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.     

Peronosclerospora maydis found on maize, sweetcorn and plume sorghum in Far North Queensland

Symptoms of a downy mildew disease were recognized on maize growing in the Atherton Tableland and Lakeland Downs areas of Far North Queensland in 1985. Quarantine measures were invoked to prevent the spread of this potentially serious disease to other parts of Australia.
The pathogen was identified as Peronosclerospora maydis in 1986 following examination of conidiophores with conidia and host range studies. An alternative host was strongly suspected and a survey near fields where the disease had been prevalent led to the identification of P. maydis on Sorghum plumosum, a grass indigenous to northern Australia. All downy mildew outbreaks on maize and sweetcorn in northern Australia since 1970 have been in areas where S. plumosum occurs. It seems likely that P. maydis has been present in Australia for many years.     

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。     

20%唑菌酯悬浮剂对黄瓜霜霉病的作用方式

采用盆栽植株喷雾-叶碟法、叶碟喷雾法分别测定20%唑菌酯悬浮剂对黄瓜霜霉病的保护、治疗作用,采用离体叶片法、叶碟法测定其对黄瓜霜霉病的铲除作用。结果表明:在200 μg/mL浓度下,20%唑菌酯悬浮剂对黄瓜霜霉病的保护、治疗效果分别为77.99%、70.08%;20%唑菌酯悬浮剂对黄瓜霜霉病菌病斑扩展、产孢及再侵染的抑制率分别为74.37%、84.56%和72.22%,与黄瓜霜霉病菌孢子囊悬浮液混合接种能显著降低孢子囊的致病性。室内试验结果表明,20%唑菌酯悬浮剂对黄瓜霜霉病具有良好的保护、治疗作用,且效果均优于对照药剂25%嘧菌酯悬浮剂,20%唑菌酯悬浮剂对黄瓜霜霉病的铲除作用与25%嘧菌酯悬浮剂效果相当。     

山东省和河北省黄瓜霜霉病菌对烯酰吗啉及双炔酰菌胺的抗性监测

 为明确黄瓜霜霉病菌对烯酰吗啉及双炔酰菌胺的抗性时空动态及这两种药剂对黄瓜霜霉病的田间防效,采用叶盘漂浮法检测了2011~2016年采自河北省和山东省黄瓜主产区的霜霉病菌1 821个菌株对烯酰吗啉及双炔酰菌胺的敏感性,并采用茎叶喷雾法对这两种药剂的田间防效进行评估。结果表明:黄瓜霜霉病菌1 821个菌株对烯酰吗啉和双炔酰菌胺抗性频率分别为88.5%和34.3%,供试菌株中对烯酰吗啉的低抗菌株及对双炔酰菌胺的敏感菌株占优势,平均抗性倍数分别为8.92和2.44,抗性指数分别为0.42和0.27。黄瓜霜霉病菌对这两种药剂的抗性频率、抗性倍数及抗性指数随着监测年限及地域的不同而波动。在河北定兴和山东寿光日光温室进行的田间药效试验中,按照药剂田间推荐剂量喷施4次,50%烯酰吗啉可湿性粉剂和250 g·L^-1双炔酰菌胺悬浮剂对黄瓜霜霉病表现出良好的防效(85%以上),与两地黄瓜霜霉病菌对烯酰吗啉产生低抗、对双炔酰菌胺敏感的监测结果一致。由此推断,50%烯酰吗啉可湿性粉剂和250 g·L^-1双炔酰菌胺悬浮剂可在对甲霜灵(或精甲霜灵)、噁霜灵及嘧菌酯普遍产生抗性的地区,作为58%甲霜灵·代森锰锌WP、68%精甲霜灵·代森锰锌WG、64%噁霜灵·代森锰锌WP 和250 g·L^-1嘧菌酯SC的替代药剂单独使用或与不同作用机理的杀菌剂混用,防治黄瓜霜霉病。     

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.     

基于流式细胞术的黄瓜霜霉病菌孢子囊计数研究

 为病原菌时空流行动态监测及病害预测模型构建提供病原菌快速计数方法,以卵菌中代表——葫芦科霜霉病病原菌古巴假霜霉菌(Pseudoperonospora cubensis)为例,研究了孢子囊悬浮液的荧光染色方法和基于流式细胞术计数方法。研究表明EDTA和SYBR Green Ⅰ 的联合作用能够对孢子囊进行荧光标记。对SYBR Green Ⅰ 染色剂的最佳染色条件进行研究,结果表明在染色时间相同时,荧光染色剂浓度越高,染色效果越好;同一浓度荧光染色剂,随着染色时间的增加,孢子囊染色率增加。使用流式细胞术对荧光染色的孢子囊进行计数,并与显微镜血球计数板计数结果进行比较,结果显示:流式细胞仪计数结果与显微镜血球计数板计数结果得到的孢子囊浓度没有显著差异(P<0.01),对两种方法获得的孢子囊浓度取对数后进行相关性分析,显示流式细胞仪测定的孢子囊浓度与显微镜血球计数板计数得到的孢子囊浓度高度相关,相关系数为 0.993 4,表明流式细胞术可以应用于古巴假霜霉菌孢子囊的计数。     

Quantitative aspects of the spread of asian soybean rust in the southeastern United States, 2005 to 2006

ABSTRACT The regional dynamics of soybean rust, caused by Phakopsora pachyrhizi, in six southeastern states (Florida, Georgia, Alabama, South Carolina, North Carolina, and Virginia) in 2005 and 2006 were analyzed based on disease records collected as part of U.S. Department of Agriculture's soybean rust surveillance and monitoring program. The season-long rate of temporal disease progress averaged approximately 0.5 new cases day(1) and was higher in nonsentinel soybean (Glycine max) plots than in sentinel soybean plots and kudzu (Pueraria lobata) plots. Despite the early detection of rust on kudzu in January and/or February each year (representing the final phase of the previous year's epidemic), the disease developed slowly during the spring and early summer on this host species and did not enter its exponential phase until late August, more than 1 month after it did so on soybean. On soybean, cases occurred very sporadically before the beginning of July, after which their number increased rapidly. Thus, while kudzu likely provides the initial inoculum for epidemics on soybean, the rapid increase in disease prevalence on kudzu toward the end of the season appears to be driven by inoculum produced on soybean. Of 112 soybean cases with growth stage data, only one occurred during vegetative crop development while approximately 75% occurred at stage R6 (full seed) or higher. The median nearest-neighbor distance of spread among cases was approximately 70 km in both years, with 10% of the distances each being below approximately 30 km and above approximately 200 km. Considering only the epidemic on soybean, the disease expanded at an average rate of 8.8 and 10.4 km day(1) in 2005 and 2006, respectively. These rates are at the lower range of those reported for the annual spread of tobacco blue mold from the Caribbean Basin through the southeastern United States. Regional spread of soybean rust may be limited by the slow disease progress on kudzu during the first half of the year combined with the short period available for disease establishment on soybean during the vulnerable phase of host reproductive development, although low inoculum availability in 2005 and dry conditions in 2006 also may have reduced epidemic potential.     

Relationship between Loci conferring downy mildew and powdery mildew resistance in melon assessed by quantitative trait Loci mapping

ABSTRACT Partial resistance to downy mildew (Pseudoperonospora cubensis) and complete resistance to powdery mildew (Podosphaera xanthii races 1, 2, 3, and 5 and Golovinomyces cichoracearum race 1) were studied using a recombinant inbred line population between 'PI 124112' (resistant to both diseases) and 'Védrantais' (susceptible line). A genetic map of melon was constructed to tag these resistances with DNA markers. Natural and artificial inoculations of Pseudoperonospora cubensis were performed and replicated in several locations. One major quantitative trait loci (QTL), pcXII.1, was consistently detected among the locations and explained between 12 to 38% of the phenotypic variation for Pseudoperonospora cubensis resistance. Eight other Pseudoperonospora cubensis resistance QTL were identified. Artificial inoculations were performed with several strains of four races of Podosphaera xanthii and one race of G. cichoracearum. Two independent major genes, PmV.1 and PmXII.1, were identified and shown to be involved in the simple resistance to powdery mildew. Three digenic epistatic interactions involving four loci were detected for two races of Podosphaera xanthii and one race of G. cichoracearum. Co-localization between PmV.1, resistance genes, and resistance genes homologues was observed. Linkage between the major resistance QTL to Pseudoperonospora cubensis, pcXII.1, and one of the two resistance genes to powdery mildew, PmXII.1, was demonstrated.     

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.     

Relationship between disease severity and escape of Pseudoperonospora cubensis sporangia from a cucumber canopy during downy mildew epidemics

Fundamental to the development of models to predict the spread of cucurbit downy mildew is the ability to determine the escape of Pseudoperonospora cubensis sporangia from infected fields. Aerial concentrations of sporangia, C (sporangia m^?3), were monitored using Rotorod samplers deployed at 0·5 to 3·0 m above a naturally infected cucumber canopy in two sites in central and eastern North Carolina in 2011, where disease severity ranged from 1 to 40%. Standing crop of sporangia was assessed each morning at 07·00 h EDT and ranged from 320 to 16 170 sporangia m^?2. Disease severity and height above the canopy significantly (P < 0·0001) affected C with mean concentration (C_m) being high at moderate disease. Values of C_m decreased rapidly with canopy height and at a height of 2·0 m, C_m was only 7% of values measured at 0·5 m when disease was moderate. Daily total flux (F_D) was dependent on disease severity and ranged from 5·9 to 2242·3 sporangia m^?2. The fraction of available sporangia that escaped the canopy increased from 0·028 to 0·171 as average wind speed above the canopy for periods of high C increased from 1·7 to 3·6 m s^?1. Variations of C_m and F_D with increasing disease were well described (P < 0·0001) by a log‐normal model with 15% as the threshold above which C_m and F_D decreased as disease severity increased. These results indicate that disease severity should be used to adjust sporangia escape in spore transport simulation models that are used to predict the risk of spread of cucurbit downy mildew.     

Occurrence and molecular characterization of strobilurin resistance in cucumber powdery mildew and downy mildew

ABSTRACT Between 1998 and 1999, control failure of powdery mildew (Podosphaera fusca) and downy mildew (Pseudoperonospora cubensis) by the strobilurin fungicides azoxystrobin and kresoxim-methyl was observed in cucumber-growing areas of Japan. Results from inoculation tests carried out on intact cucumber plants and leaf disks clearly showed the distribution of pathogen isolates highly resistant to azoxystrobin and kresoximmethyl. Fragments of the fungicide-targeted mitochondrial cytochrome b gene were polymerase chain reaction amplified from total pathogen DNA and their sequences analyzed to elucidate the molecular mechanism of resistance. A single point mutation (GGT to GCT) in the cytochrome b gene, resulting in substitution of glycine by alanine at position 143, was found in resistant isolates of downy mildew. This substitution in cytochrome b seemed to result in high resistance to strobilurins in this pathogen. The same mutation was found in some but not all resistant isolates of powdery mildew. This study suggests that a mutation at position 143 in the target-encoding gene, resulting in an amino acid substitution, was probably a major cause of the rapid development of high strobilurin resistance in these two pathogens.     

霜脲氰与不同杀菌剂混配对两种霜霉病菌的增效作用

采用叶盘法（定量喷药）测定霜脲氰与不同杀菌剂混配对黄瓜和葡萄霜霉病菌抗甲霜灵菌株的增效作用,证实了霜脲氰加代森锰锌或二者再加恶霜灵以不同比率混配对黄瓜、葡萄霜霉病菌均明显增效,发现了霜脲氰与碱式硫酸铜1：8混配对黄瓜霜霉病菌增交,代森锰锌（1：8）或再加恶霜灵（1：8：1）对葡萄霜霉病菌的增效作用随着供试菌株对甲霜灵抗性水平的提高而增强。霜脲氰与代森锰锌1：7混配以保护、治疗、铲除方式处理黄瓜叶盘,对黄瓜霜霉病菌均表现增效。     

Spatiotemporal analysis of epiphytotics of downy mildew of oilseed poppy in tasmania, australia

Downy mildew, caused by Peronospora arborescens, has become the major disease affecting oilseed poppy (Papaver somniferum) since its first record in Tasmania in 1996. Two field trials conducted in 2000 and 2001 studied the progression and spatial distribution of downy mildew epiphytotics. The logistic and exponential models best described the progression of disease incidence and severity, respectively. Incidence and severity increased rapidly following canopy closure. In 2001, incidence increased from 0.16%, prior to canopy closure, to 100% at late flowering (40 days). Spatial analyses of epiphytotics were conducted by fitting the beta-binomial and binomial distributions, median runs analysis, and the spatial analysis by distance indices. All analyses demonstrated that the distribution of incidence and severity was strongly spatially aggregated from canopy closure until at least late flowering. These results suggest that secondary spread from a few primary infections is the major factor in epiphytotics.     

Spatial analysis of lettuce downy mildew using geostatistics and geographic information systems

ABSTRACT The epidemiology of lettuce downy mildew has been investigated extensively in coastal California. However, the spatial patterns of the disease and the distance that Bremia lactucae spores can be transported have not been determined. During 1995 to 1998, we conducted several field- and valley-scale surveys to determine spatial patterns of this disease in the Salinas valley. Geostatistical analyses of the survey data at both scales showed that the influence range of downy mildew incidence at one location on incidence at other locations was between 80 and 3,000 m. A linear relationship was detected between semivariance and lag distance at the field scale, although no single statistical model could fit the semi-variograms at the valley scale. Spatial interpolation by the inverse distance weighting method with a power of 2 resulted in plausible estimates of incidence throughout the valley. Cluster analysis in geographic information systems on the interpolated disease incidence from different dates demonstrated that the Salinas valley could be divided into two areas, north and south of Salinas City, with high and low disease pressure, respectively. Seasonal and spatial trends along the valley suggested that the distinction between the downy mildew conducive and nonconducive areas might be determined by environmental factors.     

Downy mildew outbreaks on grapevine under climate change: elaboration and application of an empirical‐statistical model*

The global climate is changing. Much research has already been carried out to assess the potential impacts of climate change on plant physiology. However, effects on plant disease have not yet been deeply studied. In this paper, an empirical disease model for primary infection of downy mildew on grapevine was elaborated and used to project future disease dynamics under climate change. The disease model was run under the outputs of the General Circulation Model (GCM) and future scenarios of downy mildew primary outbreaks were generated at several sites all over the word for three future dates: 2030, 2050, 2080. Results suggested a potential general advance of first disease outbreaks, both in the Northern and Southern Hemispheres, for all three future decades considered. The advance is predicted to be from about a minimum of one day in South Africa in 2030 to a maximum of 28 days in Chile and China in 2080. The advance in the outbreak time could lead to more severe infections, due to the polycyclic nature of the pathogen. Therefore, changes in the timing and frequency of fungicide treatments could be expected in the future, with a possible increase in the costs of disease management.     

Importance of secondary inoculum of Plasmopara viticola to epidemics of grapevine downy mildew

To quantify the magnitude and the spatial spread of grapevine downy mildew secondary sporangia, 4685 Plasmopara viticola single lesion samples were collected from 18 plots spread across central Europe. Disease symptoms were collected on two to 22 sampling dates per plot between 2000 and 2002. Four multiallelic microsatellite markers were used for genotypic identification of pathogen samples. Genetic analysis showed more than 2300 site-specific P. viticola genotypes, indicating that populations are genetically rich demographic units. Approximately 70% of the genotypes were sampled once and 14% were sampled twice throughout the various epidemics. In the 18 populations only seven genotypes (0.3%) were identified more than 50 times. Three genotypes particularly successful in causing disease through secondary cycles showed mainly a clustered distribution. The distance of sporangial migration per secondary cycle was less than 20 m and their plot colonization rate was calculated at around 1–2 m² day⁻¹. Downy mildew epidemics of grapevine are therefore the result of the interaction of a multitude of genotypes, each causing limited (or a few) lesions, and of a dominant genotype able to spread stepwise at plot-scale. These findings contrast with current theories about grapevine downy mildew epidemiology, which postulate that there is massive vineyard colonization by one genotype and long-distance migration of sporangia.     

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     

Effect of downy mildew development on transpiration of cucumber leaves visualized by digital infrared thermography

ABSTRACT Disease progress of downy mildew on cucumber leaves, caused by the obligate biotrophic pathogen Pseudoperonospora cubensis, was shown to be associated with various changes in transpiration depending on the stage of pathogenesis. Spatial and temporal changes in the transpiration rate of infected and noninfected cucumber leaves were visualized by digital infrared thermography in combination with measurements of gas exchange as well as microscopic observations of pathogen growth within plant tissue and stomatal aperture during pathogenesis. Transpiration of cucumber leaf tissue was correlated to leaf temperature in a negative linear manner (r = -0.762, P < 0.001, n = 18). Leaf areas colonized by Pseudoperonospora cubensis exhibited a presymptomatic decrease in leaf tem perature up to 0.8 degrees C lower than noninfected tissue due to abnormal stomata opening. The appearance of chlorosis was associated with a cooling effect caused by the loss of integrity of cell membranes leading to a larger amount of apoplastic water in infected tissue. Increased water loss from damaged cells and the inability of infected plant tissue to regulate stomatal opening promoted cell death and desiccation of dying tissue. Ultimately, the lack of natural cooling from necrotic tissue was associated with an increase in leaf temperature. These changes in leaf temperature during downy mildew development resulted in a considerable heterogeneity in temperature distribution of infected leaves. The maximum temperature difference within a thermogram of cucumber leaves allowed the discrimination between healthy and infected leaves before visible symptoms appeared.     

葡萄抗霜霉病机制研究进展

葡萄霜霉病是危害葡萄的重要病害之一,该病害发生时会给葡萄生产造成严重的经济损失。本文综合分析了国内外对葡萄抗霜霉病机制研究的结果,认为葡萄对霜霉病抗性受多种因素影响,主要表现为生理生化物质、叶片形态结构及抗病遗传基因三个方面。由于各种因素对霜霉病病原菌作用关系复杂,目前对于葡萄抗霜霉病的决定性因子还没有明确结论。