Aggressiveness of Mycosphaerella graminicola Isolates from Susceptible and Partially Resistant Wheat Cultivars

ABSTRACT The selective effect of quantitative host resistance on pathogen aggressiveness is poorly understood. Because two previous experiments with a small number of bread wheat cultivars and isolates of Mycosphaerella graminicola had indicated that more susceptible hosts selected for more aggressive isolates, we conducted a larger experiment to test that hypothesis. In each of 2 years, six cultivars differing in their levels of partial resistance were planted in field plots, and isolates were collected from each cultivar early and late in the growing season. The isolates were inoculated as populations bulked by cultivar of origin, field replicate, and collection date on seedlings of the same six cultivars in the greenhouse. The selective impact of a cultivar on aggressiveness was measured as the difference in aggressiveness between early and late isolates from that cultivar. Regression of those differences on disease severity in the field yielded significance values of 0.0531 and 0.0037 for the 2 years, with moderately resistant cultivars selecting for more aggressive isolates. In a related experiment, the protectant fungicide chlorothalonil was applied to plots of two susceptible cultivars to retard epidemic development. When tested in the greenhouse, isolates of M. graminicola from those plots were significantly more aggressive than isolates from the same cultivars unprotected by fungicide.     

Measuring Immigration and Sexual Reproduction in Field Populations of Mycosphaerella graminicola

ABSTRACT A field experiment was conducted to determine the relative contributions of immigration and sexual reproduction to the genetic structure of Mycosphaerella graminicola populations during the course of an epidemic. The genetic structure of M. graminicola populations sampled from wheat plots inoculated artificially with 10 isolates was compared with control plots infected naturally by airborne ascospores. Restriction fragment length polymorphisms (RFLPs) were used to test the randomness of associations among loci, and DNA fingerprints were used to identify clones. All isolates in the control plots had unique genotypes and RFLP loci were at gametic equilibrium, findings consistent with random mating. The proportion of isolates in the inoculated plots with DNA fingerprints that differed from the 10 inoculated isolates increased from 3% in the early to 39 and 34% in the mid- and late season, respectively. The degree of gametic disequilibrium was higher in the mid-season than in the late-season population. By the end of the growing season, we estimate that 66% of the isolates in the inoculated plots were asexual progeny of the 10 inoculated isolates, 10% were immigrants, and 24% were sexual recombinants. The proportion of infections caused by ascospores increased over the growing season.     

Effects of Infection by Mycosphaerella graminicola on Translocation of Fluquinconazole in Wheat Seedlings

ABSTRACT Translocation of (14)C-labeled fluquinconazole was measured using combustion analysis and radio thin-layer-chromatographic analysis in seedling wheat leaves uninfected and infected with Mycosphaerella graminicola. Two isolates were used with differing sensitivity to demethylation inhibitor fungicides. Fluquinconazole was translocated acropetally, but not basipetally. Fluquinconazole accumulated around infection sites within 6 days after treatment. Accumulation occurred before M. graminicola hyphae had colonized the host mesophyll further than one host cell around the invasion site. This suggested that the accumulation was caused by a host response to infection. Infrared gas analysis showed that rates of transpiration and stomatal conductance in inoculated leaves were significantly increased very soon after inoculation but net photosynthesis was decreased. The actual mechanism of fungicide accumulation was not determined.     

Inheritance of Race-Specific Resistance to Mycosphaerella graminicola in Wheat

ABSTRACT Mycosphaerella graminicola causes Septoria tritici blotch of hexaploid and tetraploid wheat. The inheritance of high-level resistance to Septoria tritici blotch was studied in controlled environment experiments. Intraspecific reciprocal crosses were made between hexaploid wheat lines Salamouni, ST6, Katepwa, and Erik, and the tetraploid wheat lines Coulter and 4B1149. Parental, F(1), F(2), F(3), BC(1)F(1), and BC(1)F(2) populations were evaluated for reaction to isolates MG2 and MG96-36 of M. graminicola. Resistance was controlled by incompletely dominant nuclear genes in all cases. Salamouni had three independent resistance genes to isolate MG2, two of which also controlled resistance to isolate MG96-36. ST6 had a single resistance gene to isolate MG2 and none to isolate MG96-36. The resistance genes in Salamouni and ST6 were not allelic. Two independent genes control resistance to isolate MG2 in Coulter, one of which also controlled resistance to isolate MG96-36. These data are consistent with a gene-for-gene interaction in the wheat-M. graminicola pathosystem.     

Partial Resistance to Septoria Tritici Blotch (Mycosphaerella graminicola) in Wheat Cultivars Arina and Riband

ABSTRACT Partial resistance to Septoria tritici blotch (STB) and its inheritance were investigated in a doubled-haploid population of a cross between cvs. Arina and Riband. The former has good partial resistance whereas the latter is susceptible. In adult plant trials in polytunnels, STB disease scores were negatively correlated with heading date. Resistance was not specific to any of the three fungal isolates used in these tests. A quantitative trait locus (QTL) for partial resistance to STB was identified in Riband on chromosome 6B and is named QStb.psr-6B-1. No QTL controlling a major part of the Arina resistance was identified, suggesting that its resistance may be dispersed and polygenic. There was no correlation between the lines' mean disease scores at the seedling and adult stages, implying that partial resistance to STB is developmentally regulated. Seedling resistance to the isolate IPO323 was isolate-specific and controlled by a single gene in Arina, probably allelic with the Stb6 gene in cv. Flame that confers resistance to the same isolate. The implications of these results for wheat breeding programs are discussed.     

Frequency of sexual recombination by Mycosphaerella graminicola in mild and severe epidemics

The importance of sexual recombination in determining fungal population structure cannot be inferred solely from the relative abundance of sexual and asexual spores and reproductive structures. To complement a previously reported study of proportions of Mycosphaerella graminicola ascocarps and pycnidia, we investigated the share of sexual recombinants among isolates randomly derived from the same field at the same time. Early in three successive growing seasons (those ending in 1998, 1999, and 2000), field plots of the susceptible winter wheat cultivar Stephens were inoculated with suspensions of two M. graminicola isolates that each had rare alleles at restriction fragment length polymorphism (RFLP) loci. Near harvest time, leaves were randomly sampled from the same plots, and a population of over 100 monopycnidial isolates was created for each year of the experiment. Natural populations were also sampled from noninoculated plots in the 1999 and 2000 seasons, in order to compare allele frequencies. Based on RFLP haplotypes and DNA fingerprints, isolates from the inoculated plots were categorized by both inspection and Bayesian methods as inoculant clones, recombinants, or immigrants. Inoculation in the 2000 season was delayed, and the recovery rate of inoculant types was just 1 to 2%. In 1998, a high-disease year, and 1999, a low-disease year, inoculants comprised 36 and 22 to 23% of end-of-season samples, respectively. In those 2 years, recombinants as a percentage of inoculant descendants (both sexual and asexual) were 35 and 32%, respectively. By comparison, the study of fruiting bodies had found 93 and 32% of M. graminicola fruiting bodies were ascocarps in 1998 and 1999, respectively. These findings support the hypothesis that sexual recombination makes a relatively consistent contribution to M. graminicola population structure, despite differences in epidemic severity and ascocarp proportions.     

Identification and Molecular Mapping of a Gene in Wheat Conferring Resistance to Mycosphaerella graminicola

ABSTRACT Septoria tritici leaf blotch (STB), caused by the ascomycete Mycosphaerella graminicola (anamorph Septoria tritici), is an economically important disease of wheat. Breeding for resistance to STB is the most effective means to control this disease and can be facilitated through the use of molecular markers. However, molecular markers linked to most genes for resistance to STB are not yet available. This study was conducted to test for resistance in the parents of a standard wheat mapping population and to map any resistance genes identified. The population consisted of 130 F(10) recombinant-inbred lines (RILs) from a cross between the synthetic hexaploid wheat W7984 and cv. Opata 85. Genetic analysis indicated that a single major gene controls resistance to M. graminicola in this population. This putative resistance gene is now designated Stb8 and was mapped with respect to amplified fragment length polymorphism (AFLP) and microsatellite markers. An AFLP marker, EcoRI-ACG/MseI-CAG5, was linked in repulsion with the resistance gene at a distance of approximately 5.3 centimorgans (cM). Two flanking microsatellite markers, Xgwm146 and Xgwm577, were linked to the Stb8 gene on the long arm of wheat chromosome 7B at distances of 3.5 and 5.3 cM, respectively. The microsatellite markers identified in this study have potential for use in marker-assisted selection in breeding programs and for pyramiding of Stb8 with other genes for resistance to M. graminicola in wheat.     

A Gene-for-Gene Relationship Between Wheat and Mycosphaerella graminicola, the Septoria Tritici Blotch Pathogen

ABSTRACT Specific resistances to isolates of the ascomycete fungus Mycosphaerella graminicola, which causes Septoria tritici blotch of wheat, have been detected in many cultivars. Cvs. Flame and Hereward, which have specific resistance to the isolate IPO323, were crossed with the susceptible cv. Longbow. The results of tests on F1 and F2 progeny indicated that a single semidominant gene controls resistance to IPO323 in each of the resistant cultivars. This was confirmed in F3 families of Flame x Longbow, which were either homozygous resistant, homozygous susceptible, or segregating in tests with IPO323 but were uniformly susceptible to another isolate, IPO94269. None of 100 F2 progeny of Flame x Hereward were susceptible to IPO323, indicating that the resistance genes in these two cultivars are the same, closely linked, or allelic. The resistance gene in cv. Flame was mapped to the short arm of chromosome 3A using microsatellite markers and was named Stb6. Fifty-nine progeny of a cross between IPO323 and IPO94269 were used in complementary genetic analysis of the pathogen to test a gene-for-gene relationship between Stb6 and the avirulence gene in IPO323. Avirulence to cvs. Flame, Hereward, Shafir, Bezostaya 1, and Vivant and the breeding line NSL92-5719 cosegregated, and the ratio of virulent to avirulent was close to 1:1, suggesting that these wheat lines may all recognize the same avirulence gene and may all have Stb6. Together, these data provide the first demonstration that isolate-specific resistance of wheat to Septoria tritici blotch follows a gene-for-gene relationship.     

Effects of Wheat Cultivar Mixtures on Epidemic Progression of Septoria Tritici Blotch and Pathogenicity of Mycosphaerella graminicola

ABSTRACT The effects of host genotype mixtures on disease progression and pathogen evolution are not well understood in pathosystems that vary quantitatively for resistance and pathogenicity. We used four mixtures of moderately resistant and susceptible winter wheat cultivars naturally inoculated with Mycosphaerella graminicola to investigate impacts on disease progression in the field, and effects on pathogenicity as assayed by testing isolate populations sampled from the field on greenhousegrown seedlings. Over 3 years, there was a correspondence between the mixtures' disease response and the pathogenicity of isolates sampled from them. In 1998, with a severe epidemic, mixtures were 9.4% less diseased than were their component pure stands (P = 0.0045), and pathogen populations from mixtures caused 27% less disease (P = 0.085) in greenhouse assays than did populations from component pure stands. In 1999, the epidemic was mild, mixtures did not reduce disease severity (P = 0.39), and pathogen populations from mixtures and pure stands did not differ in pathogenicity (P = 0.42). In 2000, epidemic intensity was intermediate, mixture plots were 15.2% more diseased than the mean of component pure stands (P = 0.053), and populations from two of four mixtures were 152 and 156% more pathogenic than the mean of populations from component pure stands (P = 0.043 and 0.059, respectively). Mixture yields were on average 2.4 and 6.2% higher than mean component pure-stand yields in 1999 and 2000, respectively, but the differences were not statistically significant. The ability of mixtures challenged with M. graminicola to suppress disease appears to be inconsistent. In this system, host genotype mixtures evidently do not consistently confer either fitness benefits or liabilities on pathogen populations.     

Durability of Resistance to Globodera pallida I. Changes in Pathogenicity, Virulence, and Aggressiveness During Reproduction on Partially Resistant Potato Cultivars

ABSTRACT We studied changes in pathogenicity, virulence, and aggressiveness of Globodera pallida populations over time. As a measure for pathogenicity, the reproduction factor on a partially resistant host was used; for aggressiveness, the reproduction factor on a susceptible reference host was used; and, for virulence, the ratio pathogenicity/aggressiveness was used. The G. pallida populations were reared in a glasshouse for four generations on potato cultivars with different levels of resistance. The cultivar Elkana did not increase pathogenicity significantly, but the more resistant cultivars Karakter and Darwina did. This increase in pathogenicity was caused by an increase in virulence, whereas aggressiveness generally was not altered significantly. The increase in virulence appeared to be caused by an enhanced ability of eggs to develop into cysts, and not by an increase in egg production per new cyst. The observed changes in virulence could be predicted reasonably well by a simple numerical model. The rate of selection depended strongly on the nematode population. Rearing a mix of two different populations on a susceptible host decreased the virulence strongly, as predicted by the Hardy-Weinberg equilibrium, but increased the aggressiveness because of heterosis.     

Genetic structure of Mycosphaerella graminicola populations in Iran

To provide insight into the genetic structure of Mycosphaerella graminicola populations in Iran, a total of 221 isolates were collected from naturally infected wheat fields of five major wheat‐growing provinces and analysed using AFLP markers and mating‐type loci. All populations showed intermediate to high genotypic diversity. In the Golestan and Ardabil populations two mating types were found at near‐equal frequencies, whilst all populations were in gametic disequilibrium. Moreover, clonal haplotypes were identified in different sampling sites within a field in both the Khuzestan and Fars provinces, demonstrating that pycnidia are probably the primary source of inoculum. All five populations had low levels of gene diversity and had private bands. Low levels of gene flow and high genetic differentiation were observed among populations and different clustering methods revealed five genetically distinct groups in accordance with the sampling areas. The Golestan and East Azarbaijan populations were more genetically differentiated than the others. Random genetic drift, selection and geographic barriers may account for the differentiation of the populations. The results of this study indicate a population structure of M. graminicola in Iran contrasting to that of most other countries studied.     

Specificity of incomplete resistance to Mycosphaerella graminicola in wheat

We examined interactions between wheat (Triticum aestivum) and Mycosphaerella graminicola, causal agent of Septoria tritici blotch, to determine whether specific interactions occur between host and pathogen genotypes that could be involved in eroding resistance. The moderate resistance of the wheat cultivar Madsen has eroded significantly in the Willamette Valley of Oregon since its release in 1990. Foote is a replacement cultivar expressing moderate resistance and was released in 2000. Isolates of M. graminicola were collected from Foote and Madsen in 2004 and 2005 and tested on each cultivar in growth chamber and greenhouse experiments. There was a significant (P 相似文献   

The Occurrence of Mycosphaerella graminicola and its Anamorph Septoria tritici in Winter Wheat during the Growing Season

The disease septoria tritici blotch of wheat is initiated by ascospores of the teleomorph Mycosphaerella graminicola or pycnidiospores of the anamorph Septoria tritici. We report for the first time the presence of the teleomorph, M. graminicola, in Denmark. With the objective of elucidating the importance of the teleomorph for the development of septoria tritici blotch, data on the occurrence of fruit bodies of the anamorph (pycnidia) and the teleomorph (pseudothecia) stages were collected over three growing seasons. Pseudothecia were present in the springs, however, high numbers of pseudothecia compared to pycnidia were not observed until July, too late to influence the epidemic. On an individual leaf layer, pycnidia were observed well before pseudothecia. As the leaves aged, progressively higher proportions of fruit bodies were observed to be pseudothecia. The period from the appearance of pycnidia to detection of pseudothecia was estimated as 29–53 days. At harvest, high proportions of sporulating fruit bodies in the crop were pseudothecia, suggesting that the primary source of inoculum for new emerging wheat crops in autumn is likely to be ascospores.     

Population Structure of Mycosphaerella graminicola: From Lesions to Continents

ABSTRACT The genetic structure of field populations of Mycosphaerella graminicola was determined across a hierarchy of spatial scales using restriction fragment length polymorphism markers. The hierarchical gene diversity analysis included 1,098 isolates from seven field populations. Spatial scales ranged from millimeters to thousands of kilometers, including comparisons within and among lesions, within and among fields, and within and among regions and continents. At the smallest spatial scale, microtransect sampling was used to determine the spatial distribution of 15 genotypes found among 158 isolates sampled from five individual lesions. Each lesion had two to six different genotypes including both mating types in four of the five lesions, but in most cases a lesion was composed of one or two genotypes that occupied the majority of the lesion, with other rare genotypes interspersed among the common genotypes. The majority (77%) of gene diversity was distributed within plots ranging from approximately 1 to 9 m(2) in size. Genotype diversity (G / N) within fields for the Swiss, Texas, and Israeli fields was high, ranging from 79 to 100% of maximum possible values. Low population differentiation was indicated by the low G(ST) values among populations, suggesting a corresponding high degree of gene flow among these populations. At the largest spatial scale, populations from Switzerland, Israel, Oregon, and Texas were compared. Population differentiation among these populations was low (G(ST) = 0.05), and genetic identity between populations was high. A low but significant correlation between genetic and geographic distance among populations was found (r = -0.47, P = 0.012), suggesting that these populations probably have not reached an equilibrium between gene flow and genetic drift. Gene flow on a regional level can be reduced by implementing strategies, such as improved stubble management that minimize the production of ascospores. The possibility of high levels of gene flow on a regional level indicates a significant potential risk for the regional spread of mutant alleles that enable fungicide resistance or the breakdown of resistance genes.     

Incidence of Soil-borne Wheat Mosaic Virus in Mixtures of Susceptible and Resistant Wheat Cultivars

The multiplication of Soil-borne wheat mosaic virus (SBWMV) was studied in mixtures of two winter wheat (Triticum aestivum) cultivars, one susceptible (Soissons) and the other resistant (Trémie). Two seed mixtures of susceptible and resistant varieties in ratios of 1 : 1 and 1 : 3 and their component pure stands, i.e. each variety grown separately, were grown in a field infected with SBWMV. The presence of the virus was detected using DAS-ELISA from January to May. The resistant cultivar Trémie showed no foliar symptoms nor could the virus be detected in the leaves or roots. In May, about 88% of plants of susceptible cultivar Soissons grown in pure stands were infected. At this time, the disease reduction relative to pure stands was 32.2% in the 1 : 1 mixture and 39.8% in the 1 : 3 mixture. Optical density (OD) values from ELISA of the infected plants in the two mixtures were consistently lower than that of the infected plants in cultivar Soissons in pure stands. The ELISA index (EI) calculated using three scales of OD values was 65.5% in the susceptible cultivar in pure stands. The value for this index was 19.1% in the 1 : 1 mixture and 7.9% in the 1 : 3 mixture. The plants of the resistant cultivar Trémie infected in the same field and transferred in January to a growth cabinet at 15 °C multiplied the virus and produced viruliferous zoospores. These results show that the resistant cultivar Trémie plays a role in disease reduction in the cultivar mixtures in field conditions. Possible reasons for this are discussed.     

Evidence for Natural Selection in the Mitochondrial Genome of Mycosphaerella graminicola

ABSTRACT Pathogenicity assays were combined with restriction fragment length polymorphism (RFLP) markers in the mitochondrial and nuclear genomes to compare Mycosphaerella graminicola populations adapted to bread wheat (Triticum aestivum) and durum wheat (T. turgidum) in the Mediterranean Basin. The majority of isolates had unique nuclear DNA fingerprints and multilocus haplotypes. Only six mitochondrial DNA (mtDNA) haplotypes were identified among 108 isolates assayed. There were minor differences in frequencies of alleles at nuclear RFLP loci between the two host-adapted populations, but differences in the frequencies of mtDNA haplotypes were highly significant (P < 0.0001). mtDNA haplotype 1 dominated on the isolates adapted to bread wheat, and its frequency was twice as high as for the isolates adapted to durum wheat. mtDNA haplotype 4, which contained a unique approximately 3-kb insertion, was detected only in isolates showing specificity toward durum wheat and was the dominant haplotype on this species. We propose that the low mitochondrial diversity in this pathogenic fungus is due to a selective sweep and that differences in the frequencies of mtDNA haplotypes between the two host-adapted populations were due to natural selection according to host species.     

Alternative oxidase reduces the sensitivity of Mycosphaerella graminicola to QOI fungicides

Forty-six (1.5%) of nearly 3000 isolates of Mycosphaerella graminicola assayed in vitro were resistant to the QOI fungicide azoxystrobin, but on sub-culturing only ten remained resistant. Cross-resistance extended to other QOIs, but varied between different isolates. In planta the resistant isolates were not well controlled, especially at lower azoxystrobin dose rates. Propyl gallate, an inhibitor of alternative oxidase, potentiated the activity of azoxystrobin in vitro so that resistance was no longer observed. The growth of resistant strains in the presence of azoxystrobin led to alternative oxidase activation. This increased flexibility in respiration allows resistant strains to survive in the presence of a QOI fungicide. Under these conditions, selection for target-site mutations can occur. Using QOIs preventatively reduces the risk of resistance since the alternative oxidase cannot by itself generate all the energy needed for germination and early infection.     

Genetic Structure of Mycosphaerella graminicola Populations from Iran, Argentina and Australia

Restriction fragment length polymorphism (RFLP) markers were used to assess the genetic structure of populations of Mycosphaerella graminicola collected from wheat fields. A total of 585 isolates representing 10 field populations were sampled from Iran, Argentina and Australia. The genetic structure of M. graminicola populations from Iran and Argentina is described for the first time. Results were compared to previously investigated populations from Israel, Uruguay and Australia. Populations from Iran exhibited high clonality and low gene diversity, suggesting an inoculation event. Populations from uninoculated fields in Argentina had gene and genotype diversities similar to previously described European and North American populations. Genotype diversity was high for populations from Australia and tests for multilocus associations were consistent with sexual recombination in these populations. Gene diversity was low and fixed alleles were found for several loci. These findings are consistent with a relatively small founding population for Australia. These 10 new populations were integrated into a genetic distance comparison with 13 global populations that were characterized earlier.     

Update on mechanisms of azole resistance in Mycosphaerella graminicola and implications for future control

This review summarises recent investigations into the molecular mechanisms responsible for the decline in sensitivity to azole (imidazole and triazole) fungicides in European populations of the Septoria leaf blotch pathogen, Mycosphaerella graminicola. The complex recent evolution of the azole target sterol 14α‐demethylase (MgCYP51) enzyme in response to selection by the sequential introduction of progressively more effective azoles is described, and the contribution of individual MgCYP51 amino acid alterations and their combinations to azole resistance phenotypes and intrinsic enzyme activity is discussed. In addition, the recent identification of mechanisms independent of changes in MgCYP51 structure correlated with novel azole cross‐resistant phenotypes suggests that the further evolution of M. graminicola under continued selection by azole fungicides could involve multiple mechanisms. The prospects for azole fungicides in controlling European M. graminicola populations in the future are discussed in the context of these new findings. Copyright © 2012 Society of Chemical Industry