Resistance of Pseudoperonospora cubensis to flumorph on cucumber in plastic houses

Plastic-house experiments were conducted over a 2-year period (2004–05) to estimate the effects of successive applications of flumorph or a mixture of flumorph with mancozeb to cucumber plants on selection for flumorph resistance in the downy mildew oomycete, Pseudoperonospora cubensis . Application of flumorph alone favoured the selection of resistant isolates of Ps. cubensis . Resistant populations were detected at a frequency of 2·5% after six successive applications of flumorph alone in a plastic house in 2004. Resistant isolates were also detected (4·8%) after eight successive applications of the mixture of flumorph and mancozeb in 2004, although the mixture gave significantly better disease control than flumorph alone and produced a slight delay in the development of resistance. In a second cucumber crop in the same plastic houses in 2004, the frequency of resistant isolates increased to 100% after three successive applications of flumorph or four of flumorph + mancozeb. Under laboratory conditions, most flumorph-resistant isolates showed high levels of resistance and their levels of pathogenicity and sporulation were as high as that of wild-type isolates. Flumorph showed cross-resistance with dimethomorph and iprovalicarb, but not with azoxystrobin, cyazofamid, cymoxanil or metalaxyl. These studies suggest a high risk for the occurrence of resistance to flumorph in Ps. cubensis in cucumber crops under plastic-house conditions.     

寡糖类物质诱导黄瓜对霜霉病的抗性

霜霉病是黄瓜栽培中发生最为普遍的病害之一，目前生产上主要利用化学药剂来防治，存在着农药残留的现象，从而影响了无公害绿色食品的生产。寡糖是新一类生物诱抗剂，具有可自然降解，不污染环境，无残留等优点。本文以酶解方法从海带中获得寡糖类物质，利用其防治黄瓜霜霉病[Pseudoperonospora cubensis(Berk．et Curt．)Rostov．]取得了一定的效果。现将研究结果报道如下。     

黄瓜霜霉病菌保存方法

 Pseudoperonospora cubensis(Berk. & M. A. Curtis) Rostovzev, the causal agent of cucumber downy mildew, is an obligate parasitic fungus. Up to now it can not be preserved on culture medium. In this study, the sporangia of P. cubensis were preserved in protective substances of 10% dimethyl sulfoxide plus 5% skim milk, 10% dimethyl sulfoxide plus 10% skim milk, sterilized water and leaves in vitro, and stored at -20℃, -70℃, and preliminary freezing at -20℃ for 24 h prior to -70℃ respectively. The results showed that the sporangia preserved in 10% dimethyl sulfoxide plus 5% skim milk, preliminary freezing at -20℃ for 24 h prior to -70℃ were still highly pathogenic after 12 months preservation. The percentage of germination of sporangia, incidence and disease index were 46%, 50.0% and 40.0 respectively 9 d after inoculation. This method solved the problem of P. cubensis preservation.     

Resistance mechanism to carboxylic acid amide fungicides in the cucurbit downy mildew pathogen Pseudoperonospora cubensis

BACKGROUND: Pseudoperonospora cubensis, the causal oomycete agent of cucurbit downy mildew, is responsible for enormous crop losses in many species of Cucurbitaceae, particularly in cucumber and melon. Disease control is mainly achieved by combinations of host resistance and fungicide applications. However, since 2004, resistance to downy mildew in cucumber has been overcome by the pathogen, thus driving farmers to rely only on fungicide spray applications, including carboxylic acid amide (CAA) fungicides. Recently, CAA‐resistant isolates of P. cubensis were recovered, but the underlying mechanism of resistance was not revealed. The purpose of the present study was to identify the molecular mechanism controlling resistance to CAAs in P. cubensis. RESULTS: The four CesA (cellulose synthase) genes responsible for cellulose biosynthesis in P. cubensis were characterised. Resistant strains showed a mutation in the CesA3 gene, at position 1105, leading to an amino acid exchange from glycine to valine or tryptophan. Cross‐resistance tests with different CAAs indicated that these mutations lead to resistance against all tested CAAs. CONCLUSION: Point mutations in the CesA3 gene of P. cubensis lead to CAA resistance. Accurate monitoring of these mutations among P. cubensis populations may improve/facilitate adequate recommendation/deployment of fungicides in the field. Copyright © 2011 Society of Chemical Industry     

Resistance to acylalanines in Phytophthora infestans in The Netherlands1

In The Netherlands in 1980 a severe late-blight epidemic involving metalaxyl-resistant strains of Phytophthora infestans did considerable damage to the potato crop. As a consequence metalaxyl or metalaxyl-containing products were withdrawn from the Dutch fungicide market for the control of potato late blight. In 1981 the majority of the P. infestans isolates obtained from various parts of the country were sensitive to metalaxyl. In three areas where metalaxyl mixtures were used on a relatively large scale metalaxyl-resistant strains dominated the population. Incidental use of metalaxyl mixtures in 1982 and 1983 showed adequate late-blight control and, when in 1984 an early and in some areas severe epidemic developed, a metalaxyl/mancozeb mixture was reintroduced for curative application in combination with conventional fungicides. Although disease control was adequate, resistant strains were present in a small number of fields. Whether resistant strains will increase in frequency and threaten the usefulness of mixtures of acylalanines will heavily depend on how well potato farmers adopt strategies involving a limited use of mixtures of acylalanines in a spray schedule with conventional late-blight fungicides.     

Assessing the risk of resistance in Pseudoperonospora cubensis to the fungicide flumorph in vitro

BACKGROUND: The oomycete fungicide flumorph is a recently introduced carboxylic acid amide (CAA) fungicide. In order to evaluate the risk of developing field resistance to flumorph, the authors compared it with dimethomorph and azoxystrobin with respect to the ease of obtaining resistant isolates to these fungicides, the level of resistance and their fitness in the laboratory. RESULTS: Mutants with a high level of resistance to azoxystrobin were isolated readily by adaptation and UV irradiation, and their fitness was as good as that of the parent isolates. Attempts to generate mutants of Pseudoperonospora cubensis (Burk. & MA Curtis) Rostovsev resistant to flumorph and dimethomorph by sporangia adaptation on fungicide-treated leaves were unsuccessful. However, moderately resistant mutants were isolated using UV mutagenesis, but their resistance level [maximum resistance factor (MRF) < 100] was much lower than that of the azoxystrobin-resistant mutant (MRF = 733). With the exception of stability of resistance, all mutants showed low pathogenicity and sporulation compared with wild-type isolates and azoxystrobin-resistant mutants. There is cross-resistance between flumorph and dimethomorph, suggesting that they have the same resistance mechanism. CONCLUSION: The above results suggest that the resistance risk of flumorph may be similar to that of dimethomorph but lower than that of azoxystrobin and can be classified as moderate. Thus, it can be managed by appropriate product use strategies.     

基于SRAP标记的黄瓜霜霉菌遗传多样性分析

本研究对来自黑龙江、辽宁、河北、山东、江苏5个省12个黄瓜主产区的77个黄瓜霜霉菌菌株,采用SRAP分子标记进行了遗传多样性分析。从35对SRAP引物中筛选出10对引物,共产生9 554条扩增条带,其中9 132条表现多态性,占95.6%。基于SRAP分子标记,77个黄瓜霜霉菌菌株的遗传距离为0.60~1.00,表明黄瓜霜霉菌具有丰富的遗传多样性。通过聚类分析,77个黄瓜霜霉菌菌株聚类为8个类群,并且来自相同地区的菌株大多数聚集于同一类群中,表明黄瓜霜霉菌群体的遗传多样性与其地理来源密切相关。     

黄瓜霜霉病菌抗氟吗啉菌株的适合度研究

 采用离体叶盘法和活体植株法,测定了从田间分离获得的对氟吗啉具有抗药性的黄瓜霜霉病菌菌株和通过室内紫外线诱变获得的对氟吗啉及对照药剂烯酰吗啉和嘧菌酯产生抗药性的突变菌株的生存适合度。试验结果表明,室内诱导获得的氟吗啉和烯酰吗啉抗药性突变菌株的适合度明显低于敏感菌株和嘧菌酯抗药性突变体及田产氟吗啉抗药性菌株,主要表现在菌株的侵染率、产孢能力和致病力的减弱。室内获得的氟吗啉和烯酰吗啉抗药性突变体和敏感菌株混合接种、继代培养后,其抗药菌株的检出率在混合群体中逐渐降低,其中抗性突变体FR4和敏感菌株以2:8的比例混合接种、培养7代后,其无性后代中检测不到抗药性菌株的存在,表明室内氟吗啉抗药性突变体竞争力弱于敏感菌株;而田间氟吗啉抗药性菌株和室内嘧菌酯抗药性突变体则表现出和敏感菌株相似或更强的竞争力,预示田间自然情况下抗药性菌株存在着可能形成优势菌群而导致药剂防治失败的潜在风险。     

Occurrence of metalaxyl-resistant isolates of Pseudoperonospora cubensis in Israel: a 5-year survey1

The occurrence of metalaxyl-resistant isolates of Pseudoperonospora cubensis, the causal agent of downy mildew in cucurbits, was surveyed during 1980–1984 subsequent to their first appearance in December 1979. Metalaxyl use was abandoned in mid-1981 and resistant isolates diminished in late 1981. Metalaxyl use was renewed in 1982 and a gradual increase in frequency of resistant (R) strains was detected. In 1983, 31 out of 40, and in 1984, 43 out of 47 isolates tested were highly resistant to the fungicide with 65 and 91% of them, respectively, collected from plastic houses or fields not treated with metalaxyl. The survey indicates a rapid build-up of R strains of the fungus and the need for alternative chemicals for combating downy mildew in cucurbits.     

Optimization of chemical control of Pseudoperonospora cubensis on cucumber in Ukraine

Downy mildew ( Pseudoperonospora cubensis ) is one of the most harmful diseases of cucumber in Ukraine. At high disease severity, the growing season is considerably shortened and yield is reduced. The most effective method of control is the development of resistant cultivars but, in the absence of absolutely resistant forms, control with fungicides remains very important. The effectiveness of such treatment depends on the dates of application, the number of sprays and the efficacy of the fungicides used. An improved scheme is presented, based on three components: use of cultivars with increased resistance; application of highly effective systemic + contact fungicides; and reliable short-term forecasting of disease appearance. The predictors for downy mildew forecasting are: sum of effective temperatures, weather index, duration of leaf wetness, concentration of sporangia in air and latent period of disease development. These conditions determine'critical days'for disease development. The first fungicide spray on less susceptible cultivars is recommended on the first critical day, and the next on the day of appearance of first disease symptoms. This scheme allows the fungicide load to be approximately halved, while ensuring higher yield.     

Genetic and pathogenic relatedness of Pseudoperonospora cubensis and P. humuli

The most economically important plant pathogens in the genus Pseudoperonospora (family Peronosporaceae) are Pseudoperonospora cubensis and P. humuli, causal agents of downy mildew on cucurbits and hop, respectively. Recently, P. humuli was reduced to a taxonomic synonym of P. cubensis based on internal transcribed spacer (ITS) sequence data and morphological characteristics. Nomenclature has many practical implications for pathogen identification and regulatory considerations; therefore, further clarification of the genetic and pathogenic relatedness of these organisms is needed. Phylogenetic analyses were conducted considering two nuclear and three mitochondrial loci for 21 isolates of P. cubensis and 14 isolates of P. humuli, and all published ITS sequences of the pathogens in GenBank. There was a consistent separation of the majority of the P. humuli isolates and the P. cubensis isolates in nuclear, mitochondrial, and ITS phylogenetic analyses, with the exception of isolates of P. humuli from Humulus japonicus from Korea. The P. cubensis isolates appeared to contain the P. humuli cluster, which may indicate that P. humuli descended from P. cubensis. Host-specificity experiments were conducted with two reportedly universally susceptible hosts of P. cubensis and two hop cultivars highly susceptible to P. humuli. P. cubensis consistently infected the hop cultivars at very low rates, and sporangiophores invariably emerged from necrotic or chlorotic hypersensitive-like lesions. Only a single sporangiophore of P. humuli was observed on a cucurbit plant during the course of the studies. Together, molecular data and host specificity indicate that there are biologically relevant characteristics that differentiate P. cubensis and P. humuli that may be obfuscated if P. humuli were reduced to a taxonomic synonym of P. cubensis. Thus, we recommend retaining the two species names P. cubensis and P. humuli until the species boundaries can be resolved unambiguously.     

6种染色方法对黄瓜霜霉病菌不同发育阶段的染色效果比较

 比较了Calcofluor、苯胺蓝、荧光素钠3种荧光染色方法和曲利苯兰酒精-乳酚酸、考马斯亮兰和苯胺蓝组织透明染色法对黄瓜霜霉病菌不同生长发育阶段的染色效果。结果表明,在一定的染液浓度和pH值条件下,3种荧光染色方法均可进行孢子囊、休止孢、芽管、附着胞、孢囊梗等寄主组织外部菌体的原位染色,其中Calcofluor染色方法更稳定、简捷和易于观察。曲利苯兰组织透明染色法能观察黄瓜霜霉病菌的整个生长发育阶段,但该法更适宜胞间菌丝及吸器的染色。     

葡聚六糖诱导黄瓜抗霜霉病的研究

黄瓜经喷施人工合成的植物抗病性诱导剂葡聚六糖诱导后 ,黄瓜霜霉病的发病程度显著降低。葡聚六糖诱导黄瓜抗霜霉病的效果以质量浓度为10μg/L ,诱导间隔期为7d ,连续诱导 3次效果较好 ,其诱导持续期可达 7d。结果还表明葡聚六糖的诱导抗性表达不仅与它本身的作用机制有关 ,还与它的使用方法有关     

Structure and temporal shifts in virulence of Pseudoperonospora cubensis populations in the Czech Republic

The structure and temporal dynamics of the virulence of Pseudoperonospora cubensis (causal agent of cucurbit downy mildew) were studied in pathogen populations in the Czech Republic from 2001 to 2010. A total of 398 P. cubensis isolates collected from Cucumis (Cm.) sativus, Cm. melo, Cucurbita (Cr.) maxima, Cr. pepo, Cr. moschata and Citrullus lanatus were analysed for variation in virulence (pathotypes). Virulence was evaluated on a differential set of 12 genotypes of cucurbitaceous plants. All isolates of P. cubensis were characterized by their level of virulence (classified according the number of virulence factors, VF; low VF = 1–4, medium VF = 5–8, high VF = 9–12): high (75%), medium (24%) and low (1%). The structure and dynamics of virulence in the pathogen populations were expressed by pathotypes using tetrad numerical codes and a total of 67 different pathotypes of P. cubensis were determined. The most susceptible group of differentials was Cucumis spp., while the lowest frequency of virulence was recorded on Cr. pepo ssp. pepo, Ci. lanatus and Luffa cylindrica. A high proportion (c. 90%) of isolates was able to infect cucurbit species Benincasa hispida and Lagenaria siceraria, which are not commonly cultivated in the Czech Republic or elsewhere in central Europe. In the recent pathogen populations (2008–2010) there was prevailing frequency (70–100%) of isolates with high numbers (9–12) of virulence factors. ‘Super pathotype’ 15.15.15 was often observed in the study within the pathogen populations and was one of the four most frequently recorded pathotypes. Pseudoperonospora cubensis populations shifted to a higher virulence over time. From 2009 the pathogen population changed dramatically and new pathotypes appeared able to establish natural and serious infection of Cucurbita spp. and Ci. lanatus, which was not observed in 2001–2008. Generally, virulence structure and dynamics of P. cubensis populations are extremely variable in the Czech Republic.     

干旱区甜瓜霜霉病远距离传播空间结构的初步研究

 经过9年在塔里木盆地边缘干旱区对伽师县等3个相互隔离的甜瓜霜霉病流行系统的实地调查研究,提出了甜瓜霜霉病远距离传播的三级空间结构。一个甜瓜霜霉病流行系统单元是在一个绿洲内,围绕可提供初侵染病原的黄瓜温室区而形成的,其空间结构分为区系(包含若干种植区)、种植区(包含若干田块)和田块三级结构。伽师甜瓜霜霉病流行区系分为3个种植区,甜瓜霜霉病每年春季最先从位于种植一区发生霜霉病的黄瓜温室附近的甜瓜开始发生,之后甜瓜霜霉病由近向远逐步传播。种植二区和种植三区甜瓜霜霉病的初发病时间(初发病中心出现的时间)比种植一区的分别平均晚20和45d,发病中心与初菌源地的平均距离分别是56.4和106.5km,表明此地甜瓜霜霉病菌远距离传播经历了从种植一区到种植二区,再从种植二区到种植三区的传播过程。     

新型杀菌剂吲唑磺菌胺对黄瓜霜霉病的防治效果研究

2012和2013两年田间药效试验结果表明,吲唑磺菌胺200g/L悬浮剂对黄瓜霜霉病具有较好防效,使用80g(ai)/hm2的末次防效达82%以上,是防治黄瓜霜霉病较为理想的药剂,在推荐用量下使用对黄瓜安全。     

Screening of wildcucumis species against downy mildew (Pseudoperonospora cubensis) isolates from cucumbers

Under controlled inoculation, a set of 56 accessions belonging to 19 wild species of the genusCucumis was studied for resistance to seven isolates of cucurbit downy mildew (Pseudoperonospora cubensis (Berk. et Curt.) Rostow.) from cucumber. No resistance toP. cubensis was detected in theseCucumis accessions. In three host accession/pathogen isolate combinations, limited sporulation was observed. Nine newCucumis species are described as hosts forP. cubensis: C. africanus, C. ficifolius, C. figarei, C. meeusii, C. metuliferus, C. myriocarpus, C. leptodermis, C. sagittatus andC. zeyheri. Results are discussed in relation to the origin and evolution ofCucumis species.