Isolation of R-type progeny of Tapesia yallundae from apothecia on wheat stubble in England

Apothecia of Tapesia yallundae were collected from a set-aside straw stubble site in Lincolnshire in March 1993. Single ascospore isolates were obtained which produced colonies with morphologies and growth rates characteristic of the R-pathotype of Pseudocercosporella herpotrichoides. Isolates were confirmed to be R-type by the use of randomly amplified polymorphic DNA (RAPD) markers. These observations confirm that T. yallundae is the teleomorph of the R-type of P. herpotrichoides and represent the first detection of the sexual stage of the R-type in the UK.     

Observations of apothecia of Tapesia yallundae and the cultural phenotypes of their progeny

Apothecia of Tapesia yallundae, the perfect state of Pseudocercosporella herpotrichoides, were found in March 1989 on wheat and barley stubble at nine of 37 naturally infested sites and also at one inoculated site in Germany. Apothecia were found on about 1% of stems collected from the naturally infested sites and 5% of stems from the inoculated site, and were first found in the second week of March, coinciding with the onset of an early spring. Apothecia were not found on either green plants or second-year stubble. In total, 143 single-spore isolates were obtained from the apothecia and described according to a range of morphological characteristics. The majority were ascribed to P. herpotrichoides var. herpotrichoides. This is the first isolation of this variety from T. yallundae apothecia in Germany. Even single-conidial isolates from one apothecium showed much variation, some being like var. herpotrichoides, some like var. acuformis and some intermediate.     

Development of apothecia ofTapesia yallundae in contrasting populations selected by fungicides

Apothecia of the eyespot fungus,Tapesia yallundae, were found on 0–18% of straws in plots of wheat stubble in February–March 1994. The fungicides carbendazim, prochloraz or carbendazim plus prochloraz had been applied repeatedly to the same plots in each of the previous 9 years in which successive wheat crops had been grown. The factors most strongly correlated with the incidence of apothecia were the incidence and severity of eyespot in the preceding wheat crop and the frequency of carbendazim-resistant W-type fungus in populations recovered from that wheat crop. Plots treated with carbendazim, which had previously had more disease and more resistance to carbendazim in the pathogen population relative to untreated plots, therefore yielded most apothecia. Plots treated with prochloraz, which had selected for predominantly R-type fungus and decreased eyespot, yielded few apothecia. Single-ascospore isolates were all of the W-type and were more frequently carbendazim-sensitive than expected, except those from plots treated only with carbendazim. None showed decreased sensitivity to prochloraz. The implications of applying fungicides regularly for controlling eyespot on the capability of the eyespot fungus for genetic variation through sexual reproduction are discussed.     

Occurrence of Tapesia yallundae apothecia on field and laboratory-inoculated material and evidence for recombination between isolates

Tapesia yallundae was observed in early July on wheat straws, which had been inoculated with isolates of Pseudocercosporella herpotrichoides (W-type) 9 months previously and placed in plots of winter wheat, to act as inoculum in a resistance screen. A perfect state was not observed on straws inoculated with R-type isolates in the same experiment. One hundred and thirty-four single-ascospore isolates were obtained and observations on morphology in culture, pathogenicity to wheat and rye seedlings, resistance to benomyl and the pattern of restriction fragment length polymorphisms demonstrated that all isolates were W-type, although some grew at half the expected rate on agar, and at least two isolates were involved in the production of apothecia. Apothecia were produced in vitro on straws inoculated with single isolates of P. herpotrichoides only when they were incubated in contact with other straws similarly inoculated with certain other isolates. No apothecia were produced on straws with only isolate present, suggesting the need for at least two strains, possibly representing different mating types, for apothecium formation. Apothecia of T. yallundae were produced in vitro on straw pieces which had been inoculated individually with W-type isolates, 14-16 weeks after straws of different isolates were mixed. Evidence for genetic recombination between isolates was obtained when single-ascopore colonies were analysed for resistance to benomyl in culture and for restriction-length polymorphisms (RFLPs).     

Development of apothecia of the eyespot pathogen Tapesia on cereal crop stubble residue in England

A reservoir of infection of Tapesia yallundae may exist after harvest in bases of cereal stems due to the presence of apothecia capable of discharging infective ascospores. Apothecia of T. yallundae developed in a seasonal pattern on winter barley inoculated with the pathogen, with maximum numbers of apothecia produced on stubble 5–7 months after harvest. A similar pattern of development was observed on infected winter wheat. However, the peak in numbers of mature apothecia was observed 2 months later than in winter barley. Apothecia capable of discharging ascospores were present for up to 6 months on stubble. Apothecia of T. acuformis were not detected on spring or winter barley, or spring or winter wheat stubble, despite inoculation of growing crops with isolates of compatible mating type.     

Evolution of fungicide resistance in the cereal eyespot fungi Tapesia yallundae and Tapesia acuformis in France

Field isolates of the cereal eyespot pathogen can be divided into two groups which are now considered as two species: Tapesia yallundae and Tapesia acuformis. In both species the first case of acquired resistance was observed with benzimidazole fungicides in the early 1980s. At the same time, a number of sterol C-14 demethylation inhibitors (DMIs), such as the imidazole prochloraz and several triazoles, including flusilazole, were introduced. Surprisingly T. acuformis appeared intrinsically resistant to the triazole derivatives in comparison to T. yallundae, but both species were sensitive to prochloraz. The intensive use of these DMIs led to the development of acquired resistance towards triazoles in T. yallundae and towards prochloraz in T. acuformis. Today all the strains in both species appear equally sensitive to the anilinopyrimidine cyprodinil. ©1997 SCI     

Occurrence of Tapesia yallundae, teleomorph of Pseudocercosporella herpotrichoides, on unharvested wheat culms in England

Apothecia of Tapesia yallundae are reported for the first time in the UK. They were observed on decaying stems and leaf sheaths of unharvested wheat near Bristol in January 1989. Isolates from single ascospores produced cultures typical of the W-type form of the anamorph, Pseudocercosporella herpotrichoides . Pathogenicity on wheat was confirmed by inoculation and re-isolation from glasshouse-grown wheat seedlings.     

Differentiation and quantification of the cereal eyespot fungi Tapesia yallundae and Tapesia acuformis using a PCR assay

Isolates of Tapesia yallundae and Tapesia acuformis were subjected to Random Amplified Polymorphic DNA (RAPD) assay. Amplification products common to isolates of either species were cloned and primers were generated from each sequence for use in conventional PCR. The primer pair derived from a T. yallundae specific RAPD marker amplified a product only from DNA of T. yallundae isolates and not from DNA of a range of other fungal species associated with the stem base disease complex of cereals. Similarly, the primer pair generated from a T. acuformis -specific RAPD marker amplifed a product only from DNA of T. acuformis isolates. Quantitative assays were developed for both species of Tapesia from these primer pairs, using competitive PCR . Competitive PCR was used to determine the level of colonization of seedlings by each species in glasshouse- and field-inoculated cereal hosts and results compared to those for conventional seedling disease assessment.     

Translocation of benomyl,prochloraz and procymidone in relation to control of Botrytis cinerea in strawberries

Benomyl, prochloraz or procymidone, applied as an overall plant spray at the openflower stage, effectively suppressed Botrytis cinerea fruit rot, whereas no control was achieved by foliar application only. Fruit rot was prevented using procymidone applied to the soil 12 days before inoculation of the flowers, whereas benomyl or prochloraz gave little or no control, respectively when applied in the same manner. Bioassays, using Penicillium expansum on leaf and flower extracts of strawberry plants growing in soil treated with procymidone, showed the presence of an inhibitory compound with the same R_F value on thin-layer chromatography as that of procymidone. Analysis by gas chromatography and identification by gas chromatography—mass spectrometry established that the fungicide procymidone was translocated from the root system of strawberry plants to the leaves and flowers.     

A New Source of Resistance to Tapesia yallundae Associated with a Homoeologous Group 4 Chromosome in Thinopyrum ponticum

ABSTRACT Wheat (Thinopyrum ponticum line SS767; PI 611939) with 42 chromosomes previously was identified as a new source of eyespot resistance. Individual plants of SS767 were tested for reaction to Tapesia yallundae, the major pathogen of eyespot in the Pacific Northwest region of the United States. Resistance of this line was similar to the resistant winter wheat cv. Madsen (carrying gene Pch1 for eyespot resistance). Polymerase chain reaction analysis with primers specific for the J or E genomes revealed that SS767 contains Thinopyrum chromatin. Cytological and Cbanding analyses demonstrated that SS767 is a chromosome substitution line in which wheat chromosome 4D is replaced by a homoeologous group 4 chromosome of Thinopyrum ponticum. Genomic in situ hybridization using St genomic DNA from Pseudoroegneria strigosa as a probe, which can differentiate chromosomes from different genomes of Thinopyrum, indicated that this chromosome belongs to the J genome. Molecular analysis of an F(2) population segregating for chromosome 4J and resistance to eyespot confirmed that eyespot resistance in line SS767 is associated with chromosome 4J of Thinopyrum ponticum. This is the first report of genetic control of resistance to eyespot derived from Thinopyrum ponticum. This source of resistance provides a new opportunity to improve wheat resistance to eyespot by adding to the diversity of resistance sources available.     

Incidence of benomyl and iprodione resistance in isolates of Botrytis cinerea in tomato crops in England and Wales in 1986

In 1986, mass-mycelial isolates of Botrytis cinerea from 67 tomato crops in England and Wales were examined for benomyl and iprodione resistance. Of the 706 isolates obtained, 62.7% were resistant to benomyl at 2μg/ml and 43.2% were resistant to iprodione at 2 μg/ml. Iprodione resistance persisted in the absence of a dicarboximide spray programme. The incidence of benomyl resistance has not decreased since the last survey in 1984 in spite of a considerable reduction in the use of benzimidazole fungicides. There were no clear indications that the use of dichlofluanid influenced the incidence of benomyl or iprodione resistance. Disease control was poorer in crops with a higher incidence of iprodione resistance.     

Species and Mating-Type Distribution of Tapesia yallundae and T. acuformis and Occurrence of Apothecia in the U.S. Pacific Northwest

ABSTRACT Eyespot of wheat is caused by the discomycete fungi Tapesia yallundae and T. acuformis. T. yallundae is considered the most important causal agent of the disease in this region but no apothecia of either species have been found in the U.S. Pacific Northwest (PNW). Two compatible isolates of T. yallundae from the PNW were used to inoculate a field plot in the fall of 1998 and apothecia developed in the spring and fall of 2000 on standing wheat stubble. In the spring of 2000, wheat stubble from eight naturally infected fields was examined for the presence of apothecia of T. yallundae and T. acuformis. Apothecia of T. acuformis were found in two fields but no apothecia of T. yallundae were found. This is the first report of apothecia of the eyespot pathogens occurring in the PNW. Species and mating-type distribution of T. yallundae and T. acuformis in the PNW were determined from 817 isolates collected from diseased wheat over 3 years at spatial scales ranging from within fields to across states. In all, 460 isolates were identified as T. yallundae and 357 isolates were identified as T. acuformis with MAT1-1/MAT1-2 ratios not significantly different from 1:1 based on chi(2) tests at most scales tested. The apparent increase in frequency of T. acuformis from previous surveys may indicate a shift in the predominant species causing eyespot. The occurrence of apothecia under field conditions, along with the widespread distribution of mating types of both species, suggests that sexual reproduction may be occurring in both species.     

Acquired resistance to benomyl and some other systemic fungicides in a strain of Botrytis cinerea in cyclamen

A benomyl-resistant strain (R) ofBotrytis cinerea was isolated from cyclamen that had been sprayed with relatively high doses of Benlate two weeks before. In vitro mycelial growth of this strain was less inhibited on PDA containing 1000 g/ml benomyl (Benlate, 50% W.P.) than that of another, wild isolate ofB. cinerea from cyclamen on PDA with 0.5 g/ml of the fungicide.The R-strain was also resistant to methyl-thiophanate, furidazol and to a lesser extent to thiabendazole. Mycelial growth of 5 other isolates was much more inhibited by benomyl than by thiabendazole.Resistance was retained for at least 20 weeks after repeated subculturing on fungicide-free agar.Samenvatting In een kwekerij, waar bespuiting met benomyl (Benlate, 50% W.P.) drie maal was toegepast ter bestrijding vanBotrytisrot in cyclamen, bleek de laatste bespuiting niet meer effectief. Integendeel, de ziekte breidde zich sneller uit dan onder normale omstandigheden het geval is. Uit bloemstelen van de aangetaste planten werd eenB. cinerea-stam (R) geïsoleerd, die zeer resistent bleek tegen benomyl. In vitro werd de groei van deze stam op aardappel-glucose-agar met 1000 g/ml benomyl (Benlate 50% W.P.) minder geremd dan die van een willekeurigB. cinerea-isolaat van cyclamen op het medium met 0.5 g/ml van het fungicide (Tabel 1, Fig. 1).De R-stam bleek eveneens resistent tegen methyl-thiophanaat, furidazol en in mindere mate tegen thiabendazol (Tabel 2).De myceliumgroei van vijf isolaten vanB. cinerea, verkregen van verschillende waardplanten, bleek in tegenstelling tot die van de R-stam juist sterker geremd te worden door benomyl dan door thiabendazol (Tabel 3).De R-stam bleef gedurende tenminste 20 weken resistent na regelmatig overenten op voedingsbodems zonder het fungicide.     

First record of Tapesia yallundae as the teleomorph of Pseudocercosporella herpotrichoides var. acuformis, and its occurrence in the field in the Federal Republic of Germany

Tapesia yallundae is reported for the first time in the Federal Republic of Germany. Apothecia were found on wheat and rye stubble at two sites in the north of the country and one in the centre, in April 1988. Isolates of Pseudocercosporella herpotrichoides var. acuformis were obtained from germinating single ascospores, identified on the basis of colony morphology, mode of conidial production and pathogenicity to wheat and rye.     

西藏辣椒疫病菌及其对新型杀菌剂的敏感性测定

经分离鉴定,引起西藏辣椒疫病的病原菌为辣椒疫霉,菌丝生长温度为10~37℃,最适生长温度25~30℃,致死温度40℃。交配型为A2自育型。烯酰吗啉和腈嘧菌脂均对辣椒疫霉的菌丝生长具有很强的抑制作用,分别抑制菌丝的生长速度和菌丝的生长量,EC50分别为0.241 mg/L和0.302 8 mg/L,比常规杀菌剂甲霜灵EC50分别低32.6和25.9倍。     

禾谷镰孢菌对苯醚甲环唑的敏感性基线及与其他杀菌剂敏感性的相关性分析

由禾谷镰孢菌引起的赤霉病是小麦上的重要病害,可严重影响小麦的产量并降低小麦的品质。苯醚甲环唑属于三唑类杀菌剂,是甾醇脱甲基化抑制剂,具有较高的抑菌活性。采用菌丝生长速率法测定了于2016—2017采集自河南省的107株禾谷镰孢菌对苯醚甲环唑的敏感性。结果表明:苯醚甲环唑对禾谷镰孢菌菌丝的生长具有较强的抑制效果,其有效抑制中浓度(EC₅₀)值范围为0.0128~0.6079 mg/L,符合正态分布,平均EC₅₀值为(0.2239 ± 0.1192) mg/L。因此,这些敏感性数据可以作为河南省禾谷镰孢菌对苯醚甲环唑的敏感性基线。通过对苯醚甲环唑与其他7种杀菌剂氟环唑、多菌灵、氰烯菌酯、氟唑菌酰羟胺、戊唑醇、丙硫菌唑及叶菌唑对20株禾谷镰孢菌的log₁₀ EC₅₀值之间的Spearman’s rho (ρ) 相关性分析发现:苯醚甲环唑与叶菌唑之间具有较低水平的相关性,与其他供试杀菌剂之间无相关性。本研究可为监测河南省禾谷镰孢菌对苯醚甲环唑的抗药性发展和防控小麦赤霉病合理用药提供依据。     

Baseline sensitivity of Monilia yunnanensis to the DMI fungicides tebuconazole and triadimefon

Monilia yunnanensis was recently identified as a new species causing brown rot of peach in China. Sterol 14α-demethylase inhibitors (DMIs) continue to be important in the management of brown rot of Monilinia spp. worldwide. Tebuconazole and triadimefon are two kinds of DMI fungicides that may be used for brown rot control in China. To establish the baseline sensitivity of M. yunnanensis to these two DMI fungicides, 203 M. yunnanensis single spore isolates were collected. Measurements of sensitivity to the two fungicides were based on inhibition of mycelial growth. For both fungicides, the sensitivity distribution was a unimodal curve, with an EC₅₀ range (the effective concentration to inhibit mycelial growth by 50 %) of 0.0001–0.0644 μg/ml for tebuconazole and 0.2311–1.7477 μg/ml for triadimefon. The M. yunnanensis isolates were obtained from orchards where DMI fungicides have not been used for peach brown rot control, thus the fungicide sensitivity distribution established in this study can be considered as the baseline for monitoring the resistance development in M. yunnanensis once the DMI fungicides are used to control peach brown rot.     

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.     

Changes in populations of the eyespot fungi Tapesia yallundae and T. acuformis under different fungicide regimes in successive crops of winter wheat, 1984–2000

Effects of regular treatments with the fungicides carbendazim and prochloraz applied to whole plots divided into subplots with different initial population mixtures of carbendazim-sensitive or carbendazim-resistant Tapesia yallundae or T. acuformis were studied in successive crops of winter wheat from 1984/85 to 1999/2000. In unsprayed and carbendazim-sprayed whole plots, a stable coexistence of about 50% each of T. yallundae and T. acuformis developed within five seasons, but in whole plots sprayed with prochloraz or prochloraz plus carbendazim, the proportion of T. acuformis increased to > 80%. A discrete time difference equation model was derived from knowledge of the biology of eyespot and competition theory to describe the population changes. The model was fitted to the data from treatments where coexistence occurred [subplots in unsprayed (1985–92) and carbendazim-sprayed (1985–89) whole plots], using nonlinear least squares regression. The optimized value of the resource overlap coefficient was small, suggesting niche differences between the two species. Populations were nearly 100% carbendazim-resistant in carbendazim-sprayed whole plots by July 1985 (one season) and in whole plots sprayed with prochloraz plus carbendazim by July 1986 (two seasons). In prochloraz-sprayed whole plots, the proportion of carbendazim-resistant isolates decreased more rapidly than in unsprayed whole plots in the 1980s, but by July 1992 a shift in populations in unsprayed and prochloraz-sprayed whole plots towards predominantly carbendazim-resistant strains had occurred.