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1.
Recently, flavonoids were shown to modulate the outcome of clubroot development in Arabidopsis thaliana after infection with the obligate biotrophic pathogen Plasmodiophora brassicae. Therefore, the development of clubroot disease was investigated in Arabidopsis after treatment with prohexadione‐calcium (ProCa), an inhibitor of ascorbic acid/2‐oxoglutaric acid‐dependent dioxygenases such as flavanone‐3‐hydroxylase. The treatment resulted in a reduction of the flavonols quercetin and kaempferol in clubroots, whereas the precursor naringenin highly accumulated. The root system of ProCa‐treated plants was better developed although galls were still visible. Thus, ProCa treatment resulted in reduced gall size. Flavonoids are thought to inhibit polar auxin transport by modulating auxin efflux carriers. It was investigated whether the auxin response might change as a consequence of the accumulation of naringenin in ProCa‐treated plants. In the areas of gall development an auxin response was indicated by the auxin‐responsive promoter DR5 coupled to the reporter β‐glucuronidase (GUS), whereas very little staining was found in healthy root parts. No differences in GUS activity were found between P. brassicae‐infected and ProCa‐treated plants, and plants only infected with P. brassicae, indicating that the effect of ProCa treatment on clubroot reduction is not via changes in auxin responses. As ProCa is also an inhibitor of late steps in gibberellin biosynthesis, a specific gibberellin biosynthesis inhibitor, chlormequatchloride (CCC), was tested on club development. However, CCC did not reduce disease symptoms, indicating that the observed reduced gall development was not because of gibberellin biosynthesis inhibition by ProCa.  相似文献   

2.
The clubroot pathogen Plasmodiophora brassicae is an obligate biotrophic protist that lives in close relationship with its host cell. The roots of the host plants are colonized and the plant growth is altered upon infection. While shoots can be stunted and show wilt symptoms after longer infection periods, the root system is converted to a tumorous root tissue, called ‘clubroot’, by alterations of the plant growth promoting hormones auxin, cytokinin and brassinosteroid. Because the life cycle occurs largely within the host cells, this leads to dramatic changes in host root morphology and anatomy. Thus, the identification of the respective protist structures in the host tissue by microscopy is challenging. Different staining methods as well as fluorescence and electron microscopy of thin sections can reveal specific life stages of P. brassicae and can yield additional information on the changes in the host tissues concerning, for example, cell wall properties. In addition, promoter–reporter fusions, immunostaining methods and in situ hybridization techniques can be used to gain additional information on the changes in the host roots.  相似文献   

3.
Root-knot nematodes (RKN) are obligate endoparasites that severely damage the host root system. Nutrient and water uptake are substantially reduced in infested plants, resulting into altered physiological processes and reduced plant growth. The effect of nematode infestation on the morphological changes of roots and subsequent physiological plant responses of infested tomatoes with the RKN Meloidogyne ethiopica was studied in a pot experiment. Plants were infested with two inoculum densities (10 or 50 eggs per cm3 substrate) and its effect was evaluated 74 and 102 days post inoculation (DPI). Morphological changes and root growth was determined by analysing scanned images of the whole root system. Nematode infestation reduced the portion of fine roots and increased that of coarse roots due to gall formation. Fine roots of non-infested control plants represented around 51% of the area of the whole root system at 74 and 102 DPI. In comparison to controls, plants inoculated with low and high nematode density had 2.1 and 3.2-times lower surface area of fine roots at 102 DPI. Root analyses revealed that plants had a very limited ability to mitigate the effects of the root-knot nematodes infestation by altering root growth. Root galls had a major influence on the hydraulic conductivity of the root system, which was significantly reduced. The low leaf water potential of infested plants coincided with decreased stomatal conductivity, transpiration and photosynthesis. The latter two were reduced by 60–70% when compared to non-infested control plants.  相似文献   

4.
Fusarium wilt, caused by Fusarium oxysporum f. sp. cucumerinum (FO), is one of the major diseases in cucumber (Cucumis sativus) production. Root and foliar applications of 24-epibrassinolide (EBL), an immobile phytohormone with antistress activity, were evaluated for their effects on the incidence of Fusarium wilt and changes in the microbial population and community in roots of cucumber plants. EBL pre-treatment to either roots or shoots significantly reduced disease severity followed by an improved plant growth regardless of the treatment methods applied. EBL applications decreased the Fusarium population on root surfaces and in nutrient solution, but increased the population of fungi and actinobacteria on root surfaces. PCR-DGGE analysis showed that FO-inoculation had significant effects on the bacterial community on root surfaces as expressed by a decreased diversity index and evenness index, but EBL applications alleviated these changes. Moreover, several kinds of decomposing bacteria and growth-promoting bacteria were identified from root surfaces of FO-inoculated plants and EBL-pre-treated plants, respectively. Overall, these results show that the microbial community on root surfaces was affected by a complex interaction between phytohormone-induced resistance and plant pathogens.  相似文献   

5.
Pantoea agglomerans pvs. gypsophilae and betae are related gall-forming bacteria. While P. agglomerans pv. beta initiates gall formation on both beet and gypsophila, the gypsophila pathovar causes gall formation only on gypsophila. PthG is a type III effector determining host range of these pathogens, initiating the hypersensitivity response in beet, but is a virulence factor in gypsophila. The role of PthG and its mode of action in pathogenicity remain unclear. Transgenic Nicotiana tabacum plants expressing pthG were created. PthG over-expression was often lethal, and surviving pthG-bearing lines showed morphological and developmental abnormalities such as leaf deformation and abnormal vascular branching, dwarf stature, loss of apical dominance, seedling apical meristem loss, rapid germination, reduced fertility, plants which cease growth for several weeks later producing a new lateral shoot, and loss of endophyte resistance (bearing unusual saprophyte populations). Some transformants required light for seed germination and showed rapid seedling greening. In in vitro assays PthG expression modified responses to auxin and cytokinin, inhibiting root and shoot production but not callus formation. In vitro differentiation responses to light were modified by PthG expression. This effector may interfere in the plant auxin signalling pathways resulting in higher observed auxin and ethylene levels, and subsequent blockage of root and shoot development. Apparently PthG tunes the host response to high hormone levels, changing the developmental response. Since shoot and root development are delayed, we hypothesize that callus/gall formation is supported by this activity. However, interference by PthG with hormone and light signalling does not explain all the responses observed in pthG-bearing lines.  相似文献   

6.
To investigate the susceptibility of hairy root lines of Brassica species to Plasmodiophora brassicae, hairy roots were induced in a number of Brassica species with Agrobacterium rhizogenes. Turnip hairy root was highly susceptible to P. brassicae; infection rates were high and large galls formed. In contrast, the rates of root hair infection and gall formation on intact Brassica plants did not differ significantly from the control. To induce resting spore formation, turnip hairy roots were incubated at 15°, 20°, or 25°C after 3 weeks of incubation at 25°C. The number and fresh mass of the galls per hairy root were higher and formation of resting spores was greatest after a 7-week incubation at 20°C. To subculture P. brassicae using turnip hairy root, turnip hairy roots were reinoculated with resting spores and gall with resting spores then formed on the hairy roots. In this way, P. brassicae using hairy roots could be subcultured in vitro two or three times on three single-spore isolates of P. brassicae. This is the first report of in vitro subculture of P. brassicae using hairy root.  相似文献   

7.
Agrobacterium tumefaciens (AT) is the causal agent of crown gall, a major problem in the family Rosaceae and particularly for Prunus spp. Crown gall symptoms result from the bacterial infection of the cells damaged mechanically at the collar or by root parasitic nematodes. Myrobalan plum (P. cerasifera) is susceptible to AT and is not a host for the root-knot nematode (RKN), M. hapla. Some clones of this plum carry single Ma resistance genes that control M. arenaria, M. incognita and M. javanica. The four above mentioned RKN and Myrobalan progenies segregating for Ma were used in experiments aimed at obtaining a better knowledge of the interaction between AT and RKN in relation to the RKN resistance genes. Prunus rooted cuttings, naturally infected with the bacterium were repotted, grown and inoculated individually with RKN. In a first experiment, Prunus plants were (i) either inoculated with 10,000 juveniles (J2s) of M. arenaria to provide a short inoculum pressure (SIP) or (ii) inoculated by association with one M. arenaria-galled tomato root system that produced a high and durable inoculum pressure of the same nematode species. Four months after RKN inoculation, plants were rated for nematode and bacterial root galling symptoms. RKN and AT galls were more numerous and more homogenous under DIP than under SIP. Nevertheless, for both inoculum regimes, AT galls were present in the RKN-susceptible clones (= carrying none of the Ma genes) and absent in the RKN-resistant clones. Subsequent experiments, conducted under DIP with M. arenaria, M. incognita, M. javanica and M. hapla, also showed, for the three first species, the presence of AT galls only in RKN-susceptible clones whereas Prunus plants inoculated with M. hapla and nematode-free controls were free of AT galls. Consequently RKN act as a wound agent in the AT infection process of Myrobalan plum only when the plant develops a compatible reaction (i.e. when it lacks the Ma resistance genes). Considering that J2s do penetrate the roots of resistant plants, the absence of crown gall symptoms on this material even under durable inoculum pressure strengthens the hypothesis that this nematode stage has a very weak effect on plant cells during the infection process. This is the first evidence of the protective effect of a RKN resistance gene against the expression of root crown gall consecutive to RKN infection. The protective effect of Ma and presumably of other RKN resistance genes against AT is a strong argument for their introgression into Prunus and other Rosaceae or bacterium-susceptible crops.  相似文献   

8.
Fusarium species are soil-borne fungal pathogens that produce a variety of disease symptoms when attacking crop plants. The mode of root colonization of Eucalyptus viminalis seedlings by a pathogenic F. oxyporum strain (Foeu1) at the ultrastructural level and changes in cell wall pectin during host pathogen interactions are described. Root systems of E. viminalis plants were inoculated with F. oxysporum in an in vitro model system. Hyphae of F. oxysporum adhered to the outer epidermal cell walls through fibrillar material, and after penetration they spread into the internal tissues. They developed intercellularly and intracellularly in the root cortex and invaded vascular tissues. Papillae were induced, and the host plasma membrane ruptured in colonized cells, causing rapid host tissue and cell damage. Changes in distribution and occurrence of nonesterified and methyl-esterified pectins were evaluated after root colonization by F. oxysporum using two monoclonal antibodies, JIM 5 and JIM 7, respectively. Nonesterified pectin in control roots was mainly localized in the epidermal cell walls and middle lamellae in parenchymal cortex, whereas methyl-esterified pectin accumulated more in primary cell walls of the cortex and phloem. Decreases in immunodetected nonesterified and methyl-esterified pectins were associated with extensive plant tissue degradation after root colonization by the pathogenic fungus.  相似文献   

9.
Since 2003, Torenia fournieri plants grown for experimental purposes were repeatedly infected by powdery mildew in a laboratory in Hungary. Based on morphological characteristics, the pathogen belonged to the mitosporic genus Oidium subgen. Reticuloidium, the anamorph stage of Golovinomyces. The rDNA ITS sequence was identical to that of two other powdery mildew fungi, infecting Arabidopsis and Veronica, respectively, in different parts of the world. According to a previous phylogenetic analysis of ITS and 28S rDNA sequences, those two powdery mildews belong to a recently evolved group of Golovinomyces characterized by multiple host range expansions during their evolution. Both the ITS sequence and the morphological data indicate that the powdery mildew anamorph infecting Torenia also belongs to this group. It is likely that the powdery mildew infections of the experimental T. fournieri plants, native to south-east Asia, were the result of a very recent host range expansion of a polyphagous Golovinomyces because (i) T. fournieri is absent from our region, except as an experimental plant grown in the laboratory, (ii) the powdery mildew fungus infecting this exotic plant belongs to a group of Golovinomyces where host range expansion is a frequent evolutionary scenario, (iii) cross-inoculation tests showed that this pathogen is also able to infect other plant species, notably A. thaliana and tobacco, and (iv) no Golovinomyces species are known to infect T. fournieri anywhere in the world. Although host range expansion has often been proposed as a common evolutionary process in the Erysiphales, and also in other biotrophic plant pathogens, this has not been clearly demonstrated in any case studies so far. To our knowledge, this is the first convincing case of a host range expansion event in the Erysiphales.  相似文献   

10.
Nonpathogenic isolates of Fusarium oxysporum can be successful antagonists of pathogenic forms of the same fungal species that commonly attacks crop plants. The characteristics that distinguish nonpathogenic from pathogenic forms are not well understood. In this study, the mode of root colonization of Eucalyptus viminalis seedlings by a nonpathogenic F. oxysporum strain is described at the ultrastructural level. Root systems of E. viminalis plants were inoculated with nonpathogenic F. oxysporum strain Fo47 in an in vitro model system. Changes in the occurrence of nonesterified and methyl-esterified pectins in colonized E. viminalis roots were evaluated by in situ immunolabeling using two monoclonal antibodies, JIM 5 and JIM 7. Modes of penetration and root colonization patterns in E. viminalis seedlings by the nonpathogenic fungus were similar to those described for pathogenic forms of F. oxysporum. However, root interactions differed in that the nonpathogenic fungus did not induce host tissue damage. No papilla-like appositions were observed in host cells in response to invading hyphae, which did not disrupt the host plasma membrane in many cases, suggesting that a biotrophic relationship was established. Root colonization by the nonpathogenic strain did not induce alteration in JIM 7 labeling of methyl-esterified pectin in E. viminalis cell walls, whereas nonesterified pectin was detected to a significantly greater extent in cell walls of roots colonized by the fungus. Pectin components decreased slightly only at points of hyphal contact with host cells. Because nonpathogenic strains utilize pectin in pure culture, host control over enzyme activity or production by the fungi may at least partly explain their compatible interactions with host tissues.  相似文献   

11.
Crown gall disease of tobacco was found in Iwate Prefecture, Japan in 1995. Ten bacterial isolates, obtained from the galls of tobacco, were identified as Agrobacterium tumefaciens (Smith and Townsend 1907) Conn 1942 biovar 1 based on their ability to induce galls on the 14 tested plants, including tobacco after needle-prick inoculation, and on 12 cultural, physiological, and biological characteristics. The growth of the causal organism was not inhibited in vitro by agrocin of A. radiobacter strain K84. This report is the first on the natural occurrence of crown gall caused by A. tumefaciens on tobacco plants.  相似文献   

12.
Oomycetes contain some of the economically most important pathogens of flowering plants. Most have a rather narrow host range, often being restricted to single host species. In downy mildews and other obligate biotrophic plant parasites, like powdery mildews and rusts, delimitating species on grounds of morphological characteristics is often hardly possible and thus often based on only subtle differences. This has led to the widespread application of a broad species concept for these organisms. Consequently, despite the fact that morphological differences were reported for Pseudoperonospora cubensis from different host species, the corresponding new pathogen species were not accepted as being independent, and the host range of Pseudoperonospora cubensis is reported to encompass more than 50 host species in the Cucurbitaceae in temperate to tropical climates. However, recent studies have reported narrow host ranges for other downy mildew genera and advocated a narrow species concept. Here, we report successful colonisation of five different tribes of the Cucurbitaceae by a strain of Pseudoperonospora cubensis and demonstrate that the host matrix has a major impact on the morphology of the pathogen. On the basis of five morphological criteria significant differences could be found for all hosts. These differences were more pronounced in phylogenetically unrelated than in related hosts. Our results provide evidence for a broad host range of Pseudoperonospora cubensis and demonstrate that species delimitation based on morphological characters is not feasible in Pseudoperonospora on Cucurbitaceae. Also in other biotrophic plant pathogens, the situation could be similar, thus necessitating thorough morphological, molecular phylogenetic and cross inoculation experiments for species recognition.  相似文献   

13.
A nonpathogenic strain of Agrobacterium vitis VAR03-1 was tested as a biological control agent against crown gall of grapevine (Vitis vinifera L.). A mixture of the nonpathogenic strain VAR03-1 and a tumorigenic strain G-Ag-27 of A. vitis at cell ratios of 1 : 1, 3 : 1, 9 : 1, and 99 : 1 significantly inhibited gall formation and size on stems of tomato (Lycopersicon esculentum Mill.). Strain VAR03-1 also inhibited gall formation on stems of both tomato and grapevine at a 1 : 1 cell ratio with several tumorigenic A. vitis strains isolated from different fields of grapevine in Japan. In biological control tests, when roots of grapevine and tomato seedlings were soaked in a cell suspension of strain VAR03-1 for 24 h before a 1-h soaking in a cell suspension of the pathogen and subsequent planting in pots of infested soil, strain VAR03-1 significantly reduced the incidence of gall formation on both plants.  相似文献   

14.
Ascospores, discharged naturally from apothecia growing on rachis debris, were used as inoculum to examine the invasion of ash tissues by Hymenoscyphus fraxineus in order to understand the critical, but poorly understood, early interactions between host and pathogen. Methods were developed to collect ascospores for controlled infection assays on detached leaves, petioles and stem internode tissues. Light microscopy, using plasmolytic techniques, allowed the invasion of living plant cells to be observed. Ascospores were readily available from late May to September. On the plant surface, most spores differentiated directly to form appressoria without germ‐tube growth. Direct penetration was followed by a significant period of biotrophic fungal growth before lesions developed. Following the formation of a vesicle‐like structure after penetration, bulbous and elongated intracellular hyphae were produced in living plant cells. The use of ascospore inoculum, rather than mycelia, will allow natural and rapid screening of ash genotypes for resistance to the devastating dieback disease. The identification of the biotrophic phase of infection suggests that host range is controlled by effector‐triggered immunity.  相似文献   

15.
The obligate biotrophic nature of rust fungi calls for an in planta selection scheme as a means of developing a rust transformation technology. We show that the fungicides benomyl (used as its formulated product benlate) and carboxin suppress morphogenesis of the rust fungus Uromyces fabae in vitro and disease in planta, the latter without affecting the health of the host. The limits of their applicability were determined regarding concentration, method of application and optimal time intervals of treatment. Besides procedures for selection, a stable transformation system will also need to include genetic markers allowing to enrich for transformed cells within a large background of untransformed cells. Since the molecular targets of benlate and carboxin had been identified as -tubulin and succinate dehydrogenase, respectively, the corresponding genes (Uf-TBBIand Uf-SUCDHI) were cloned and characterized. Molecular phylogenies demonstrate that both are typical homologs to those of other Basidiomycota. RT-PCR analysis confirmed that both genes are constitutively expressed in all developmental stages of the mitotic uredospore multiplication cycle. Since homologs of Uf-TBB1and Uf-SUCDH1 have been successfully used as selection markers in other fungal systems, they provide valuable tools to develop additional corner stones of a stable transformation system for rust fungi.  相似文献   

16.
The differential interactions of V. longisporum (VL) and V. dahliae (VD) on the root surface and in the root and shoot vascular system of Brassica napus were studied by confocal laser scanning microscopy (CLSM), using GFP tagging and conventional fluorescence dyes, acid fuchsin and acridin orange. VL and VD transformants expressing sGFP were generated by Agrobacterium-mediated transformation. GFP signals were less homogenous and GFP tagging performed less satisfactory than the conventional fluorescence staining when both were studied with CLSM. Interactions of both pathogens were largely restricted to the root hair zone. At 24 h post-inoculation (hpi), hyphae of VL and VD were found intensely interwoven with the root hairs. Hyphae of VL followed the root hairs towards the root surface. At 36 hpi, VL hyphae started to cover the roots with a hyphal net strictly following the grooves of the junctions of the epidermal cells. VL started to penetrate the root epidermal cells without any conspicuous infection structures. Subsequently, hyphae grew intracellularly and intercellularly through the root cortex towards the central cylinder, without inducing any visible plant responses. Colonisation of the xylem vessels in the shoot with VL was restricted to individual vessels entirely filled with mycelium and conidia, while adjacent vessels remained completely unaffected. This may explain why no wilt symptoms occur in B. napus infected with VL. Elevated amounts of fungal DNA were detectable in the hypocotyls 14 days post-inoculation (dpi) and in the leaves 35 dpi. Root penetration was also observed for VD, however, with no directed root surface growth and mainly an intercellular invasion of the root tissue. In contrast to VL, VD started ample formation of conidia on the roots, and was unable to spread systemically into the shoots. VD did not form microsclerotia in the root tissue as widely observed for VL. This study confirms that VD is non-pathogenic on B. napus and demonstrates that non-host resistance against this fungus materializes in restriction of systemic spread rather than inhibition of penetration.  相似文献   

17.
A wilt disease of the model legume Lotus japonicus was observed in a greenhouse in Tokyo, Japan in May 2004. Roots of diseased plants were rotted and dark brown with lesions spreading to lower stems and leaves, resulting in rapid plant death. The causal agent was identified as Fusarium solani based on the morphology. Sequence analysis of rDNA supported the identification. Inoculation of roots of healthy plants with conidia reproduced characteristic disease symptoms, and F. solani was reisolated from lesions, satisfying Koch’s postulates. The isolate also caused chlorotic to necrotic lesions on leaves of healthy plants after wound-inoculation. Infection by F. solani of leaves of L. japonicus was confirmed histologically. Mycelia were observed in the intercellular spaces of parenchymatous tissues in the lesion area and the surrounding tissues. This is the first report of fungal disease on L. japonicus satisfying Koch’s postulates. We named it “Fusarium root rot of L. japonicus” as a new disease. The compatibility of L. japonicus and F. solani is expected to form a novel pathosystem for studying interactions between legumes and fungal pathogens. The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under accession numbers AB258993 and AB258994.  相似文献   

18.
Lettuce downy mildew caused by Bremia lactucae has long been a model for understanding biotrophic oomycete–plant interactions. Initial research involved physiological and cytological studies that have been reviewed earlier. This review provides an overview of the genetic and molecular analyses that have occurred in the past 25 years as well as perspectives on future directions. The interaction between B. lactucae and lettuce (Lactuca sativa) is determined by an extensively characterized gene-for-gene relationship. Resistance genes have been cloned from L. sativa that encode proteins similar to resistance proteins isolated from other plant species. Avirulence genes have yet to be cloned from B. lactucae, although candidate sequences have been identified on the basis of motifs present in secreted avirulence proteins characterized from other oomycetes. Bremia lactucae has a minimum of 7 or 8 chromosome pairs ranging in size from 3 to at least 8 Mb and a set of linear polymorphic molecules that range in size between 0.3 and 1.6 Mb and are inherited in a non-Mendelian manner. Several methods indicated the genome size of B. lactucae to be ca. 50 Mb, although this is probably an underestimate, comprising approximately equal fractions of highly repeated sequences, intermediate repeats, and low-copy sequences. The genome of B. lactucae still awaits sequencing. To date, several EST libraries have been sequenced to provide an incomplete view of the gene space. Bremia lactucae has yet to be transformed, but regulatory sequences from it form components of transformation vectors used for other oomycetes. Molecular technology has now advanced to the point where rapid progress is likely in determining the molecular basis of specificity, mating type, and fungicide insensitivity.  相似文献   

19.
Investigations on the efficacy of various methods of managing root-knot nematodes in microplots and under field conditions revealed that soil solarization, Furadan 5G and Tagetes erecta applied separately or in combination with other control methods, were the most effective in reducing the numbers of three root-knot nematodes, and root gall and egg mass indices. These management methods also resulted in significant increases (P0.05) in number of pods per plant, number of seeds per pod and 100-seed weight, and increased seed yield by up to 96.7 percent. Farmyard manure and Crotalaria ochroleuca were the least effective treatments. The use of T. erecta was the most economical root-knot nematode control method.  相似文献   

20.
The polyphagous obligate parasites Meloidogyne spp. devastate a wide range of crop plants including bananas and plantains. Their infestations impact agriculture worldwide. Therefore, an effective combating regime against this nematode species and an in-depth understanding of plant-nematode interaction are essential. Early detection of infection by visual inspection is not possible. This hampers early control strategy efforts and makes in-depth research of the early infection and plant defence unfeasible. A simple and robust in planta PCR-based nematode detection method is described here as the first crucial step. This PCR-based detection assay exploits the existence of the Internal Transcribed Spacer 1 (ITS 1) region of the ribosomal DNA (rDNA) gene family in the nematodes for early detection of nematode penetration into the roots. The results demonstrate that this detection assay is suitable to serve as a molecular screening tool for plant root diagnostic purposes.  相似文献   

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