Secalis Isolates Research Articles

Leaf rust (Puccinia recondita f. sp. secalis) triggers substantial changes in rye (Secale cereale L.) at the transcriptome and metabolome levels

BackgroundRye (Secale cereale L.) is a cereal crop highly tolerant to environmental stresses, including abiotic and biotic stresses (e.g., fungal diseases). Among these fungal diseases, leaf rust (LR) is a major threat to rye production. Despite extensive research, the genetic basis of the rye immune response to LR remains unclear.ResultsAn RNA-seq analysis was conducted to examine the immune response of three unrelated rye inbred lines (D33, D39, and L318) infected with compatible and incompatible Puccinia recondita f. sp. secalis (Prs) isolates. In total, 877 unique differentially expressed genes (DEGs) were identified at 20 and 36 h post-treatment (hpt). Most of the DEGs were up-regulated. Two lines (D39 and L318) had more up-regulated genes than down-regulated genes, whereas the opposite trend was observed for line D33. The functional classification of the DEGs helped identify the largest gene groups regulated by LR. Notably, these groups included several DEGs encoding cytochrome P450, receptor-like kinases, methylesterases, pathogenesis-related protein-1, xyloglucan endotransglucosylases/hydrolases, and peroxidases.The metabolomic response was highly conserved among the genotypes, with line D33 displaying the most genotype-specific changes in secondary metabolites. The effect of pathogen compatibility on metabolomic changes was less than the effects of the time-points and genotypes. Accordingly, the secondary metabolome of rye is altered by the recognition of the pathogen rather than by a successful infection. The results of the enrichment analysis of the DEGs and differentially accumulated metabolites (DAMs) reflected the involvement of phenylpropanoid and diterpenoid biosynthesis as well as thiamine metabolism in the rye immune response.ConclusionOur work provides novel insights into the genetic and metabolic responses of rye to LR. Numerous immune response-related DEGs and DAMs were identified, thereby clarifying the mechanisms underlying the rye response to compatible and incompatible Prs isolates during the early stages of LR development. The integration of transcriptomic and metabolomic analyses elucidated the contributions of phenylpropanoid biosynthesis and flavonoid pathways to the rye immune response to Prs. This combined analysis of omics data provides valuable insights relevant for future research conducted to enhance rye resistance to LR.

НАСЛЕДОВАНИЕ УСТОЙЧИВОСТИ К РИНХОСПОРИОЗУ У ЧЕТЫРЕХ ОБРАЗЦОВ МЕСТНОГО ЯЧМЕНЯ

Background. Scald (caused by Rhynchosporium secalis (Oudem.) J.J. Davis) is one of the most harmful barley diseases. The pathogen is characterized by the differential interaction with the host plant genotypes. Therefore, the most rational plant breeding strategy should envisage broadening of genetic diversity of cultivated varieties. It has been found that the majority of the recently released barley varieties and of the previously identified sources of resistance to R. secalis are now susceptible to the pathogen. Only the resistance gene Rrs9 maintains efficiency against the pathogen populations in the Russian Northwest. Earlier, we have found scald resistant barleys to be frequent among barley landrace accessions. It was demonstrated by the hybridological analysis that accessions k-3307, k-15868, k-18989 and k-3481 are protected by effective scald resistance genes that differ from each other, and which are not allelic to the Rrs4, rrs6, rrs7 or Rrs9 genes. The goal of the present work was to study genetic control to the dangerous pathogen in four other barley landraces. Materials and methods . The inheritance of scald resistance was analyzed in barley accessions k-16233, k-27205 (China), k-27768 (India) and k-22299 (Ecuador). The study employed two experimental techniques, that is, the pathogen/host plant differential interactions analysis in the lab, and the hybridological analysis in the field. The experiments were carried out using R. secalis isolates and clones from the pathogen populations sampled in the North Caucasus and the Russian Northwest. Resistance was scored against the scales from 0 (no symptoms) to 4 points (in the lab), and from 0 to 5 points (in the field). Plants that scored 0-2 points were regarded as resistant. Results and conclusions . The analysis of the pathogen clone/host plant interaction has shown that the landraces k-16233, k-27205 (China), k-27768 (India) and k-22299 (Ecuador) are protected by effective genes of scald resistance which differ from each other and are non-allelic to the previously effective genes Rrs4, rrs6, rrs7 and the currently effective Rrs9. According to the hybridological analysis data, k-27205 has one dominant gene of scald resistance; k-16233, k-27768 and k-22299 are protected by two dominant effective resistance genes, all of which manifest themselves at all the stages of plant ontogenesis.

Non-parent of Origin Expression of Numerous Effector Genes Indicates a Role of Gene Regulation in Host Adaption of the Hybrid Triticale Powdery Mildew Pathogen.

Powdery mildew is an important disease of cereals. It is caused by one species, Blumeria graminis, which is divided into formae speciales each of which is highly specialized to one host. Recently, a new form capable of growing on triticale (B.g. triticale) has emerged through hybridization between wheat and rye mildews (B.g. tritici and B.g. secalis, respectively). In this work, we used RNA sequencing to study the molecular basis of host adaptation in B.g. triticale. We analyzed gene expression in three B.g. tritici isolates, two B.g. secalis isolates and two B.g. triticale isolates and identified a core set of putative effector genes that are highly expressed in all formae speciales. We also found that the genes differentially expressed between isolates of the same form as well as between different formae speciales were enriched in putative effectors. Their coding genes belong to several families including some which contain known members of mildew avirulence (Avr) and suppressor (Svr) genes. Based on these findings we propose that effectors play an important role in host adaptation that is mechanistically based on Avr-Resistance gene-Svr interactions. We also found that gene expression in the B.g. triticale hybrid is mostly conserved with the parent-of-origin, but some genes inherited from B.g. tritici showed a B.g. secalis-like expression. Finally, we identified 11 unambiguous cases of putative effector genes with hybrid-specific, non-parent of origin gene expression, and we propose that they are possible determinants of host specialization in triticale mildew. These data suggest that altered expression of multiple effector genes, in particular Avr and Svr related factors, might play a role in mildew host adaptation based on hybridization.

Two new species of Rhynchosporium

Rhynchosporium consists of two species, R. secalis and R. orthosporum. Both are pathogens of grasses with R. secalis infecting a variety of Poaceae hosts and R. orthosporum infecting Dactylis glomerata. Phylogenetic analyses of multilocus DNA sequence data on R. secalis isolates originating from cultivated barley, rye, triticale and other grasses, including Agropyron spp., Bromus diandrus and Hordeum spp., resolved the monophyletic groups into three species according to their respective hosts. Host specificity according to phylogenetic lineages was confirmed with pathogenicity studies. Because R. secalis was described first on rye this name is retained for Rhynchosporium isolates infecting rye and triticale. Rhynchosporium isolates infecting cultivated barley and other Hordeum spp. and Bromus diandrus belong to a distinct species, R. commune. Similarly isolates infecting Agropyron spp. represent a distinct species of Rhynchosporium, namely R. agropyri. A PCR-RFLP assay was developed as a rapid tool for species identification of R. secalis and R. commune.

Infection of Rrs1 barley by an incompatible race of the fungus Rhynchosporium secalis expressing the green fluorescent protein

Scald disease of barley, caused by the fungal pathogen Rhynchosporium secalis, is one of the most serious diseases of this crop worldwide. Disease control is achieved in part by deployment of major resistance (Rrs) genes in barley. However, in both susceptible and resistant barley plants, R. secalis is able to complete a symptomless infection cycle. To examine the R. secalis infection cycle, Agrobacterium tumefaciens‐mediated transformation was used to generate R. secalis isolates expressing the green fluorescent protein or DsRed fluorescent protein, and that were virulent on an Rrs2 plant (cv. Atlas), but avirulent on an Rrs1 plant (cv. Atlas 46). Confocal laser scanning microscopy revealed that R. secalis infected the susceptible cultivar and formed an extensive hyphal network that followed the anticlinal cell walls of epidermal cells. In the resistant cultivar, hyphal development was more restricted and random in direction of growth. In contrast to earlier models of R. secalis infection, epidermal collapse was not observed until approximately 10 days post‐inoculation in both cultivars. Sporulation of R. secalis was observed in both susceptible and resistant interactions. Observations made using the GFP‐expressing isolate were complemented and confirmed using a combination of the fluorescent probes 5‐chloromethylfluorescein diacetate and propidium iodide, in the non‐transformed wild‐type isolate. The findings will enable the different Rrs genes to be better characterized in the effect they exert on pathogen growth and may aid in identification of the most effective 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.

Virulence spectrum to barley (Hordeum vulgare L.) in isolates of Rhynchosporium secalis from Syria

Scald caused by Rhynchosporium secalis, is an economically important disease found worldwide. Information on the variability of virulence in R. secalis is essential for the development of spot-resistant barley cultivars. In order to establish the number and distribution of R. secalis virulence types, 115 isolates collected during 1997–2006 from major barley growing areas in Syria were evaluated on ten differential barley genotypes. Eleven pathotypes (Pt1–11) were identified based on the infection response of the genotypes with mean disease rating from 1.7 to 4.2. Cultivar had a significant influence (P < 0.001) on the severity of scald for all pathotypes tested. Pathotype Pt9 exhibited low virulence on all used genotypes, and pathotype Pt6 was highly virulent. The two most common pathotypes were Pt1 (22 isolates, 19.1%) and Pt3 (24 isolates, 20.9%). The pathotype distribution suggests there has been previous dispersal of R. secalis isolates across the country. To incorporate adequate levels of resistance into future barley genotypes, disease evaluations should be made with R. secalis isolates that express the full spectrum of virulence found in Syria.

Molecular evidence for recent founder populations and human-mediated migration in the barley scald pathogen Rhynchosporium secalis

Rhynchosporium secalis is an important pathogen of barley globally. Fourteen polymorphic microsatellites were analyzed for 1664 R. secalis isolates sampled from 37 field populations to infer their demographic history. The results falsified the hypothesis that R. secalis co-evolved with its barley host in the Middle East. Populations from Scandinavia had significantly higher allelic diversities, the greatest number of private alleles and the highest genotypic diversities. All but three of the analyzed populations had an excess of gene diversity compared to the number of alleles, consistent with a recent population bottleneck. The remaining populations had a gene diversity deficit consistent with a population expansion following a recent population bottleneck in the last ±100 years. A coalescent analysis revealed that the effective population sizes based on θ, of the analyzed populations were small relative to their ancestral population sizes, indicating that only a fraction of the diversity present in the ancestral populations was transmitted into current populations. These findings are consistent with the hypothesis that the pathogen population on barley experienced a selection bottleneck imposed by the host and/or are founder populations. The mean estimate of migration rates was 2.2 (avg 90% confidence interval = 1.3–3.1). Major migration routes were identified among populations separated by long distances, eg between South Africa and Australia, as well as among North Africa, the Middle East and California, suggesting contemporary exchange of infected barley seed. In contrast with earlier findings, most populations exhibited significant gametic disequilibrium, probably as a result of genetic drift. We conclude that the majority of R. secalis populations have experienced human-mediated migration that led to numerous and relatively recent founder events around the world.

Sequence conservation in the mitochondrial cytochrome b gene and lack of G143A QoI resistance allele in a global sample ofRhynchosporium secalis

Barley scald caused by Rhynchosporium secalis is often controlled by fungicides, most recently by those in the strobilurin-based (QoI) class. Since the launch of QoIs in 1996 a range of important plant pathogens, including Blumeria graminis, Mycosphaerella fijiensis and Plasmopara viticola, developed resistance. Present monitoring data indicate that R. secalis populations remain sensitive. The primary molecular mechanism of QoI resistance in several fungi is a point mutation at codon 143 in the mitochondrial-encoded cytochrome b gene (cytb), which causes the substitution of glycine by alanine (G143A). We characterised the cytb gene of R. secalis, assessed the intraspecific and interspecific sequence diversity, developed a PCR-RFLP diagnostic tool to detect the most common allele associated with QoI resistance, screened a global collection of 841 R. secalis isolates for this allele and tested a representative sample of isolates for QoI resistance in vitro. The results indicated a high degree of conservation for the cytb gene at both intra- and interspecific levels, and complete QoI sensitivity in all R. secalis populations tested.

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.

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.

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.

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 L.). II. Diallel analysis of near-isogenic lines

Near-isogenic lines (NILs) in the BC7 generation for resistance to barley scald (Rhynchosporium secalis Oudem. J.J. Davis) have been developed with ‘Ingrid’ as the recurrent parent (RP). Starting from 23 differential varieties and a classical BC program with disease tests followed by selection for the RP phenotype, NILs from the following 9 sources have been selected for analysis: ‘Turk’, ‘Brier’, ‘CI 8162’, ‘La Mesita’, ‘Hispont’, ‘Atlas46’, ‘Modoc’, ‘Hudson’ and ‘Abyssinian’. The 9 NILs were crossed in a half-diallel design. The NILs, their F1s and F2s were tested along with the donors and RP against R. secalis isolates ‘4004’ and ‘2’ to understand the degree of resistance in each NIL, allelism/linkage and dominance relationships among the NILs. With isolate ‘4004’, 13 of the 36 crosses segregated in the F2 generation. A common feature was that either ‘Atlas 46’-NIL or ‘Hudson’-NIL was one of the parents. Thus it seems that 7 out of the 9 lines are allelic, probably at the Rrs1 complex locus on chromosome 3H, one carries the Rrs2 locus on chromosome 7H and one is independent of both. Segregation with isolate ‘2’ was harder to establish because of a weak resistance gene in the RP which acted additively with the other resistance genes and resulted in a narrower phenotypic range. Degree of dominance/recessiveness depended on the crosses as well as the isolate used. In general GCA effects were much stronger than SCA.