Variability of Peronospora Sparsa (syn. P. rubi) in Finland as Measured by Amplified Fragment Length Polymorphism

Downy mildew caused by Peronospora sparsa (syn. P. rubi) is a serious threat to commercial cultivation of arctic bramble (Rubus arcticus subsp. arcticus) in Finland. P. sparsa is distributed throughout the country in cultivated and wild arctic bramble and in cloudberry (R. chamaemorus). A total of 36 isolates of P. sparsa collected from these hosts was analysed for amplified fragment length polymorphism (AFLP). Of the 226 markers scored, 223 were polymorphic and all isolates of P. sparsa had unique AFLP fingerprints, which indicated high levels of genetic variability. An UPGMA clustering analysis of the isolates did not reveal any genetically distinguishable strains. The isolates were grouped neither according to the geographic origin nor the host from which they were isolated. Isolates of P. sparsa obtained from wild arctic bramble and one from cloudberry readily infected the leaves of the cultivated arctic bramble (cultivar Pima). Also, P. sparsa isolated from cultivated arctic bramble infected the leaves of wild arctic bramble. These data suggest that P. sparsa may be disseminated from wild arctic bramble and cloudberry to cultivated arctic bramble in the field.     

Induction of resistance to downy mildew (Peronospora parasitica) in cauliflower by DL-β-amino-n-butanoic acid (BABA)

Of the three isomers of aminobutyric acid, only the β isomer (BABA) was effective in inducing resistance against Peronospora parasitica , the causal agent of downy mildew, in cauliflower ( Brassica oleracea var. botrytis ). A single foliar spray applied to 7-day-old seedlings protected the plants against Peronospora parasitica for at least 15 days. Of the enantiomers (R and S), only the R was effective. Resistance was accompanied by a hypersensitive-like reaction (necrotic spots) which was evident before inoculation. BABA was systemically effective when applied to the roots, but failed to protect cotyledons adjacent to treated ones. Unlike other chemical inducers, BABA was effective when applied several hours postinoculation. It had no effect on P. parasitica spore germination. In cauliflower seedlings, BABA did not induce the accumulation of the pathogenesis-related protein PR-1, PR-2, PR-3, PR-5 and PR-9. Only treated and challenged seedlings accumulated PR-2.     

New genes for resistance to downy mildew (Peronospora parasitica) in oilseed rape (Brassica napus ssp. oleifera)

The response of cotyledons of oilseed rape ( Brassica napus ssp. oleifera ) accessions to infection by isolates of Peronospora parasitica under controlled conditions was assessed on a 0–7 scale (disease reaction). In interactions scored 0–3, 4–5 and 6–7, the host was considered resistant, partially resistant and susceptible, respectively. Accession RES-26, selected from the spring oilseed rape cultivar Janetzkis, was partially resistant to isolate R1 and resistant to isolate P003 of P. parasitica , which distinguishes it from three previously described differential response groups ('A', 'B' and 'C') of accessions in B. napus . The resistance of RES-26 to isolate P003 seemed to be conditioned by a single, partially dominant gene and the resistance of RES-02, which belongs to group 'A' (resistant to R1 and P003), by two independent partially dominant genes. The gene for resistance to P003 in RES-26 is either closely linked, allelic or identical to one of the two genes for resistance in RES-02. Resistance of RES-02 to R1 is conditioned by a single, incompletely dominant gene. The genes for resistance to isolates R1 and P003 in RES-02 are either closely linked, allelic or identical. The cotyledonary leaves of each seedling responded independently when inoculated simultaneously each with a different isolate of the pathogen.     

Proteomic analysis of a compatible interaction between Pisum sativum (pea) and the downy mildew pathogen Peronospora viciae

A proteomic approach was used to identify host proteins altering in abundance during Peronospora viciae infection of a susceptible cultivar of pea (Pisum sativum cv. Livioletta). Proteins were extracted from fully developed pea leaflets at 4 days post-inoculation, before visible symptoms were apparent. Cytoplasmic proteins and membrane- and nucleic acid-associated proteins from infected and control leaves were examined using two-dimensional difference gel electrophoresis. The majority of proteins had a similar abundance in control and infected leaves; however, several proteins were altered in abundance and twelve were found to have increased significantly in the latter. These proteins were selected for either matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry or electro-spray ionisation quadrupole time-of-flight tandem mass spectrometry analysis following trypsin digestion, with sequence identity being assigned to eight of the proteins. These included the ABR17 stress-response protein, the pathogen-induced PI176 protein, three photosynthetic proteins, a glycine-rich RNA binding protein and two glyceraldehyde 3-phosphate dehydrogenases (cytosolic and chloroplastic) which can be induced by a range of abiotic and biotic stresses in many plant species. The possible roles of these proteins in the response of the pea plant during P. viciae infection are discussed. This study represents the first proteomic analysis of downy mildew infection of pea leaves, and provides the basis for further work to elucidate molecular mechanisms of compatibility in P. viciae infections.     

Seedling and adult plant resistance to downy mildew (Peronospora parasitica) in cauliflower (Brassica oleracea convar. botrytis var. botrytis)

Seedlings of six cauliflower cultivars ( Brassica oleracea convar. botrytis var. botrytis ) were assessed for resistance to a Danish isolate of Peronospora parasitica , under controlled conditions. Resistance, characterized by restricted sporulation and necrotic dark flecks at the inoculation site on the cotyledons, was expressed in the hybrids 9306 F1, 9311 F1, and the open pollinated cultivar Perfection. Testing of the parent lines and F2 generations of the two resistant hybrids suggested that resistance was a dominantly inherited trait controlled by a single gene. Inoculation of the cultivars with seven isolates, from different geographical origins, showed that the resistance was isolate specific. The two hybrid cultivars expressing cotyledon resistance and two hybrids expressing susceptibility were assessed for adult plant resistance under field conditions. The AUDPC (Area Under the Disease Progress Curve), based on disease incidence and severity, revealed significant differences between the cultivars. At harvest, the cultivars exhibited significantly different levels of defoliation and curd attack. The cultivars 9306 F1 and 9311 F1 showed high levels of resistance in all assessments, whereas the two cultivars exhibiting susceptibility at the seedling stage, 9304 F1 and 9305 F1, also exhibited susceptibility through the adult plant stage. Thus, the resistance exhibited under field conditions resembled that identified at the seedling stage under controlled conditions. The results suggest that cotyledon resistance similar to that described could provide resistance throughout the adult plant stage, including curds.     

番茄细菌性溃疡病菌的实时荧光PCR检测

 由Clavibacter michiganensis subsp.michiganensis(Cmm)引起的番茄细菌性溃疡病是一种严重危害番茄生产的种传细菌性病害。根据ITS序列多态性设计引物及TaqMan探针进行实时荧光PCR检测的结果表明,这组引物一探针能检测出所有供试的Cmm菌,对照菌均未检测到荧光信号。用接种但未显示症状的番茄苗叶片及人工处理的带菌种子提取的核酸作为模板,均能检测到病菌,其检测灵敏度比常规PCR高约100倍。实验中不需病原菌的分离培养及PCR的后续处理。该方法快速、简便、安全、准确,适用于出入境检验检疫及种子、种苗健康检测领域。     

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.     

The identification of a gene for race-specific resistance to Peronospora parasitica (downy mildew) in Brassica napus var. oleifera (oilseed rape)

The oilseed rape cultivar Cresor was resistant to 14 isolates of Peronospora parasitica derived from crops of Brassica napus in the UK. Segregation for resistance to one isolate among F₂ plants and F₃ progeny of crosses between Cresor and the susceptible cultivars Victor and Jet Neuf indicated that resistance was controlled by a single gene. There was evidence that genetic background and environment could influence the phenotypic expression of this resistance. Two sexual progeny isolates derived from a homothallic isolate of P. parasitica avirulent on Cresor were completely virulent on this cultivar. This suggested that the parental isolate was heterozygous at a matching locus or loci for avirulence and demonstrated the race-specific nature of the resistance.     

Development and physiology of infection by the downy mildew fungus Peronospora parasitica (Pers. ex Fr.) Fr. in susceptible and resistant Brassica species

The growth of two isolates of the downy mildew fungus Peronospora parasitica , one obtained from cauliflower ( Brassica oleracea ) and the other from oilseed rape ( B. napus ) was assessed in their respective hosts of origin, and also in the alternative combination. Both isolates were capable of infecting either host, but there were marked contrasts in the time course and extent of mycelial development, the amounts of associated host-cell necrosis, and eventual intensity of sporulation. Oilseed rape, which was partially resistant to the isolate from cauliflower, exhibited extensive necrosis of mesophyll cells, in conjunction with reduced mycelial development, and delayed and reduced sporulation by the pathogen. The isolate from oilseed rape was virulent on both host species. Pathogenesis in the susceptible combinations was accompanied by large increases in electrolyte leakage, and increased activity of the enzymes β-glucosidase, ribonuclease, and peroxidase. Effects on chlorophyll content were variable and activities of acid phosphatase and acid phosphodiesterase were unaffected. Electrophoretic analyses of extracts from fungal sporangia and infected seedlings indicated that the large increases in β-glucosidase were of pathogen origin, while evidence from inhibitor studies suggested that enhanced ribonuclease activity was due to a new post-infectional form of the enzyme. The significance of these findings is discussed in relation to pathogenesis and host resistance mechanisms.     

Variation in aggressiveness of Plasmopara halstedii (sunflower downy mildew)

Journal of Plant Diseases and Protection - Variability in aggressiveness was studied in seven Plasmopara halstedii (sunflower downy mildew) parental isolates of races 100, 300, 304, 314, 704, 710...     

Variation for virulence among Scandinavian isolates of Peronospora viciae f.sp. pisi (pea downy mildew) and responses of pea genotypes

Variation for virulence among Scandinavian isolates of Peronospora viciae f.sp. pisi (downy mildew) on different pea genotypes was investigated. Variation for virulence was found within and between mass-conidial isolates which were originally obtained from oospore populations. Virulence towards pea cultivars that have not been commerically cultivated in Scandinavia was observed. The specific resistance of the pea cultivars Puget, Cobri, Gastro, Starcovert and Starnain was confirmed. One pea breeding line showed a stable partial resistance to different isolates of the fungus and it was also highly resistant to race 8'from The Netherlands which is considered to be virulent on most pea genotypes carrying known specific resistance factors.     

Seed transmission of maize downy mildew (Peronosclerospora sorghi) in Nigeria

In an area of Nigeria where downy mildew of maize is present, histological assessment of maize seed revealed the presence of mycelium and oospores of Peronosclerospora sorghi in the kernels. Seed transmission of downy mildew of maize was demonstrated when grain purchased at local markets gave mean seedling infection rates of 12·3% (untreated seeds) and 10·0% (in metalaxyl-treated seeds) within 7 days of emergence, after storage in a desiccator for 30 days. When untreated seeds taken from nubbin ears of systemically infected plants from four states in southern Nigeria were planted at 9 days (17–22% moisture content) and 27 days (9–22% moisture content) after harvest, 20·0% infected seedlings resulted in both trials. Seeds from Borno state in northern Nigeria had 26·6% systemic seedling infection after 9 months of storage at 11% moisture content. When seeds harvested from maize plants inoculated with P. sorghi through silks were examined histologically, hyphae of P. sorghi were observed mostly in the scutellum of the embryo. Transmission of disease to seedlings was observed when the silk-inoculated seeds (9% moisture content) were planted in pots in a greenhouse; however, no disease transmission was observed when such seeds were planted in the field. The epidemiological significance of seed transmission is discussed with particular reference to survival of inoculum and development of epidemics. Also noteworthy is the overall significance of seed transmission in Nigeria, where the major source of seed is that saved by farmers from their grain crop, occasionally supplemented by seed bought from the local market.     

应用real-time PCR定量检测小麦条锈菌潜伏侵染量方法的建立

 小麦条锈病是我国小麦主要病害之一。快速、及时地诊断与定量监测处于潜育状态下的病叶,对准确估计越冬、越夏后的病情,制定正确的防治方案具有重要的意义。根据小麦条锈菌Puccinia striiformis的β-tubilin基因序列设计对该病原菌的种具有特异性的引物betaf/betar,并分别在普通PCR和real-time PCR扩增时对该引物的特异性和灵敏性进行了测定。结果表明该引物对小麦条锈菌特异性高,可稳定扩增出243 bp的目标条带。Real-time PCR的灵敏度为普通PCR的100倍。应用此特异性引物,建立了real-time PCR测定系统,定量测定了条锈菌在小麦叶片接种后组织内的DNA随时间的变化。结果表明,在接种后12 h,可在小麦叶片内检测到条锈菌,且条锈菌在小麦叶片内潜育期间随时间呈指数增长。接种第6 d后叶片内的菌量有明显的增加。建立的小麦条锈菌的real-time PCR早期定量测定方法,为及时、快速监测小麦条锈病在潜育期间的发病规律以及为该病的预测、防治提供依据。     

Quantification of Cylindrocarpon destructans f. sp. panacis in soils by real-time PCR

Ginseng ( Panax quinquefolius ) is an important cash crop in various regions of North America, but yields are often reduced by various root pathogens. A quantitative real-time PCR (qPCR) assay for Cylindrocarpon destructans f. sp. panacis (CDP), the cause of a root rot and replant disease which discourages successive cropping of ginseng on the same site, was developed to quantify the levels of this pathogen in soils previously cropped with ginseng. DNA was extracted from 5-g samples of soil. In pasteurized soils which were re-infested with varying levels of the pathogen, qPCR estimates of pathogen DNA were significantly correlated with disease severity ( r  = 0·494) and with counts of colony-forming units ( r  = 0·620) obtained with an agar medium. In several naturally infested field soils, qPCR estimates of CDP-DNA concentration were significantly correlated with disease severity ( r  = 0·765) and these concentrations were estimated to range from 0 to 1·48 ng g⁻¹ dried soil. A principal components analysis did not show any strong relationships between soil chemistry factors and the concentration of pathogen DNA. The approach outlined here allows the quantification of current populations of CDP in soil many years after ginseng cultivation and the prediction of disease severity in future crops. The method should be generally applicable to root diseases of many crops.     

应用real-time PCR 定量检测田间小麦条锈菌潜伏侵染的研究

 为快速、及时地定量检测处于潜育状态下的小麦条锈菌菌量,本研究于2009－2010年实施田间试验,在北京和甘肃各设3个试验小区,将小区分割成1 m×1 m或1.5 m×1.5 m的小格。初春刚返青叶片出现症状前,在每小格内采集30片叶,提取叶片基因组DNA,应用已建立的real-time PCR定量测定方法,计算每个样本的分子病情指数(molecular-detected disease index, MDI）。在本试验的天气条件和试验设计下,采样10 d后,调查田间标记采样点的发病情况,计算病情指数DI,比较MDI和DI的关系,建立回归模型。结果表明：甘肃甘谷3个试验小区的MDI和DI存在极显著的相关性（R₂¹=0.813 6,R₂²=0.884 5,R₂³=0.592 8）;北京上庄3个试验小区的MDI和DI也存在极显著的相关性（R₂¹=0.742 3,R₂²=0.578 4,R₂³=0.858 4）。本研究证明了应用分子生物学方法可估测小麦条锈病潜育侵染程度的可能性,为建立分子定量测定田间潜育侵染量系统提供了可靠依据。     

Role of oospores as primary inoculum for epidemics of downy mildew caused by Peronospora arborescens in opium poppy crops in Spain

This study investigated the role of Peronospora arborescens oospores as primary inoculum for downy mildew of opium poppy and infection types that they may give rise to in Spain using an integrative experimental approach that combined pathogenicity tests in growth chambers and field microplots, together with molecular detection of P. arborescens infection by specific nested-PCR assays. The results demonstrated that oospores in infested soil or leaf debris were effective inoculum for ingress of the pathogen through underground plant tissues early in poppy seedling growth. This gave rise to systemic infections that reproduced the stunting, chlorotic syndrome frequently observed in affected plants in commercial fields. Additionally, infection of underground tissues of older plants by oospore inoculum could remain asymptomatic. Results also suggested that sporangia formed on infected plants are effective in producing secondary local infections that later may become systemic and either symptomatic or asymptomatic. Finally, and more importantly, those delayed symptomatic or asymptomatic systemic infections, as well as secondary local infections of capsules, can give rise to infected seeds. This research on the biology of P. arborescens on poppy plants and epidemiology of downy mildew may help to develop knowledge-based disease-management strategies of use in the protection of yields of opium poppy crops in Spain and elsewhere.