its reactions to 10 geographically diverse P. teres f. teres isolates. Quantitative trait locus (QTL) mapping led to the identification of at least 10 genomic regions associated with disease, with chromosomes 3H and 6H harbouring major QTLs for resistance/susceptibility. SPN1 was associated with all the 6H QTLs, except one. Collectively, this information indicates that the barley–P. teres f. teres pathosystem follows, at least partially, an NE‐triggered susceptibility (NETS) model that has been described in other necrotrophic fungal disease systems, especially in the Dothideomycete class of fungi.     

Pyrenophora teres,an agent causing wheat leaf spot

In 2007–2008, the barley net blotch agent Pyrenophora teres was found to infect spring wheat in northwestern Russia, causing symptoms similar to wheat tan spot caused by P. tritici-repentis. The frequency of occurrence of P. teres on spring wheat cultivars was 12–29%. P. teres isolates were more virulent to some wheat cultivars than P. tritici-repentis ones. P. teres was not found on wheat in the south of Russia (Krasnodar krai, Dagestan).