Levels of resistance of Erysiphe graminis f.sp. hordei to the systemic fungicide triadimenol

A random sample of conidiospores ofErysiphe graminis f.sp.hordei was obtained from the atmosphere above Cambridge, England, by incubating them on barley seedlings exposed on three dates in 1985. The asexual progeny of each spore was tested for its response to a range of doses of a systemic triazole fungicide, triademenol. Principal components analysis of the data revealed that the majority of isolates had a distinct level of response to triadimenol, being resistant to triadimenol at the rate apoplied to seed commercially. The remaining isolates appeared either to be sensitive to the lowest dose of triadimenol used, or to have one of two intermediate levels of resistance. There was a significant increase in the frequency of isolates with higher levels of resistance to triadimenol during 1985. This is likely to have been a response to the continuing widespread use of demethylation inhibitor (DMI) fungicides by British farmers.     

Genetic control of the resistance of Erysiphe graminis f.sp. hordei to five triazole fungicides

The genetics of resistance to ergosterol demethylation inhibitor (DMI) fungicides of the triazole (conazole) group was examined in a cross between two isolates of the barley powdery mildew fungus, Erysiphe graminis (=  Blumeria graminis ) f.sp. hordei . One isolate, E1, was previously identified as being resistant to the triazole fungicide triadimenol, while the other, HL3/5, was sensitive. The 56 progeny tested were classified into two distinct groups, either being resistant to triadimenol, like E1, or sensitive, like HL3/5. The segregation ratio was not significantly different from 1:1, consistent with responses to triadimenol being controlled by a single gene. In further tests with cyproconazole, epoxiconazole, propiconazole and tebuconazole, all the progeny classified as resistant to triadimenol were also more resistant to each of these other triazole fungicides than were any of the triadimenol-sensitive progeny. This is consistent with the triadimenol resistance allele also conferring cross-resistance to the other triazoles. The ratio between the responses of the resistant and sensitive progeny (the resistance factor, RF) was greatest for triadimenol, followed by tebuconazole, propiconazole, epoxiconazole and cyproconazole, in that order. The RF for triadimenol was much greater when the fungicide was applied as a seed treatment than when it was sprayed. Five isolates, covering the five levels of responses to triadimenol identified previously in the UK population of E. graminis f.sp. hordei , were used as standards; a triadimenol-sensitive isolate and one with the lowest level of resistance were sensitive to all four of the other fungicides, while three isolates with higher levels of triadimenol-resistance were also more resistant to the other chemicals.     

Sensitivity of Erysiphe graminis f. sp. hordei to ethirimol in relation to field performance in UK1

When introduced as a seed treatment 5 years ago, triadimenol (Baytan) was more effective against Erysiphe graminis f. sp. hordei than was the alternative ethirimol (Milstem). This contrasts with the current situation on the powdery mildew-susceptible cv. Golden Promise, where ethirimol is generally better than triadimenol. In part this reflects a decline in the performance of triadimenol seed treatment, as a result perhaps of some decrease in sensitivity to this fungicide. However, laboratory assays of field isolates also revealed that negatively correlated cross-resistance between triadimenol and ethirimol might exist. In field trials the sensitivity of E. graminis to triadimenol was indeed increased after treatment with ethirimol, but triadimenol hardly altered sensitivity to ethirimol. The possible causes of these changes in fungicide sensitivity are discussed.     

Genetics of avirulences in Erysiphe graminis f.sp. hordei

The genetics of avirulences towards barley mildew resistances were analysed in crosses of the Ervsiphe graminis f.sp. hordei isolate DH14 with CC107 and with CC138. Nine avirulences, Av r_a9, Avr _a10, Avr _a11, Avr _a12, Avr _Ab, Avr _CP, Avr _h, Avr _k and Avr _La, segregated as single genes in one or other cross. However. F₁ segregation data were consistent with avirulence matching the Mla7 resistance gene being controlled by two genes, designated Avr _a7 1 and Avr _a7 2. Infection types of avirulent isolates differed on varieties in which Mla7 had been derived from each of the four known sources of that resistance. Linkage was detected between Avr _a7 1 and Avr _h in the cross CC107 × DH14, and between Avr _a10 and Avr _k, Avr _a11 and Avr _La, and Avr _h and the triadimenol response gene Tdl2 in CC138 × DH14.     

Structure and evolution of a population of Erysiphe graminis f.sp. hordei

Samples of single colony isolates of Erysiphe graminis f.sp. hordei were collected in Cambridge on three dates in 1985, and tested for the presence of 12 virulence alleles and resistance to two fungicides, triadimenol and ethirimol. The frequency of the virulence V-(La) fell during 1985, while the frequency of V-h. virulence on cv. Triumph and higher levels of resistance to each fungicide and combined resistance to both fungicides rose. Two phenotypes, both of which possessed virulence on cv. Triumph and three unnecessary virulence alleles and had similar sensitivities to ethirimol, but differed in their level of resistance to triadimenol, accounted for 35.9% of all isolates. The high frequency of these phenotypes accounted for most of the observed gametic phase disequilibria between pathogenicity characters. Most individuals with these two phenotypes may be members of the same clone. It was estimated that 25% of the spore population which initiated the autumn epidemic of E.g. f.sp. hordei originated from ascospores formed by sexual reproduction in the summer. It is proposed that genetic drift followed by hitch-hiking selection, due to intense selection for a clone virulent on a newly-introduced cultivar, is a major factor influencing the frequency of unnecessary virulence alleles.     

Comparison of resistance-inducing abilities of virulent and avirulent races of Erysiphe graminis f.sp. hordei and a race of Erysiphe graminis f.sp. tritici in barley

Resistance to powdery mildew was induced in barley by preinoculation with virulent and avirulent races of barley powdery mildew ( Erysiphe graminis f.sp. hordei ), and with a race of wheat powdery mildew ( E. graminis f.sp. tritici ). Four inducer densities were tested in 13 different induction periods between 1 and 24 h. Generally, the resistance induced by barley powdery mildew increased up to 10-12 h of induction and was maintained in longer induction periods. The inducing abilities of virulent and avirulent races could not be distinguished up to 10-12 h of induction, after which the inducing ability of avirulent races increased significantly in relation to virulent races. Wheat powdery mildew was able to induce more resistance than barley powdery mildew in induction periods up to 8 h. In a single inoculation procedure the number of haustoria developing from virulent barley powdery mildew decreased as inoculum density increased. The effect was ascribed to induction of resistance. This reduction of infection efficiency in the compatible interaction was compared to induced resistance. However, the inoculum density needed for 50% resistance induction in the double inoculation procedure was approximately 40 times higher than the inoculum density needed for 50% reduction in infection efficiency in the single inoculation procedure.     

The spread oi Erysiphe graminis f. sp. hordei in mixtures of barley varieties

A model is proposed of mechanisms which might affect the progress of Erysiphe graminis f. sp. hordei in mixtures of barley varieties. Results obtained from two field trials indicate that the efffect of mixtures may be panitioned into three categoriesof the influence of the reduced density of the susceptible plants, the barrier effect of the resistant plants, and the induced resistance due to the non-virulent pathogen biotypes. In the early stages of plant growth the lower density of susceptible plants accounted for most of the reduction in pathogen development in mixtures. As the epidemic progressed, the barrier and induced resistance effects increased in importance and the total mixture effect was at a maximum mid-way through epidemic development. Towards the end of the trials the overall mixture effect declined though the influence of induced resistance was at its maximum. The reasons for these changes and their implications for the use of host varietal mixtures in disease control are discussed.
Mixtures also protected the crop against a pathogen other than the target organism.     

Virulence patterns in populations of Erysiphe graminis f.sp. hordei in Europe in 1986

A total of 3166 single colony progeny of Erysiphe graminis f.sp. hordei , constituting random samples of the aerial inoculum from 68 regions of Western Europe, were analysed in the laboratory for virulence to barley resistance genes Mlra, Mlg, Mla6+Mlg. MILa, Mla7+Mlk, Mlall, Mla9, MlaI3 + MlRu3. Mlal, Mla3 and mlo. No isolate virulent to mlo was detected. Distinct regional patterns were observed for the frequencies of most of the other virulence genes. They stressed the role of selection exerted by the cultivars grown, as well as the effect of wind dissemination of mildew populations. These two phenomena are particularly well exemplified by the data on early stages in the evolution of the newly emerging Val3 virulence in Denmark. In contrast with the preceding evolution, there was a considerable decrease of the frequency of a broadly distributed virulence ( Va6); the most probable reasons for this are outlined.     

The use of DNA polymorphisms to test hypotheses about a population of Erysiphe graminis f.sp. hordei

The distribution of restriction-fragment-length polymorphisms (RFLPs) among isolates of Erysiphe graminis f.sp. hordei was used to test hypotheses about the structure of the pathogen population. There was a large diversity in the lengths of DNA restriction fragments homologous to E9, a chromosomal fragment of unknown function. It was shown that a large group of isolates, which shared identical or very similar virulence phenotypes, including virulence on the barley cv. Triumph, were members of a single clone. Several other clones were identified, including a second group of identical isolates which were virulent on cv. Triumph but were highly distinct from the more common clone of Triumph-virulent isolates. Isolates which shared the same level of resistance to the triazole fungicide triadimenol were genetically diverse. Such diversity is consistent with triadimenol resistance being under oligogenic, rather than polygenic, control. Conclusions obtained from analysis of the most easily identifiable subset of fragment lengths were very similar to those obtained from analysis of the complete set.     

Relative tolerances of wild and cultivated barleys to infection by Blumeria graminis f.sp. hordei (Syn. Erysiphe graminis f.sp. hordei). I. The effects of infection on growth and development

The two lines of wild barley, B19909 and I-17-40 and the cultivated barley, cv. Prisma used in this investigation were found to be the most susceptible to infection of 25 wild and four cultivated barley lines when exposed to the local population of Blumeria graminis f.sp. hordei. All three lines were susceptible during the early stages of growth but expressed some level of adult plant resistance although this level of resistance was significantly higher in line I-17-40 than in either of the other two.The relative tolerances of the lines to the mildew were determined by comparing the effects of infection on their growth and development in growth cabinet experiments. Mildew developed more slowly on line I-17-40 than on the other two lines and by the third week after inoculation, when mildew cover on B19909 and cv. Prisma had reached about 27%, only about 15% of the green leaf area of line I-17-40 was covered. Mildew continued to increase on line B19909 and cv. Prisma so that 6 weeks after inoculation it covered 40% of their leaf-blades. On line I-17-40 30% of the green leaf area was colonised by 4 weeks after inoculation but because of adult plant resistance coupled with the loss of the earlier infected leaves through senescence mildew cover then reduced falling to 15% by 6 weeks. Although total mildew biomass, measured as conidial production was higher on line B19909 than on cv. Prisma all its growth parameters were reduced less indicating that it was the more tolerant line. Conidial production on the lower susceptible leaves of line I-17-40 was slightly lower than on cv. Prisma yet the reaction of these leaves to infection was the same on both lines indicating that tissues of I-17-40 were slightly less tolerant than those of the cultivated barley. However, during the later stages of growth when its upper leaves expressed high levels of ‘adult plant resistance’ dry matter production in this line increased to levels higher even than in the controls. This capacity for compensatory photosynthesis ensured that by the end of growth few differences in any of the measured growth parameters between infected and uninfected plants of line I-17-40 were significant.The greater tolerance of line B19909 over the other two lines and of cv. Prisma over line I-17-40 during the early stages of growth appears to be due to a lower sensitivity to infection of those processes which regulate dry matter accumulation and its distribution around the plant.     

The influence of the primary germ tube on infection of barley by Erysiphe graminis f. sp. hordei

In each of three separate genotype combinations of Erysiphe graminis f. sp. hordei and barley the probability of infection success by an appressorium was reduced by close proximity to a primary germ tube interacting with the same host cell, whether this germ tube was produced by the same or another conidium. Infection success of appressoria was unaffected by events in an adjacent cell, whether associated with other conidia or with the primary germ tube of the same conidium. This interaction between germ tubes and underlying host cells should be considered when mechanisms of resistance to E. graminis f. sp. hordei are investigated.     

Selection of pathotypes of Erysiphe graminis f.sp. hordei in pure and mixed stands of spring barley

Pathotypes of Erysiphe graminis f.sp. hordei were monitored at fortnightly intervals in pure and mixed stands of spring barley during the course of mildew epidemics in two field trials. Mixtures were composed of cultivars with Arabische (gene Mla12 ), Laevigatum ( Ml(La) ), and Monte Cristo ( Mla9 ) resistance, respectively. The three-way mixtures were either random or, in 1989, laid out as one-row mixtures (i.e., regularly alternating rows of different genotypes) or three-row mixtures (i.e., regularly alternating three-row strips of different genotypes), respectively. In 1990 only random mixtures and six-row mixtures were compared with pure stands. The virulence complexity (i.e., the average number of virulence factors per isolate with reference to Mla12, Ml(La) , and Mla9 ) was always maximal in the random mixtures. In 1989, linear regression of complexity on mildew generations gave significant b -values (slopes) of 0·049, 0·031, and 0·025 in the random mixture, one-row mixture, and three-row mixture, respectively; the b -value from pure stands was not significant. In 1990, another sampling technique allowed selection to be observed on each genotype in the mixtures separately. In the random mixture b -values were 0·048, 0·064 and 0·017 (not significant) on Mla12, Ml(La) , and Mla9 cultivars respectively. In six-row mixtures and in pure stands, there was no significant increase in complexity ( b > 0) on any of the mixture components. Although the frequency and relative fitness of complex pathotypes were higher in all types of mixtures than in pure stands, selection towards complex races was much less intense in row mixtures than in random mixtures in both field trials.     

Genetic variation in Danish populations of Erysiphe graminis f.sp. hordei: estimation of gene diversity and effective population size using RFLP data

Genetic variation of the barley powdery mildew fungus ( Erysiphe graminis f.sp. hordei ) was estimated in three Danish local populations. Genetic variation was estimated from the variation amongst clones of Egh , and was therefore an estimate of the maximum genetic variation in the local populations. The average gene diversity, Ĥ , was estimated as 0.84. The effective population size was estimated as: log₁₀ ( N^ _e ) = 0.64 − log₁₀(μ), or 4.4 × 10⁹, assuming a nucleotide mutation rate ( μ) of 10⁻⁹ per base per generation. There was no significant genetic differentiation between locations.     

The effect of fungal density on fungicide dose–response curves in barley powdery mildew (Erysiphe graminis f.sp. hordei)

The effect of inoculation density on fungicide dose–response curves and the estimated ED₅₀ value (the fungicide concentration needed to halve the infected leaf area relative to the infected area of leaves that had not been treated with a fungicide) were investigated theoretically, and predictions were tested experimentally using powdery mildew of barley ( Erysiphe graminis [syn. Blumeria graminis ] f.sp. hordei ). A host leaf was assumed to consist of a number of independent compartments, in each of which only a single fungal spore could germinate successfully. The number of fungal spores landing in a compartment was assumed to be Poisson-distributed. The spores were assumed to vary in their sensitivity toward a fungicide, and the sensitivity of the spore population was assumed to be normally distributed around a mean sensitivity. Under these assumptions, the ED₅₀ value was shown to be a positively biased estimate of the mean sensitivity in the fungal population, and the bias increased with inoculum density. Consequently, the estimated ED₅₀ value is expected to vary between experiments and laboratories if the inoculation density varies over a considerable range. The correlation between inoculum density and estimated ED₅₀ value is expected to be strongest when the assayed fungal population consists of several genotypes differing in sensitivity. This expectation was tested by estimating the ED₅₀ values for one barley powdery mildew isolate at different inoculation densities for three different fungicides. These ED₅₀ values were positively correlated strongly with the inoculum density for fenpropimorph and more weakly for triadimenol, whereas no correlation was observed for propiconazole.     

Utility of defeated resistance genes to powdery mildew,Erysiphe graminis f. sp. hordei,in spring barley variety mixtures

In 1980 and 1981 experiments have been performed to study the utility of defeated resistance genes to powdery mildew in spring barley variety mixtures. For this purpose the epidemic development of powdery mildew in pure stands and mixtures of four spring barley varieties was monitored. In three varieties the resistance was overcome several years ago. One variety is still resistant. Changes in frequencies of corresponding virulence genes were also studied.It was found that the variety mixtures slowed down the mildew epidemic only at the beginning of the season. Some reduction was lost later on since the pure stands reached their saturation level of infection at an earlier date than the more healthy mixtures. The level of reduction of infection depended on the composition of the mixtures and the number of components. Two-way mixtures appeared to be very risky due to the high frequency of corresponding combinations of virulence genes in the pathogen. Furthermore, two-way mixtures appeared to be unable to reduce the infection rate sufficiently when the infection pressure was high.In 1981 the frequencies of combinations of corresponding virulence genes increased very much in the Diva-Mazurka mixture but declined in the pure stands of both varieties.On the basis of these results it is concluded that the utility of defeated resistance genes involved in this study is very limited.The low number and the irregular distribution of the Dutch spring barley varieties over the different resistance groups prohibit the use of variety mixtures in the Netherlands.Samenvatting In 1980 en 1981 is onderzoek verricht over de bruikbaarheid van doorbroken resistentiegenen tegen meeldauw,Erysiphe graminis f. sp.hordei, in zomergerstrassenmengsels. Daartoe werd de epidemieontwikkeling in monocultures en mengsels van vier zomergerstrassen bestudeerd. Van drie rassen is de resistentie reeds verscheidene jaren doorbroken. Eén ras is nog resistent. De frequenties van compatibele meeldauwvirulentiegenen werden eveneens bepaald.Het bleek dat in vergelijking met de monocultures in de rassenmengsels de meeldauwepidemie alleen in het begin van het seizoen werd afgeremd. Later in het seizoen ging weer enige reductie in aantasting van de mengsels verloren doordat in de monocultures de aantasting eerder het verzadigingspunt bereikte dan in de minder aangetaste mengsels. De mate waarin de aantasting gereduceerd werd was afhankelijk van de aard en het aantal componenten in het mengsel. Twee-componentenmengsels bleken vrij riskant te zijn vanwege de aanwezigheid van combinaties van corresponderende virulentiegenen in de meeldauwpopulatie. Verder bleken tweecomponentenmengsels niet in staat de infectiesnelheid afdoende af te remmen wanneer de infectiedruk hoog werd.De frequentie van combinaties van corresponderende virulentiegenen nam sterk toe in het Diva-Mazurka-mengsel doch nam af in de monocultures van deze beide rassen.Gezien de verkregen resultaten lijkt de bruikbaarheid van de in dit onderzoek betrokken doorbroken resistentiegenen erg beperkt. Het geringe aantal zomergerstrassen op de rassenlijst en de ongelijke verdeling over de te onderscheiden resistentiegroepen belemmeren de toepassing van rassenmengsels in de Nederlandse akkerbouw.     

Racial diversity and complexity in regional populations of Erysiphe graminis f.sp. hordei in France over a 5-year period

The evolution of race patterns in three French regional populations of the barley powdery mildew pathogen Erysiphe graminis f.sp. hordei over a 5-year period showed rapid adaptation to newly introduced host resistance genes. In all three regions, the main change consisted of the replacement of initially abundant races by pathotypes differing markedly from them by their virulence gene combinations. This explained the increase in diversity during the first 3 years of the survey, when the second group of pathotypes became more common in the populations, and its subsequent decrease due to the decline of the first group of races. The mean number of virulence genes per isolate did not vary noticeably over time in the three populations, remaining at about four out of 12 genes tested. However, the distribution of the isolates into virulence complexity classes was greatly modified, fitting a binomial distribution by the end of the study, although significant deviations were apparent in the first 2 years (1986 and 1987). The data indicate that selection, migration and recombination are the most important factors shaping race structure and evolution in powdery mildew populations, and that mutation is of limited significance. No convincing evidence was obtained for the existence of stabilizing selection sensu Vanderplank as the mechanism limiting virulence complexity. Implications regarding spatial and temporal deployment of race-specific resistance genes to control powdery mildew are discussed.     

Histology of infection by Erysiphe graminis f.sp. hordei in spring barley lines with various levels of partial resistance

The histology of resistance to infection by conidia of Erysiphe graminis f.sp. hordei has been studied in barley lines with various levels of partial resistance. Resistance reduced the proportion of conidia which formed haustoria and was expressed more strongly in fifth than in first formed leaves. Where attempted penetration from the first appressorial lobe failed, a second and sometimes a third lobe could be formed. Most penetrations were from first lobes, a few from second lobes and a negligible number from third lobes. Adult plant resistance decreased penetration success from both first and second lobes. An unidentified component of resistance appeared to impede the development of less vigorous conidia before they stimulated a host cell response. However, in all hosts the majority of failed penetration attempts stimulated a localized host response implicating this response in resistance. Some attempts caused hypersensitive death of epidermal cells in both resistant and susceptible hosts; the proportion of dead cells was higher in fifth than first leaves. Stomatal subsidiary cells were more susceptible to penetration than interstomatal cells and in resistant leaves the majority of successful penetrations were into subsidiary cells. The pathogen was also able to avoid resistance to penetration by establishing endophytic infection following entry of germ tubes through stomates.     

Virulence phenotypes of Blumeria graminis f. sp. hordei in South Africa

Virulence analysis of 224 isolates of Blumeria graminis f. sp. hordei (barley powdery mildew) from South Africa was performed. The isolates were collected from eight fields and a greenhouse in 2004 and 2007. The isolates were tested for virulence on a set of 20 differential varieties. All isolates were virulent on the resistance genes Mla8 and Ml(Ch) and avirulent for the resistance genes Mla3, Mla6, Mla7, Mla9, Mla13, Mla23, Mlp1 and MlaN81. Virulence frequencies of field isolates for the resistance genes Mla12 + MlaEm2, Mlat, Mla22, Mlk1 and Mlh were 52.9–99.5 % and for Mla1 + MlaAl2, MlLa, Mlra, Mlg + MlCP and Ml(Ru2) were 0.5–23.5 %. In total, 46 pathotypes were detected in the field and seven other pathotypes in the greenhouse. Only nine pathotypes were found in both years, but they included 61.8 % of the isolates. The predominant pathotype represented 15.9 % of the isolates, and was the only one common to all three field populations. The average relative virulence complexity per field isolate increased from 0.405 in 2004 to 0.486 in 2007. Two powdery mildew metapopulations in geographically distant and separated areas (North West and Western Cape) were deduced. The South African population of Blumeria graminis f. sp. hordei had unique virulence frequencies and virulence associations when compared to populations from other parts of the world.