Isolates Of Rhynchosporium Secalis Research Articles

The use of differential isolates of Rhynchosporium secalis to identify resistance to leaf scald in barley

The presence of seedling resistance to leaf scald in Australian barley varieties and breeding lines has been determined using differential pathotypes of Rhynchosporium secalis. The previously described resistance genes Rrs1, Rrs2 and Rh4/Rh10 have been identified as well as other resistance sources that show different reactions against the isolates. Seedling resistance in the barley variety Skiff, RhSkiff, has been found to react in an identical fashion to seedling resistance in the variety Barque suggesting the same gene/allele is present in these varieties. Virulence in R. secalis has been detected against almost all barley resistance sources tested, reinforcing previous experience that single gene resistance to scald is not a safe option. The differential isolates are being used to detect resistance genes in breeding lines as a service to barley breeding programs. They are also being used to detect novel resistances and will prove useful to breeders in building gene pyramids to try and develop varieties with more durable resistance.

Pathotype and microsatellite analyses reveal new sources of resistance to barley scald in Tunisia

We examined the variation and relationships between pathogenicity and a microsatellite-based haplotype in 79 Tunisian Rhynchosporium secalis isolates that were collected from the most commonly cultivated barley populations in Tunisia, Rihane cv. and local landraces, with the goal of finding genes that might be used to monitor resistance to scald. Isolates could be classified into three distinct virulence groups based on artificial inoculation of 19 differential cultivars with known scald resistance genes. The resistance gene BRR2 carried by the Astrix differential cultivar appeared to be the most effective in Tunisia. Pathotypes sampled from the Rihane host were more virulent than those sampled from local barley landraces. Because some differential cultivars that carried the same resistance genes showed different reaction patterns to 48 of the isolates, we postulated that other unknown resistance gene(s) specific to Tunisian isolates may be prevalent and could be used in Tunisian barley breeding programs. Microsatellite fingerprinting allowed the detection of 11 alleles linked to the virulence and pathogenic identification of 52% of the tested isolates. Thus, microsatellite analysis may provide a rapid tool for pathogen detection, without an inoculation step that requires long incubation periods before ultimate disease assessment.

Genetic diversity of Rhynchosporium secalis in Tunisia as revealed by pathotype, AFLP, and microsatellite analyses

Genetic variability among 122 Rhynchosporium secalis isolates collected from barley in three regions of Tunisia was investigated using host differentials, amplified fragment length polymorphism (AFLP), and microsatellite markers. The isolates were collected from a widely grown scald-susceptible barley cultivar Rihane and a range of local landrace cultivars in geographically distinct regions with different agroclimatic conditions. Pathotypic diversity (the proportion of unique pathotypes) was high in R. secalis populations from the high (100% diversity), moderate (95%), and low (100%) rainfall areas of Tunisia, and from both Rihane (which is the sole variety grown in the high rainfall region) and local landraces (which predominate in the low rainfall area). This may reflect a general adaptability for aggressiveness and suggests that the widely grown cultivar Rihane has exerted little or no selection pressure on the pathogen population since its release in 1983. Genotypic diversity (GD), defined as the probability that two individuals taken at random had different genotypes, was high for populations from Rihane, local landraces, and different agro-ecological zones (GD = 0.96-0.99). There was low genetic differentiation among pathogen populations from different host populations (G(ST) < or = 0.08, theta < or = 0.12) and agro-ecological zones (G(ST) < or = 0.05, theta < or = 0.04), which may be partly explained by gene flow due to the movement of infected stubble around the country. There was no correlation (r = 0.06, P = 0.39) between virulence phenotype and AFLP haplotype. A phenetic tree revealed groups with low bootstrap values that did not reflect the grouping of isolates based on host, pathotype, or agro-ecological region. The implications of these findings for R. secalis evolutionary potential and scald-resistance breeding in Tunisia are discussed.

Differential Selection on Rhynchosporium secalis During Parasitic and Saprophytic Phases in the Barley Scald Disease Cycle

ABSTRACT Competition among eight Rhynchosporium secalis isolates was assessed during parasitic and saprophytic phases of the disease cycle in field experiments conducted at two locations and over two growing seasons. The eight isolates were inoculated onto six barley populations exhibiting varying degrees of resistance. Microsatellite analysis of 2,866 isolates recovered from the field experiments showed significant, and sometimes opposite, changes in the frequencies of R. secalis genotypes during the growing season (parasitic phase) and between growing seasons (saprophytic phase). Isolates that showed the most complex virulence in greenhouse seedling assays had the lowest fitness in the field experiment. Significant differences in isolate fitness were found on different host populations and in different environments. Selection coefficients were large, indicating that evolution can occur rapidly in field populations. Although inoculated isolates had the lowest overall fitness on the moderately resistant landrace cv. Arabi Aswad, some isolates were more virulent and consistently increased in frequency on this landrace, suggesting a risk of directional selection and possible erosion of the resistance following its widespread deployment in monoculture. These results provide the first direct evidence that R. secalis pathogen genotypes differ in their saprophytic ability and parasitic fitness under field conditions.

Global Hierarchical Gene Diversity Analysis Suggests the Fertile Crescent Is Not the Center of Origin of the Barley Scald PathogenRhynchosporium secalis

A total of 1,366 Rhynchosporium secalis isolates causing scald on barley, rye, and wild barley (Hordeum spontaneum) were assayed for restriction fragment length polymorphism loci, DNA fingerprints, and mating type, to characterize global genetic structure. The isolates originated from 31 field populations on five continents. Hierarchical analysis revealed that more than 70% of the total genetic variation within regions was distributed within a barley field. At the global level, only 58% of the total genetic variation was distributed within fields, while 11% was distributed among fields within regions, and 31% was distributed among regions. A significant correlation was found between genetic and geographic distance. These findings suggest that gene flow is common at the local level while it is low between regions on the same continent, and rare between continents. Analyses of multilocus associations, genotype diversity, and mating type frequencies indicate that sexual recombination is occurring in most of the populations. We found the highest allele richness in Scandinavia followed by Switzerland. This suggests that R. secalis may not have originated at the center of origin of barley, the Fertile Crescent, nor in a secondary center of diversity of barley, Ethiopia.

Molecular Analysis of Rhynchosporium secalis Population Collected from Barley Cultivars having Different Resistance Specificities

Molecular analysis was performed to detect genetic diversity in 106 Rhynchosporium secalis isolates collected from different regions of Canada using random amplified polymorphic DNA (RAPD) markers. The isolates collected from barley cultivars having different resistance specificity to R. secalis and grown in geographically distinct regions, exhibited reproducible variation for 2–3 polymorphic PCR products per decamer primer. Analysis of 1960 RAPD markers data obtained with five primers formed 5 groups with different genetic similarity. High genetic variation was observed in R. secalis isolates obtained from resistant and susceptible cultivars of barley. Isolates collected from susceptible cultivars showed a tendency to group together, whereas isolates from resistant cultivars were divergent. R. secalis isolates infecting different barley cultivars released as resistant to the barley scald formed a specific group with UPGMA, even though all these isolates were collected from the same epidemiological region. Analysis of 15 isolates collected from one resistant cultivar Duke formed three clusters with low bootstrap values indicating high genetic diversity among the isolates present on a single host cultivar.

Pathogenic Variation among Isolates of Rhynchosporium secalis from Cultivated Barley Growing in Central Anatolia, Turkey

Pathogenic Variation among Isolates of Rhynchosporium secalis from Cultivated Barley Growing in Central Anatolia, Turkey

Amplified fragment length polymorphism among Rhynchosporium secalis isolates collected from a single barley field in Syria

SummaryAFLP markers were used to measure the amount and distribution of genetic variation among Rhynchosporium secalis isolates on a microgeographical scale in Syria. Forty isolates hierarchically sampled from a single barley field were assayed for AFLP variation using primer combinations not previously tested in populations of the pathogen from Syria. In contrast to a previous study, which showed high clonality within field populations of R. secalis in Syria, the present study revealed a much higher level of genetic diversity, stressing the important roles that sampling strategies and the choice of primers/primer combinations play in the evaluation of genetic variation in R. secalis populations at a microgeographical scale. A high level of genetic variation was found to occur on a fine scale throughout the pathogen population examined, with 40 different haplotypes being identified among the 40 isolates sampled. Data were consistent with the hypothesis that the primary inoculum originated from a genetically diverse founding population, which may have consisted of ascospores of an as yet undescribed teleomorph and/or asexual spores of a highly mutable local population.

Alberta Regional Meeting 2004

Alberta Regional Meeting 2004

The effect of fungicide programmes based on epoxiconazole on the control and DMI sensitivity of Rhynchosporium secalis in winter barley

To prolong the effective life of the newer, more active DeMethylation Inhibitor (DMI) fungicides, two-spray programmes were evaluated for their effectiveness in combining good control of rhynchosporium on winter barley with prevention of the build-up of DMI-resistant pathogen strains. Programmes were based on the DMI epoxiconazole alone or in combination or alternation with three partner fungicides with different modes of action, fenpropimorph (a morpholine), cyprodinil (an anilinopyrimidine), azoxystrobin (a QoI/strobilurin fungicide). In the majority of six field trials in Northern Ireland and South-west England (1998–2000), DMI sensitivity of Rhynchosporium secalis isolates was lower after fungicide treatment than before and the least sensitive isolates came from the plots treated with two half-rate applications of epoxiconazole alone. All fungicide programmes reduced disease compared with the untreated control. Epoxiconazole used alone gave the poorest disease control, but did increase the yield. The three partner fungicides all improved disease control and yield; two applications of half-rate epoxiconazole with half-rate azoxystrobin or cyprodinil performed best overall. With the range of pathogen sensitivities found in the present study, a DMI such as epoxiconazole remains a useful component of a control programme for R. secalis, but should be supplemented by a partner fungicide with a different mode of action such as a strobilurin or anilinopyrimidine.

Assessment of the Seedling Reactions of Turkish Barley Cultivars to Scald

AbstractThe reactions of 37 cultivars and two candidate lines of barley to five isolates of Rhynchosporium secalis and their mixtures were determined in assays under greenhouse conditions. Fungal isolates were collected from Ankara province and surrounding areas in Turkey. There were clear differences in the reactions of the cultivars to the fungal isolates and isolates differed in pathogenicity for each cultivar. Of all genotypes tested, seven cultivars (cvs Erginel 90, Şahin 91, Kıral 97, Akhisar 98, Çetin 2000, Çumra 2001, Avcı 2002) and one candidate line (candidate 4) were found to be resistant to all isolates and their mixtures. A5 was found to be the most virulent fungal isolate.

Dynamics of Rhynchosporium secalis pathotypes in relation to barley cultivar resistance

Rhynchosporium secalis isolates E97-2 and H97-2, represented the major pathotypes in populations on barley in Alberta, Canada, but differed widely in their virulence. Following greenhouse co-inoculation with the two pathotypes, E97-2, originally isolated from resistant cv. 'CDC Earl', predominated over H97-2, isolated from the susceptible cv. 'Harrington', from the first to the last of four infection cycles on both 'CDC Earl' and 'Harrington'. These results indicated that the host can rapidly influence pathotype composition and that pathotype E97-2 may have a competitive advantage over H97-2 on these cultivars. DNA polymorphisms were found between isolates from single or mixed inoculations on cvs 'CDC Earl' and 'Harrington' for four successive cycles. Co-inoculation with the two isolates resulted in a shift to a molecular phenotype more similar to E97-2 than H97-2. The competitive advantage of E97-2 over H97-2, combined with the selective pressure exerted by the host, would explain the increased susceptibility of cv. 'CDC Earl' and other cultivars with similar sources of scald resistance, in fields across Alberta. However, H97-2 will likely remain one of the major pathotypes in Alberta due to the relatively large acreage of cv. 'Harrington' in this province.

The effect of fungicide programmes based on epoxiconazole on the control and DMI sensitivity of Rhynchosporium secalis in winter barley

To prolong the effective life of the newer, more active DeMethylation Inhibitor (DMI) fungicides, two-spray programmes were evaluated for their effectiveness in combining good control of rhynchosporium on winter barley with prevention of the build-up of DMI-resistant pathogen strains. Programmes were based on the DMI epoxiconazole alone or in combination or alternation with three partner fungicides with different modes of action, fenpropimorph (a morpholine), cyprodinil (an anilinopyrimidine), azoxystrobin (a QoI/strobilurin fungicide). In the majority of six field trials in Northern Ireland and South-west England (1998–2000), DMI sensitivity of Rhynchosporium secalis isolates was lower after fungicide treatment than before and the least sensitive isolates came from the plots treated with two half-rate applications of epoxiconazole alone. All fungicide programmes reduced disease compared with the untreated control. Epoxiconazole used alone gave the poorest disease control, but did increase the yield. The three partner fungicides all improved disease control and yield; two applications of half-rate epoxiconazole with half-rate azoxystrobin or cyprodinil performed best overall. With the range of pathogen sensitivities found in the present study, a DMI such as epoxiconazole remains a useful component of a control programme for R. secalis, but should be supplemented by a partner fungicide with a different mode of action such as a strobilurin or anilinopyrimidine.

Further evidence for sexual reproduction in Rhynchosporium secalis based on distribution and frequency of mating-type alleles

Rhynchosporium secalis, the causal agent of scald on barley, is thought to be exclusively asexual because no teleomorph has been found. Partial sequences of the HMG-box and α-domain of Rhynchosporium secalis isolates were identified and used to develop a PCR assay for the mating-type locus. PCR amplification of only one of these two domains was possible in each strain, suggesting that R. secalis has a MAT organization that is similar to other known heterothallic fungi. A multiplex PCR with primers amplifying either a MAT1-1- or MAT1-2-specific amplicon was used to determine the distribution of mating types in several R. secalis populations. In total, 1101 isolates from Australia, Switzerland, Ethiopia, Scandinavia, California, and South Africa were included in the analysis. Mating types occurred in equal frequencies for most of these populations, suggesting frequency-dependent selection consistent with sexual reproduction. In addition, both mating types were frequently found occupying the same lesion or leaf, providing opportunities for isolates of opposite mating type to interact and reproduce sexually. We propose that R. secalis should be considered a sexual pathogen, although the sexual cycle may occur infrequently in some populations.

Molecular variation in Rhynchosporium secalis isolates obtained from hotspots

A unique sampling strategy was used to search for genetic similarity among isolates of Rhynchosporium secalis, the causal agent of leaf scald of barley. A field of barley cv. Skiff with discrete ‘hotspots’ of leaf scald was identified and infected leaves were collected for this study. The isolates were genotyped using amplified fragment length polymorphism (AFLP) analysis. In contrast to previous reports of high diversity of R. secalis isolates in areas of only 1 m2, we found close genetic similarity between isolates from the same hotspot as determined by AFLP marker alleles, with a higher level of variation between isolates from different hotspots. A UPGMA phenogram showed that most isolates clustered with members of the same hotspot. This pattern is consistent with the inoculum source for the hotspots being airborne ascospores or wind-dispersed conidia from a local founder population, borne in splash-created aerosols. AFLP variation within hotspots suggests a high rate of mutation within a few cycles of fungal infection. A collection of isolates from cv. Skiff plots in a straw-inoculated disease nursery 60 km from the hotspot field was fingerprinted with AFLPs for comparison and was found to have much greater genotypic diversity.

Resistance to Scald (Rhynchosporium secalis) in Barley (Hordeum vulgare) Studied by Near-Isogenic Lines: I. Markers and Differential Isolates

ABSTRACT Near-isogenic lines (NILs) with resistance for scald in seventh generation backcross with 'Ingrid' as recurrent parent (RP) were tested with seven differential isolates of Rhynchosporium secalis in Norway and Canada. NILs of 'Turk', 'Brier', 'CI 8162', 'La Mesita', 'Hispont', 'Atlas 46', 'Modoc', 'Hudson', 'Abyssinian', 'Steudelli', and 'CI 2222' also were evaluated for field reactions. The genetic characterization of the NILs (degree of isogeneity with Ingrid) and with each other was carried out. The molecular marker pattern shows that the backcrossing program has resulted in from 86.3 to 100% RP genome in the NILs, depending on the marker system. On an average, 96% RP genome was found in the NILs. There were certain consistent (pairwise) differences between the NILs and RP on chromosomes 3H and 7H. Both chromosomes are known to contain loci conferring resistance to R. secalis, indicating successful introgression from the donors into the NILs. Approximately two-thirds of the observed RP-NIL polymorphisms were linked to the assumed resistance in the NIL. Based on the marker and phenotypic analyses of the NILs, suggestions for a more appropriate and updated terminology of genes for resistance to R. secalis in barley are made. The proposed changes in nomenclature also indicate the differentials that are available as NILs and those lacking.

Pathogenic variation of Rhynchosporium secalis in Alberta

Leaf samples with scald symptoms were taken from various barley cultivars in 1997 and 1998 at nine locations in Alberta for examination of pathogenic variability. Two hundred and fifty-six single-spore isolates of Rhynchosporium secalis were differentiated into 52 pathotypes using 12 differentials, which consisted of seven accessions with major resistance genes and five commercial cultivars. Fifty-two percent of isolates were virulent on cv. Harrington only (pathotype 1); five pathotypes, consisting of close to 25% of isolates, were virulent only on 4 of 5 commercial cultivars; and about 45 pathotypes, consisting of about 25% of isolates, were virulent on commercial cultivars and accessions. No differential was resistant to all pathotypes, and many pathotypes were represented by single isolates. There was a difference in pathotype diversity and complexity among locations in Alberta. Calmar, Lacombe, and Edmonton sites comprised the greatest number of pathotypes that were virulent on up to seven differentials; at the Beaverlodge, Carstairs, Stettler, and Westlock sites, pathotype 1 comprised more than 60% of total isolates with a few other pathotypes. The Trochu and Vegreville sites were intermediate in the number of pathotypes. Discriminant analysis based on the reactions of the 12 differentials also showed divergence in R. secalis virulence among locations. Pathogenic variability associated with location in Alberta is important in the context of breeding for resistance and use of the most resistant cultivars by producers.

Mapping of a gene for leaf scald resistance in barley line ‘B87/14’ and validation of microsatellite and RFLP markers for marker‐assisted selection

AbstractSeedlings of the barley line ‘B87/14’ were resistant to 22 out of 23 Australian isolates of Rhynchosporium secalis, the causal agent of leaf scald.‘B87/14’‐based populations were developed to determine the location of the resistance locus. Scald resistance segregated as a single dominant trait in BC1F2 and BC1F3 populations. Bulked segregant analysis identified amplified fragment length polymorphisms (AFLPs) with close linkage to the resistance locus. Fully mapped populations not segregating for scald resistance located these AFLP markers on chromosome 3H, possibly within the complex Rrs1 scald locus. Microsatellite and restriction fragment length polymorphism markers adjacent to the AFLP markers were identified and validated for their linkage to scald resistance in a second segregating population, with the closest marker 2.2 cM from the resistance locus. These markers can be used for selection of the Rrs.B87 scald‐resistance locus, and other genes at the chromosome 3H Rrs1 locus.

DMI sensitivity and cross-resistance patterns of Rhynchosporium secalis isolates from South Africa

Isolates of R. secalis were collected yearly from the Rûens area of the Western Cape during the 1993–1995 growing seasons. These isolates were evaluated in vitro to determine sensitivity to triazole fungicides (triadimenol, tebuconazole, flusilazole and propiconazole). The sensitivity fluctuated but in 1995 isolates were significantly less sensitive towards triadimenol than in the previous two years. In a second experiment, isolates collected from two fields with a 5–6 year history of triadimenol seed treatments and tebuconazole applications were evaluated for their fungicide sensitivity. A significant positive correlation was observed between tebuconazole and triadimenol sensitivity among R. secalis isolates from these fields. However, such a correlation was not found within the R. secalis population collected during 1993–1995 where shorter crop rotation patterns and a range of fungicides were applied. In a third experiment, the fungicide sensitivity of local R. secalis isolates was evaluated towards two new triazole fungicides, namely bromuconazole and triticonazole. Correlation coefficients observed between these new triazoles and those previously applied in South Africa were not significantly positive. The lack of significant cross-resistance has important practical implications for management of fungicide resistance.

Pathotypes and NIP1 gene sequences of Finnish Rhynchosporium secalis isolates from barley, couch grass and rye

Rhynchosporium secalis (Oud.) J.J. Davis, causal agent of scald of barley (Hordeum vulgare L.), was isolated from barley, couch grass (Elymus repens L.) and rye (Secale cereale L.). Isolates were used to inoculate seedlings of a differential barley series containing several sources of major gene resistance to the disease. The series included Atlas46, (resistance gene Rrs1) and the isogenic line Atlas, which lacks the gene (rrs1). The necrosis-inducing peptide NIP1 has been suggested to be the product of the avirulence gene AvrRrs1 (NIP1) that with the barley resistance gene Rrs1 determines the incompatibility of the pathogen-host interaction. All R. secalis isolates were virulent only on the susceptible barley cultivars Arve and Chan, irrespective of spore concentration or original host species. There were no indications of redundant virulence among the R. secalis isolates. The NIP1 gene was sequenced from each isolate and there was no correlation with source of the isolate or sequence modification and virulence. Four isolates, from barley and couch grass, were characterised by a basic Type I NIP1sequence. The recorded NIP1 sequence changes are consistent withR. secalis populations not receiving selection through deployment of the Rrs1 gene in commercial barley cultivars in Finland.