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1.
The potato brown rot, caused by the bacterium Ralstonia solanacearum biovar 2/race 3 (potato race) was classified as quarantine pathogen by the European Union (EU) due to the risks it would pose for the cultivation of potatoes and tomatoes. Quarantine regulations stipulate control surveys and tests on potatoes used as seed, for food as well as industrial purposes and in surface water to check for contamination with R. solanacearum. Toward this end, the Institute for Plant Protection of the Bavarian State Research Center for Agriculture has been conducting an intensive survey for the presence of the potato brown rot pathogen in Bavarian rivers since 1997. An important component of this monitoring is the testing of potential weed hosts growing near river banks. Every year, from June to September, water- and plant samples have been collected from rivers in potato cultivation areas and near potato processing plants. Since the start of the survey, a total of 70 rivers has been tested. The presence of the pathogen was checked by immunofluorescence antibody staining (IFAS) and polymerase chain reaction (PCR). Positive IFAS or PCR tests were checked again by biotests and pathogenicity tests on eggplant (Solanum melongena). So far R. solanacearum could be found in nine Bavarian rivers. Concerning wild plants growing near the river banks the bacterium could be isolated only from roots of the bittersweet nightshade (Solanum dulcamara). However, from stinging nettle (Urtica dioica), nodding beggartick (Bidens cernua), black nightshade (Solanum nigrum), great yellowcress (Rorippa amphibia) and gipsywort (Lycopus europaeus) R. solanacearum could not be isolated. In contaminated rivers, with S. dulcamara growing on the river banks, R. solanacearum could be detected repetitively for several years. In 1997 it was found that rivers can be contaminated by the pathogen, if insufficiently treated sewage from potato processing plants is released into waterways. Potato cultivation is threatened by surface water only, if contaminated water is used to irrigate potato fields. Consequently, surface water should never be used for irrigation of potato fields or application of plant protection products.  相似文献   

2.
Ralstonia solanacearum is a known bacterial pathogen of eucalypt and potato plants in Africa. A survey was undertaken to detect this pathogen in eucalypt plantations in South Africa, the Democratic Republic of Congo, and Uganda. Numerous bacterial strains were isolated from trees with symptoms typical of bacterial wilt, but only seven were positively identified as R. solanacearum. A polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique, based on the hrp (hypersensitive response and pathogenicity) gene region was used to determine and group the biovars of these R. solanacearum strains. The eucalypt isolates and one potato isolate formed a biovar 3 cluster, whereas the two other potato isolates formed a cluster that corresponded to biovar 2. Amplified fragment length polymorphism (AFLP) analysis confirmed these clusters. Therefore, PCR-RFLP can be used as a reliable diagnostic technique to enable researchers to rapidly identify the pathogen.  相似文献   

3.
Bacillus amyloliquefaciens IUMC7 and its culture supernatant, which we previously found to suppress Fusarium wilt, were found here to have antimicrobial activity against Ralstonia solanacearum. In in vivo experiments, mushroom compost inoculated with IUMC7 significantly reduced disease severity in tomato plants over that in control soils. The R. solanacearum population decreased in soil inoculated with IUMC7. A TLC–bioautography assay showed that one of the antimicrobial substances produced by IUMC7 is likely an iturin-like lipopeptide. These results suggest that these antimicrobial compounds are responsible for disease suppression and that mushroom compost containing IUMC7 has potential as a biocontrol product.  相似文献   

4.
Ralstonia solanacearum, the devastating causal agent of potato bacterial wilt, is a soil-borne bacterium that can survive in the soil for a long time. The development of sensitive on-field detection methods for this pathogen is highly desirable due to its widespread host range and distribution. A novel nanobiosensor was thus developed to detect unamplified genomic DNA of R. solanacearum in farm soil. Gold nanoparticles functionalized with single-stranded oligonucleotides served as a probe to detect R. solanacearum genomic DNA. The advantages of this strategy include rapidity, facile usage and being a visual colorimetric method.  相似文献   

5.
Ralstonia solanacearum is the causal organism of bacterial wilt of more than 200 species representing 50 families of plants in tropical, subtropical, and warm temperate regions in the world. Traditionally classified into five races based on differences in host range, R. solanacearum has also been grouped into six biovars on the basis of biochemical properties. With recent developments in molecular biology, various DNA-based analyses have been introduced and used to confirm that this binary system does not completely represent the diversity within R. solanacearum strains. Therefore, a new hierarchical classification scheme has been suggested, which defines R. solanacearum as a species complex and reorganized the concept of the species as a monophyletic cluster according to a phylogenetic analysis based on genomic sequence data. Here we discuss the current bacterial wilt situation and genetic relationships based on the recent classification system of Japanese R. solanacearum strains as well as worldwide strains. We also review the genetic, biochemical, and pathological characteristics of R. solanacearum strains, in particular, those affecting potato and Zingiberaceae plants as distinctly important pathogens in relation to continuously problematic and recent emergent diseases in Japan.  相似文献   

6.
This study investigated the effect of calcium nutrition on tomato bacterial wilt caused by Ralstonia solanacearum and the regulation of resistance mechanisms. Plants cultured in nutrient solution with calcium concentrations of 0.5, 5.0, and 25.0 mM, were inoculated with R. solanacearum by the root dip method. Severity of disease development, Ca concentration in tomato root and shoot tissues, hydrogen peroxide (H2O2) concentration, peroxidase (POD, EC 1.11.1.7) and polyphenol oxidase (PPO, EC 1.10.3.2) in tomato leaves were analyzed. Disease severities of low, medium and high Ca treatments were 100 %, 77.1 % and 56.8 % respectively. Plant growth in high Ca treatment was significantly better than those in low Ca treatment in height, stem diameter and biomass. Tomato plants absorbed significantly more Ca in roots and shoots as the level of Ca in the nutrient solution increased. In addition, H2O2 level in high Ca treatment rose faster and reached a higher peak with 10.86 μM gFW?1(31.32 % greater than medium Ca plants). The activities of POD and PPO also have a greater increase in high Ca treatment with 99.09 U gFW?1 and 107.24 U gFW?1 compared to 40.70 U gFW?1 and 77.45 U gFW?1 in low Ca treatment. A negative correlation was found between Ca concentration, level of H2O2, POD, PPO in tomato, and disease severity, indicating that they played an important role in resistance of tomato to this disease. These results suggested that Ca was involved in the regulation of H2O2 concentration, and activity of POD and PPO in tomato.  相似文献   

7.
Leaves of powdery mildew-susceptible barley (Hordeum vulgare cv. Ingrid) and related near-isogenic lines bearing various resistance genes (Mla12, Mlg or mlo5) were inoculated with Blumeria graminis f. sp. hordei race A6. Fungal attack induced several-fold increases in ethylene emission and electrolyte leakage in leaves of susceptible Ingrid beginning 3 days after inoculation. Activities of peroxidase, superoxide dismutase, glutathione S-transferase, ascorbate peroxidase and glutathione reductase enzymes were induced markedly in susceptible leaves 5–7 days after inoculation. Similar, but less pronounced pathogen-induced changes were detected in inoculated leaves of Mla-type resistant plants that show hypersensitive cell death upon inoculation, and, to an even lesser extent, in the Mlg and mlo lines, where no visible symptoms accompanied the incompatible interaction. Glutathione content increased only in susceptible barley 7 days after inoculation. Catalase activity, total ascorbate content and redox state were not influenced by inoculation in any of the genotypes. The activity of dehydroascorbate reductase was significantly reduced 3–5 days after inoculation in the susceptible parental plants and after 5 days in Mla and Mlg lines, while it was stable in the mlo barley. Slightly elevated levels of H2O2 were observed in the inoculated resistant plants. In contrast, H2O2 content decreased in the susceptible line 7 days after pathogen attack. These data indicate that high levels of antioxidants are involved in the compatible interaction of susceptible barley and powdery mildew by protecting the pathogen from oxidative damage.  相似文献   

8.
Pathogenic characters of Japanese potato strains of Ralstonia solanacearum   总被引：1，自引：0，他引：1  
Ralstonia solanacearum (Rs) strains in phylotypes I and IV isolated from potato in Japan were investigated for pathogenicity on potato, tomato, eggplant, Solanum integrifolium, tobacco, groundnut, and pumpkin. The strains were divided into 17 types based on differences in their pathogenicity on the tested plants. Particularly, the pathogenicity of most phylotype I strains on eggplant was distinctly different from that of the phylotype IV strains. When nine potato varieties (included two breeding lines) were inoculated with several Rs strains, phylotype IV strains were highly virulent on the breeding lines that are regarded as resistant to phylotype I strains.  相似文献   

9.
Solanum commersonii is a wild species related to the cultivated potato. Some S. commersonii genotypes have been proven to be resistant to the pathogenic bacteria Ralstonia solanacearum, which causes damage in potato and other economically important crops. Here an expression analysis of the response of a resistant S. commersonii genotype against R. solanacearum was performed using microarrays. The aims of this work were to elucidate the molecular processes involved in the interaction, establish the timing of the response, and contribute to identify genes related to the resistance. The response to the treatment was already initiated at 6 h post-inoculation (hpi) and was established at 24 hpi; during this period, a high number of genes was differentially expressed and several candidate genes for the resistance of S. commersonii to R. solanacearum were identified. At an early stage, the photosynthetic process was highly repressed and several genes encoding proteins related to reactive oxygen species (ROS) production were differentially expressed. The induction of ERF and ACC-oxidase genes related to the ethylene pathway and PR1 related to the salicylic acid pathway suggested the induction of both pathways, and back up the previously reported hemibiotrophic nature of the pathogen. Five genes related to plant defence and observed to be differentially expressed at the first two time points were validated by real time PCR. This work gives a glimpse to the molecular processes involved in S. commersonii resistance and identifies the species as a valuable genetic source for potato breeding against bacterial wilt.  相似文献   

10.
Surveys over three seasons of irrigation, drainage and artesian well water throughout the major potato-growing areas of Egypt indicated that Ralstonia solanacearum bv. 2 race 3 (phylotype II sequevar 1), cause of potato brown rot, was limited to the canals of the traditional potato-growing areas in the Nile Delta region, with positive findings more commonly associated with the network of smaller irrigation canals flowing through potato-growing areas. Pathogen populations in the canals of the Delta (~100–200 cfu l?1) were generally variable throughout the year with presence linked to potato cultivation in the immediate area. The pathogen was not detected in irrigation or drainage water associated with potato cultivation in the newly reclaimed desert areas (designated as Pest-Free Areas, PFAs) or in the main branches of the Nile upstream from these areas. In vitro studies showed that temperature and microbial activity were the main factors affecting survival of the pathogen in canal water. In experiments at temperatures of 4, 15, 28 and 35°C, survival was longest at 15°C and shortest at 35°C. Survival at 4 and 28°C tended to be intermediate between these extremes as was survival when the bacterium was grown at fluctuating temperatures. Aeration, solarisation and pH variation between 4 and 9 appeared to have little effect on survival. Survival in autoclaved or filter-sterilised canal water was longer than in untreated water irrespective of other factors with survival times exceeding 300 days at 15°C in some experiments. Evidence is presented indicating that survival in water-saturated sediment may be longer than in the overlying water suggesting that sediment may provide a protective niche for the pathogen in some circumstances. The maximum survival time in non-sterile Egyptian canal water at high inoculum pressure was estimated to be up to 300 days at optimum temperature for survival (15–30°C) suggesting the potential for long-distance spread in Egyptian surface waters from sources of contamination.  相似文献   

11.
Clavibacter michiganensis subsp. sepedonicus and Ralstonia solanacearum (Smith) Yabuuchi et al. race 3 are the causal agents of ring-rot and brown-rot of potato respectively. These diseases represent a serious threat to potato production in temperate climates. Both bacteria are listed as A2 pests in the EPPO region and as zero-tolerance quarantine organisms in the European Union. All the detection tests developed so far were only focused on the detection of a single pathogen while the absence of both bacteria has to be certified in the seed tubers. We have therefore developed a new multiplex real-time PCR assay to simultaneously detect both bacteria in a single assay. Additionally, the reliability of this molecular diagnostic test has been improved by the simultaneous amplification of an internal control, corresponding to a potato gene co-extracted from the sample. The polyvalence and the specificity of each set of bacterial primers and probes were evaluated on more than 90 bacterial strains. The limit of detection of this triplex real-time protocol was similar to those observed with other molecular protocols previously developed for the individual detection of one of these bacteria. A concordance of 100 % was obtained in a blind test mimicking the routine application of the technology. In conclusion, this new protocol represents a straightforward and convenient method potentially adapted to primary screening of potato tubers.  相似文献   

12.
Bacterial wilt or brown rot is one of the most devastating diseases of potato caused by a bacterium Ralstonia solanacearum (Smith 1986) Yabuuchi et al. (Microbiol Immunol 39:897–904 1995). Traditionally, R. solanacearum is classified into five races (r) on the basis of differences in host range and six biovars (bvs) on the basis of biochemical properties. Recently using molecular methods, R.?solanacearum has been classified into phylotypes based on the intergenic transcribed sequence of the ribosomal RNA genes 16S and 23S and into sequevars based on the endoglucanase gene (egl) sequence. In the present study, 75 bacterial strains, isolated from wilt infected potatoes from various potato growing regions of India, were classified by traditional and molecular methods. The identity of all the strains was confirmed as R. solanacearum as expected single 280-bp fragment resulted in all the strains following PCR amplification using R. solanacearum specific universal primer pair 759/760. Biovar (bv) analysis, based on utilization of disaccharide sugars and hexose alcohols, categorised the 75 strains into bv2 (78.7 %), 2 T (5.3 %), 3 (5.3 %) and 4 (10.7 %). The phylotype specific multiplex PCR assigned 78.7 % strains to phylotype II, 16.0 % to phylotype I and 5.3 % to phylotype IV. Phylogenetic analysis of egl gene sequences clustered all fifty nine phylotype II (bv2) strains with reference strain IPO1609 (IIB-1), all four phylotype IV (bv2T) strains with reference strain MAFF301558 (IV-8), three phylotype I (bv3) strains with reference strain MAFF211479 (I-30) and all eight phylotype I (bv4) and one phylotype I (bv3) strain with reference strain CIP365 (I-45). The study concluded that the Indian potato strains of R. solanacearum belong to three out of four phylotypes namely: the Asian phylotype I, the American phylotype II, and the Indonesian phylotype IV. This is the first study to address the diversity of R. solanacearum from potato in India using phylotype and sequevar scheme. We also report here for the first time the occurrence of phylotype IV sequevar 8 (bv2T) strain of R. solanacearum causing potato bacterial wilt in mid hills of Meghalaya in India.  相似文献   

13.
This paper describes a comparison study of test methods and supports the use of real‐time polymerase chain reaction (PCR) for the detection of Clavibacter michiganensis subsp. sepedonicus and Ralstonia solanacearum in potato tubers in routine testing. These 2 bacteria are quarantine organisms under European Union (EU) regulatory control and testing for (latent) infections of these bacteria in seed potatoes is mandatory. Real‐time PCR tests were performed on 276 routine potato tuber samples, including samples infected with either C. michiganensis subsp. sepedonicus or R. solanacearum, and the performance of these real‐time PCR tests was compared with that of immunofluorescence (IF). Real‐time PCR tests, using different primer sets and extraction and PCR protocols, proved to be sensitive and specific for the detection of C. michiganensis subsp. sepedonicus and R. solanacearum in potato tubers in routine testing, and performed at least as well as IF. Real‐time PCR is a good addition to the detection protocols as laid down in EU regulations (EU Council Directives 2006/56/EC and 2006/63/EC).  相似文献   

14.
During the last decade, a new bacterial disease has impaired the yield of vegetable sweet potato (30–80%) in Taiwan. Infected plants developed stunting, root and stem rot, vascular discoloration and wilting. Ten bacterial isolates that caused the same symptoms in sweet potatoes after inoculation were reisolated and classified as Ralstonia solanacearum phylotype I biovar 4 based on physical and molecular analyses. Moreover, these isolates also caused wilting in convolvulaceous, solanaceaous and cruciferous plants. This report is the first of bacterial wilt of sweet potato caused by R. solanacearum in Taiwan.  相似文献   

15.
Members of the Ralstonia solanacearum species complex (RSSC), causing potato bacterial wilt or brown rot, are highly contagious and there are no known cultivars with durable resistance to the pathogen. This study hypothesized (a) that crops intercropped or rotated with potato, plants in the same family, and plants grown in the neighbouring fields can host the pathogen and they can be potential sources of primary inoculum, and (b) that potato cultivars currently multiplied by the public tissue culture laboratory in Rwanda are less susceptible to the pathogen. Fourteen plant species and potato, and nine potato cultivars were tested for susceptibility to an RSSC phylotype II strain under greenhouse conditions. The bacteria induced symptoms on potato, tomato, tree tomato, sweet pepper, and eggplant only. Among the plant species with symptoms, potato, tomato, and tree tomato wilted completely. There was a significant difference in days to symptom expression and to complete wilting (p < .0001). While all tested potato cultivars were found to be susceptible, the number of days to first symptom expression, days to complete wilting, area under the disease progress curve (AUDPC), and the number and weight of harvested tubers varied considerably. Cultivars Cruza, Kinigi, and CIP-58 were less susceptible whereas the cultivars Gikungu, Kirundo, and Victoria were highly susceptible. There is a strong need to search for other sources of resistance. The results indicate that some plant species that might serve as a reservoir of the bacterium should be avoided in the vicinity of potato crops.  相似文献   

16.
Polymerase chain reaction (PCR) methods for detection and differentiation of Ralstonia solanacearum strains were compared. The 16S–23S rRNA gene ITS sequence data revealed the two main sequence clusters (divisions I and II) of R. solanacearum and further subclusters of division II. Based on this sequence data, primers were designed which differentiated divisions I and II. Furthermore, to improve reliability of the PCR assay for routine detection of R. solanacearum in host plants, a novel multiplex PCR assay was developed in which the pathogen-specific sequences are coamplified with host plant DNA as an internal PCR control (IPC). The assay was validated during routine testing of potato samples submitted in official surveys. Of 4300 samples from 143 cultivars, 13 tested positive in both multiplex PCR and immunofluorescence (IF) assays and could be confirmed by bioassay in tomato seedlings and reisolation of the pathogen. The IPC was successfully amplified from all samples tested. A further 12 samples gave positive IF results which were not confirmed by either the multiplex PCR or tomato bioassay, indicating a greater specificity of the latter two assays.  相似文献   

17.
Survival of Ralstonia solanacearum race 3 biovar 2 (phylotype II sequevar 1) in Egyptian soils and compost was studied under laboratory and field conditions. Survival of the pathogen under laboratory conditions varied with temperature, water potential and soil type, with temperature being the major determinant of survival of the pathogen. The effects of temperature and moisture content were variable between different experiments, but survival was generally longer at 15°C than at 4, 28 and 35°C respectively. Survival was also longer when moisture levels were constant compared with varying moisture levels at all temperatures. In experiments to compare the effects of progressive drying in sandy and clay soils there was a difference in survival times between the two soil types. In sandy soils, the pathogen died out more rapidly when soil was allowed to dry out than in controls where the soil was kept at constant water potential. In clay soils there was little difference between the two treatments, possibly due to the formation of a hard impermeable outer layer during the drying process, which retarded water loss from within. Survival in mature composts at 15°C was of the same order of magnitude as in soils but shorter at 28°C, possibly owing to increased biological activity at this temperature, or a resumption of the composting process, with concomitant higher temperatures within the compost itself. The maximum survival time recorded over all soil types and conditions during in vitro studies was around 200 days. In field studies, the maximum survival time in both bare sand and clay was around 85 days at depths up to 50 cm. The survival time was reduced in field experiments carried out in summer to less than 40 days and in one study when the ground was flooded for rice cultivation, the bacterium could not be detected 14 days after flooding. The maximum survival time of R. solanacearum in infected plant material or in infested soil samples incorporated into compost heaps was less than 2 weeks. At the culmination of field soil and compost experiments, no infection was detected in tomato seedlings up to 10 weeks after transplanting into the same soils or composts under glasshouse conditions at a temperature of 25°C.  相似文献   

18.
Ralstonia solanacearum, the causal agent of bacterial wilt of tomato, grows in infected plants and migrates from the roots into the soil. We investigated the effectiveness of bacterial wilt-resistant tomato rootstock in reducing the migration of R. solanacearum from susceptible scions into the soil. Rootstock stems were either 3–5 cm tall (low-grafted, LG) or ≥?10 cm tall (high-grafted, HG). After inoculation of scions of the susceptible cultivar (SC) with R. solanacearum below the first flower, there was no difference in disease progression among LG, HG, and ungrafted SC plants, and plants had wilted by 2 weeks. However, the rate of detection of R. solanacearum in the soil of wilted plants was reduced by grafting. The size of the R. solanacearum population in the soil of fully wilted plants increased in the order of HG?<?LG?<?SC. These results show that grafting onto resistant rootstock strongly suppressed the migration of R. solanacearum into the soil by the time of full wilting, and the effect was stronger with a longer rootstock. Migration of R. solanacearum into soil increased with increasing disease severity in SC, LG and HG. These facts suggest that early uprooting of slightly infected plants could control the spread of the bacteria into the soil.  相似文献   

19.
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20.
Ralstonia solanacearum causes bacterial wilt disease in many plant species, including mulberry. Here, we used a suppression subtractive hybridization (SSH) approach to identify specific DNA fragments in R. solanacearum race 5-biovar 5. The genome of the R. solanacearum M7 strain was subtracted from that of the GMI1000 strain, resulting in the identification of 85 subtracted fragments. The primer set MG67-F/R for identification of Ralstonia solanacearum race 5-biovar 5 strains was designed on the basis of the clone MG67 sequence. Furthermore, a multiplex PCR was developed by using the primer set MG67-F/MG67R in combination with the species-specific primer pair 759/760. A 156 bp r5-bv5-specific fragment, together with a 282 bp species-specific fragment, was amplified from all tested R. solanacearum r5-bv5 strains. The sensitivity of the multiplex PCR made it possible to detect concentrations as low as 102 CFU ml?1 of pure culture. Moreover, the r5-bv5-specific multiplex PCR was successfully applied to detect Ralstonia solanacearum race 5-biovar 5 strains in diseased mulberry samples. Therefore, the multiplex PCR assay can be used as a reliable diagnostic technique to enable researchers to rapidly identify isolates of R. solanacearum race 5-biovar 5.  相似文献   

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