Involvement of jasmonic acid signalling in bacterial wilt disease resistance induced by biocontrol agent Pythium oligandrum in tomato

When the biocontrol agent Pythium oligandrum (PO) colonizes the rhizosphere, it suppresses bacterial wilt disease in tomato (Solanum lycopersicum cv. Micro‐Tom) caused by Ralstonia solanacearum, and a homogenate of its mycelia exhibits elicitor activity, inducing an ethylene (ET)‐dependent defence response in Micro‐Tom. Since salicylic acid (SA) and jasmonic acid (JA) play an important role in plant defence responses to pathogens, the involvement of SA‐ and JA‐dependent signal transduction pathways in resistance to R. solanacearum was investigated in tomato roots treated with a mycelial homogenate of PO. Bacterial wilt disease was also suppressed in tomato cv. Moneymaker treated with the PO homogenate. However, the SA‐inducible PR‐1(P6) gene was not up‐regulated in either Micro‐Tom or Moneymaker. SA did not accumulate in homogenate‐treated roots in comparison with distilled water‐treated controls, even 24 h after inoculation. Induced resistance against R. solanacearum was not compromised in SA‐non‐accumulating NahG transgenic plants treated with the PO homogenate. On the other hand, the expression of the JA‐responsive gene for the basic PR‐6 protein was induced in both tomato cultivars treated with the PO homogenate. Furthermore, quantitative disease assays showed that the induced resistance against R. solanacearum was compromized in PO homogenate‐treated jai1‐1 mutant plants defective in JA signalling. These results indicated that the JA‐dependent signalling pathway is required for PO‐induced resistance against R. solanacearum in tomato.     

Biological control of Fusarium oxysporum f. sp. lycopersici

Fungi known to produce lytic enzymes were used in an attempt to control wilt of tomato caused by Fusarium oxysporum f. sp. lycopersici (FOL). Some of the fungal species (Penicillium oxalicum, Penicillium purpurogenum and Aspergillus nidulans) damaged hyphae of FOL in vitro and reduced the numbers of microconidia in the soil. Treatments with fungi did not result in a reduction in either chlamydospores of FOL in soil or populations of FOL in the rhizosphere of tomato. P. oxalicum was the most effective agent of biocontrol, and it reduced disease severity in both non-autoclaved (20% decrease) and sterile soil. In sterile soil, P. oxalicum reduced disease with different levels of severity (27% decrease at high levels and 50% decrease at low levels). Disease control by A. nidulans and P purpurogenum was only achieved when disease severity was low in sterile soil (55% and 45%, respectively).     

Development of a rapid,accurate glasshouse bioassay for assessing fusarium wilt disease responses in cultivated Gossypium species

A rapid glasshouse‐based bioassay method to screen large numbers of cotton plants for responses to Fusarium oxysporum f. sp. vasinfectum (Fov) was developed. Different Fov inoculum concentrations and methods of inoculation were assessed using resistant and susceptible cotton cultivars. Cotton seeds were planted directly into Fov‐inoculated soil. Studies of seed germination, seedling establishment, seedling mortality and fusarium wilt symptoms (i.e. stunting, foliar symptoms and vascular browning) were performed to optimize the bioassay parameters. Growing seedlings in Fov‐inoculated soils at 5 × 10⁴ or 1 × 10⁵ CFU g^?1 soil, in individual seedling tubes with 12 h at 28–30°C and 12 h at 15–18°C, gave consistent results when assessing Fov disease responses 6 weeks after inoculation. When fusarium wilt resistance ranks (FWRRs) and vascular browning index (VBI) means of 18 Australian and other cotton cultivars from the Fov glasshouse bioassay were compared against their fusarium field performance ranks (F‐ranks), assessed on adult plants for cotton cultivar release, Pearson’s correlation was highly significant for both comparisons. The level of congruence between field and glasshouse data indicated that this protocol should be an effective tool for large‐scale screening for Fov‐resistance responses in diverse germplasm and breeding populations and for advancing genetic research to develop molecular markers for Fov resistance in cotton.     

Detection of race 1 strains of <Emphasis Type="Italic">Ralstonia solanacearum</Emphasis> in field samples in Taiwan using a BIO-PCR method

Bacterial wilt caused by race 1 strains of Ralstonia solanacearum is endemic on tomato produced in diverse agro-ecosystems in Taiwan. Using a new BIO-PCR protocol developed in this study, R. solanacearum was detected in soil, weed, and water samples collected from eight fields with different disease histories and cropping systems located in major tomato production areas. The sensitivity of the BIO-PCR was 1.9 CFU ml⁻¹ and 17 CFU g⁻¹ of soil for pure suspension and infested soil, respectively. The positive detection frequency of the BIO-PCR method was 66.6, 39.6, 23.1, and 31.8% for all tested samples of soil, weed rhizosphere soil, weed root, and water, respectively, and was higher than plating on MSM-1 medium. Detection of R. solanacearum from field soil indicated that spatial distribution of the pathogen in the field was not even regardless of the presence or absence of the disease and the different agro-ecosystems where the sampled fields were located, and the degree of unevenness was higher when tomato was absent from the field. Weed rhizosphere soils could be good sampling targets to monitor the pathogen in the field, because a higher positive detection proportion and population of R. solanacearum were found in the rhizosphere rather than the root of the collected weed samples. Symptomless weeds and contaminated irrigation, standing, or drainage waters were found to be important for the over-season survival and dissemination of R. solanacearum.     

Effects of timing and method of application of Penicillium oxalicum on efficacy and duration of control of Fusarium wilt of tomato

The effects of timing and method of application of Penicillium oxalicum on the control of fusarium wilt of tomato were investigated. Application of P. oxalicum to tomato seedlings in seedbeds reduced disease caused by Fusarium oxysporum f.sp. lycopersici in a growth chamber by 45–49% and in glasshouse experiments by 22–69%. Disease suppression was maintained for 60–100 days after inoculation with the pathogen in the glasshouse. No disease reduction was observed in tomato plants where P. oxalicum was applied to seeds. Treatment with P. oxalicum did not affect the population of F. oxysporum f.sp. lycopersici in the rhizosphere.     

Rapid detection of Fusarium oxysporum f. sp. lactucae on soil,lettuce seeds and plants using loop‐mediated isothermal amplification

Fusarium oxysporum f. sp. lactucae (FOL) is a soil‐ and seedborne pathogen and the causal agent of fusarium wilt on lettuce. Four races have been identified within FOL, with different worldwide distribution. Several molecular techniques have been used to detect and identify this pathogen; however, not all of them have the optimal characteristics in terms of sensitivity to perform FOL detection in plant and seed material. A loop‐mediated isothermal amplification (LAMP) assay was developed based on the sequence‐characterized amplified region (SCAR) obtained in a previous rapid amplification of polymorphic DNA (RAPD) study. The LAMP assay has been validated according to the EPPO standard PM7/98. The LAMP assay was tested with lettuce seeds, soil and plant material, and can be used successfully to amplify DNA from each of these matrices. In seed lots artificially inoculated with FOL, the detection limit of the LAMP test was 0.004% infected seed.     

Selection and Screening of Endorhizosphere Bacteria from Solarized Soils as Biocontrol Agents Against Verticillium dahliae of Solanaceous Hosts

Verticillium dahliae antagonistic endorhizosphere bacteria were selected from root tips of tomato plants grown in solarized soils. Fifty-three out of the 435 selected bacterial isolates were found to be antagonistic against V. dahliae and several other soilborne pathogens in dual cultures. Significant biocontrol activity against V. dahliae in glasshouse trials was demonstrated in three of 18 evaluated antagonistic isolates, provisionally identified as Bacillus sp. Although fluorescent pseudomonads were also isolated from root tips of tomato plants, none of the tested isolates exercised any significant antagonistic activity against V. dahliae in dual cultures. So these isolates were not tested in glasshouse trials in this study. Finally, two of the most effective bacterial isolates, designated as K-165 and 5-127, were shown to be rhizosphere colonizers, very efficient in inhibiting mycelial growth of V. dahliae in dual cultures and successfully controlling Verticillium wilt of solanaceous hosts. In glasshouse experiments, root dipping or soil drenching of eggplants with bacterial suspension of 10⁷cfu ml^–1 resulted in reduced disease severity expressed as percentage of diseased leaves (40–70%) compared to the untreated controls under high V. dahliae inoculum level (40 microsclerotia g^–1 soil). In heavily Verticillium infested potato fields, experiments with potato seeds dusted with a bacterial talc formulation (10⁸cfu g^–1 formulation), showed a significant reduction in symptom development expressed as percentage of diseased potato plants and a 25% increase in yield over the untreated controls. As for their effectiveness in increasing plant height, both bacterial isolates K-165 and 5-127 produced indolebutyric, indolepyruvic and indole propionic acids. Both antagonists are considered as plant growth promoting rhizobacteria bacteria since significantly increased the height of treated plants compared with the untreated controls. Chitinolytic activity test showed that both isolates were able to produce chitinase. Testing rhizospheric and endophytic activity of the antagonists it was shown that although the bacteria are rhizosphere inhabitants they also preferentially colonize the endorhizosphere of tomatoes and eggplants. Fatty acid analysis showed that isolate K-165 could belong to Paenibacillus alvei while 5-127 to Bacillus amiloliquefaciens.     

Accumulation of phytoalexins in susceptible and resistant near-isogenic lines of tomato infected with Verticillium albo-atrum or Fusarium oxysporum f.sp. lycopersici

The time course of accumulation of two phytoalexins, the terpenoid rishitin and the polyacetylene cis-tetradeca-6-ene-1,3-diyne-5,8-diol, was determined in near-isogenic susceptible and resistant tomato lines inoculated with either Verticillium albo-atrum or Fusarium oxysporum f.sp. lycopersici.Cultivars containing the Ve gene for verticillium wilt resistance accumulated phytoalexins at a rate similar to that in susceptible plants following stem inoculation with V. albo-atrum. Higher amounts of phytoalexins were isolated from susceptible than from resistant plants at 11 days after inoculation. Inoculum concentrations of 10⁵, 10⁶, 10⁷ and 10⁸ conidia ml⁻¹ had no differential effect on phytoalexin accumulation at 3 days after inoculation. Also, no differences were observed between fungal growth in susceptible and resistant cultivars during that period.A cultivar containing the I-1 gene for fusarium wilt resistance contained more rishitin than did susceptible plants at 2 and 3 days after inoculation with 10⁷ conidia of F. oxysporum f.sp. lycopersici ml⁻¹, but at 7 and 11 days after inoculation more rishitin had accumulated in the susceptible plants.No difference was observed between the rate of accumulation of phytoalexin in stem segments from resistant and susceptible plants inoculated by vacuum-infiltration.To estimate the concentration of phytoalexins in the xylem fluid, sap was expressed from vascular tissue and amounts of phytoalexins were determined in the sap and in the expressed tissue. Less than 5% of the phytoalexins present in stem segments was recovered from the sap, indicating that their concentration in the xylem fluid may be relatively low.The role of phytoalexins in resistance to verticillium and fusarium wilt is discussed.     

Suppression of fusarium wilt of radish by co-inoculation of fluorescentPseudomonas spp. and root-colonizing fungi

In an earlier study, treatment of radish seed with the bacteriumPseudomonas fluorescens WCS374 suppressed fusarium wilt of radish (Fusarium oxysporum f. sp.raphani) in a commercial greenhouse [Leemanet al., 1991b, 1995a]. In this greenhouse, the areas with fusarium wilt were localized or expanded very slowly, possibly due to disease suppressiveness of the soil. To study this phenomenon, fungi were isolated from radish roots collected from the greenhouse soil. Roots grown from seed treated with WCS374 were more abundantly colonized by fungi than were roots from nonbacterized plants. Among these were several species known for their antagonistic potential. Three of these fungi,Acremonium rutilum, Fusarium oxysporum andVerticillium lecanii, were evaluated further and found to suppress fusarium wilt of radish in a pot bioassay. In an induced resistance bioassay on rockwool,F. oxysporum andV. lecanii suppressed the disease by the apparent induction of systemic disease resistance. In pot bioassays with thePseudomonas spp. strains, the pseudobactin-minus mutant 358PSB^– did not suppress fusarium wilt, whereas its wild type strain (WCS358) suppressed disease presumably by siderophore-mediated competition for iron. The wild type strains of WCS374 and WCS417, as well as their pseudobactin-minus mutants 374PSB^– and 417PSB^– suppressed fusarium wilt. The latter is best explained by the fact that these strains are able to induce systemic resistance in radish, which operates as an additional mode of action. Co-inoculation in pot bioassays, ofA. rutilum, F. oxysporum orV. lecanii with thePseudomonas spp. WCS358, WCS374 or WCS417, or their pseudobactin-minus mutants, significantly suppressed disease (except forA. rutilum/417PSB^– and all combinations with 358PSB^–), compared with the control treatment, if the microorganisms were applied in inoculum densities which were ineffective in suppressing disease as separate inocula. If one or both of the microorganism(s) of each combination were applied as separate inocula in a density which suppressed disease, no additional suppression of disease was observed by the combination. The advantage of the co-inoculation is that combined populations significantly suppressed disease even when their individual population density was too low to do so. This may provide more consistent biological control. The co-inoculation effect obtained in the pot bioassays suggests that co-operation ofP. fluorescens WCS374 and indigenous antagonists could have been involved in the suppression of fusarium wilt of radish in the commercial greenhouse trials.Abbreviations CFU colony forming units - KB King's B - PGPR plant growth-promoting rhizobacteria - CQ colonization quotient     

Siderophore-mediated competition for iron and induced resistance in the suppression of fusarium wilt of carnation by fluorescent Pseudomonas spp

The mechanisms of suppression of fusarium wilt of carnation by two fluorescentPseudomonas strains were studied.Treatments of carnation roots withPseudomonas sp. WCS417r significantly reduced fusarium wilt caused byFusarium oxysporum f. sp.dianthi (Fod). Mutants of WCS417r defective in siderophore biosynthesis (sid^–) were less effective in disease suppression compared with their wild-type. Treatments of carnation roots withPseudomonas putida WCS358r tended to reduce fusarium wilt, whereas a sid^– mutant of WCS358 did not.Inhibition of conidial germination of Fod in vitro by purified siderophores (pseudobactins) of bothPseudomonas strains was based on competition for iron. The ferrated pseudobactins inhibited germination significantly less than the unferrated pseudobactins. Inhibition of mycelial growth of Fod by bothPseudomonas strains on agar plates was also based on competition for iron: with increasing iron content of the medium, inhibition of Fod by thePseudomonas strains decreased. The sid^– mutant of WCS358 did not inhibit Fod on agar plates, whereas the sid^– mutants of WCS417r still did. This suggests that inhibition of Fod by WCS358r in vitro was only based on siderophore-mediated competition for iron, whereas also a non-siderophore antifungal factor was involved in the inhibition of Fod by strain WCS417r.The ability of thePseudomonas strains to induce resistance against Fod in carnation grown in soil was studied by spatially separating the bacteria (on the roots) and the pathogen (in the stem). Both WCS417r and its sid^– mutant reduced disease incidence significantly in the moderately resistant carnation cultivar Pallas, WCS358r did not.It is concluded that the effective and consistent suppression of fusarium wilt of carnation by strain WCS417r involves multiple mechanisms: induced resistance, siderophore-mediated competition for iron and possibly antibiosis. The less effective suppression of fusarium wilt by WCS358r only depends on siderophore-mediated competition for iron.     

Effect of cold-water irrigation on bacterial wilt pathogen of tomato

Bacterial wilt caused by Ralstonia solanacearum is one of the most devastating bacterial diseases of plants worldwide. Management of bacterial wilt in tomato and other crops has been difficult, and so novel but easily implemented control methods are being sought. To evaluate the effect of cold-water irrigation on bacterial wilt of tomato, four treatments were used in which CF (chemically fertilized) soil and CF + FYM (chemical fertilizer + farmyard manure [FYM]) soil were inoculated with a bacterial suspension (R. solanacearum strain YU1Rif43) at 10⁶ colony forming units (CFU) g^?1 soil. Tomato seedlings were grown in Agri-pots in a plant growth chamber. The soil was irrigated with water that was kept at the same temperature in each treatment: 4, 10, 20, or 30°C. Incidence and severity of wilt, counting of the colonies of the culturable population of pathogen, and dry-mass and height of the plants were examined. After 45 days and in both kinds of soil, most of the plants had wilted in soil irrigated at 30°C. Wilt incidence was substantially reduced when transplanted seedlings were irrigated at lower temperatures (4–20°C). Survival of R. solanacearum was also reduced after being irrigated with water at lower temperatures, indicating that the reduced incidence of wilt was linked to reduced survival of the pathogen. Dry-mass and plant height were slightly higher under control conditions than in soils irrigated at lower temperatures. This study suggests that cold-water irrigation could significantly reduce bacterial wilt of tomato and have an adverse effect on survival of the wilt pathogen.     

Visualization of <Emphasis Type="Italic">Ralstonia solanacearum</Emphasis> cells during biocontrol of bacterial wilt disease in tomato with <Emphasis Type="Italic">Pythium oligandrum</Emphasis>

The biocontrol agent Pythium oligandrum (PO) can suppress bacterial wilt caused by Ralstonia solanacearum (RS) in tomato. To understand the primary biocontrol mechanisms of bacterial wilt by PO, we pretreated tomato plants with sterile distilled water or preinoculated them with PO, followed by inoculation with RS, then observed PO and RS in fixed sections of tomato tissues using a confocal laser-scanning microscope and fluorescence labeling until 14 days after the inoculation with RS. Horizontal and vertical movement of RS bacteria was frequently observed in the xylem vessels of roots and stems of tomato plants (cv. Micro-Tom) that had not been inoculated with PO. In plants that were preinoculated with PO, the movement of RS was suppressed, and bacteria appeared to be restricted to the pit of vessels, a reaction similar to that observed in resistant rootstocks. PO colonization was mainly observed at the surfaces of taproots, the junctions between taproots and lateral roots, and the middle sections of the lateral roots. PO was not observed near wound sites or root tips where RS tended to colonize. However, RS colonization was significantly repressed at these sites in PO preinoculated plants. These observations suggest that the induction of plant defense reactions is the main mechanism for the control of tomato bacterial wilt by PO, not direct competition for infection sites.     

Biological control agents (BCAs) of verticillium wilt: influence of application rates and delivery method on plant protection,triggering of host defence mechanisms and rhizosphere populations of BCAs

Verticillium dahliae causes severe yield reductions in a variety of important annual crops worldwide. Control of verticillium wilt has relied on soil fumigation; however, the use of the main soil fumigant, methyl bromide, has been banned in the European Union since 2010, creating a demand for novel crop protectants. As such, the use of biocontrol agents (BCAs) is an appealing management strategy. Prerequisites for the development of a successful BCA are an understanding of the modes of action of the antagonist, its ecological fitness and an efficient and economically feasible delivery system. Therefore, two BCAs (Paenibacillus alvei K165 or the nonpathogenic Fusarium oxysporum F2) and two release strategies (seed coating or amendment of the transplant soil plug) were assessed against verticillium wilt of aubergine (eggplant). Mixing the transplant soil plug with K165 or F2, at a rate of 10 and 20% (v/v), respectively, reduced verticillium wilt symptom development. Furthermore, a positive correlation was revealed between the release strategy and the BCA rhizosphere population. Correlation analysis also showed that disease severity was negatively correlated to the rhizosphere size of the BCA population. In addition, qPCR analysis showed that both BCAs induced the expression of the pathogenesis‐related (PR) proteins PR1 and PR4 in the stem of aubergines before and after inoculation with V. dahliae in a manner that suggests a link with the rhizosphere size of the BCA population.     

Fusarium wilt‐resistant lines of Brazil banana (Musa spp., AAA) obtained by EMS‐induced mutation in a micro‐cross‐section cultural system

This study combined the micro‐cross‐section cultural system with in vitro mutagenesis induced by ethyl methanesulphonate (EMS) to screen for fusarium wilt‐resistant lines of Brazil banana (Musa spp., AAA). The results indicated that the optimum EMS concentration and duration for the treatment of micro‐cross‐sections cut from the pseudostem of tissue‐cultured plantlet were 300 mm and 60 min, respectively. Under the optimal treatment, an average of 2·2 regenerated shoots were produced from each explant. One hundred regenerated plantlets were used for screening for fusarium wilt‐resistant lines by the early screening technique. The initial disease symptom – yellowing in lower leaves of susceptible plantlets – was observed 2 weeks after inoculation. After 2 months, only six plants survived – the putative fusarium wilt‐resistant lines. The fusarium wilt pathogen Fusarium oxysporum f. sp. cubense race 4, was identified in the preliminary test field by a SCAR marker technique. Of the six putative resistant lines, five survived the preliminary field test. The regenerated plantlets from these five fusarium wilt‐resistant lines were subjected to early screening again, where they showed markedly reduced disease incidences compared with regenerated plantlets of Brazil banana (control). It was concluded that EMS‐induced mutation of banana through the micro‐cross‐section cultural system is potentially useful for banana improvement.     

Hormetic UV-C seed treatments for the control of tomato diseases

Hormesis is a dose response phenomenon in which low, non-damaging doses of a stressor bring about a positive response in the organism undergoing treatment. Evidence is provided here that hormetic UV-C treatments of tomato seed can control disease caused by Botrytis cinerea, Fusarium oxysporum f. sp. lycopersici (FOL) and f. sp. radicis-lycopersici (FORL) on tomato (Solanum lycopersicum). Treating seeds with a 4 kJ m⁻² dose of UV-C significantly reduced both the disease incidence and progression of B. cinerea, with approximately 10% reductions in both on cv. Shirley. Disease severity assays for FOL and FORL on cv. Moneymaker showed dose-dependent responses: UV-C treatments of 4 and 6 kJ m⁻² significantly reduced the disease severity scores of FOL, whilst only the 6 kJ m⁻² showed significant reductions for FORL. To determine the effects of treatment on germination and seedling growth, UV-C doses of 4, 8 and 12 kJ m⁻² were performed on cv. Shirley. No negative impacts on germination or seedling growth were observed for any of the treatments. However, the 8 kJ m⁻² treatment showed significant biostimulation, with increases in seedling, root and hypocotyl dry weight of 11.4%, 23.1% and 12.0%, respectively, when compared to the control. Furthermore, significant increases in the root-mass fraction (10.6%) and root:shoot ratio (13.1%) along with a decrease in shoot-mass fraction (2.0%) indicates that the 8 kJ m⁻² treatment stimulated root growth to the greatest extent. There was no effect on hypocotyl and primary root length or the number of lateral roots, indicating no adverse effects to basic root architecture or seedling growth.     

Studies on the colonization of axenically grown tomato plants by a GFP-tagged strain of Clavibacter michiganensis subsp. michiganensis

In this study, colonization and disease development of axenically-grown tomato plants by Clavibacter michiganensis subsp. michiganensis (Cmm), the causative agent of bacterial wilt and canker, was investigated. For this, a spontaneous rifampicin resistant strain of Cmm was tagged with a marker that expressed a green fluorescent protein (GFP) in a stable way and which possessed a similar virulence to the parental strain. In vitro plants were drop-inoculated at the stem base and the population dynamics was determined by dilution pour-plating in a selective medium. At 3 h after inoculation, Cmm was already present in low densities in roots, stems and leaves. At 16 dpi, Cmm was found throughout the entire plant in high densities of ca. 10¹⁰ cfu g^?1. Symptoms developed in the in vitro plants typical for Cmm, such as canker, wilting and growth reduction. The presence of Cmm in vascular and parenchymatic tissue of in vitro tomato plants was confirmed by epifluorescence stereo- and confocal laser scanning microscopy. This study showed that in vitro tomato plants can be effectively used for detailed studies on interactions between Cmm and its host, in particular if a GFP-tagged strain of the pathogen is used.     

木霉分生孢子和厚垣孢子对黄瓜叶片抗氧化系统及枯萎病防效的影响

采用前期筛选出的对黄瓜枯萎病菌有较好拮抗作用的3株木霉菌,即哈茨木霉菌(Trichoderma harzianum)809、拟康氏木霉菌(Trichoderma pseudokoningii)886和棘孢木霉菌(Trichoderma asperellum)525,利用盆栽试验,测定了木霉菌分生孢子和厚垣孢子对黄瓜幼苗...     

Specific Detection of Tomato Pathotype of Verticillium dahliae by PCR Assays

Verticillium dahliae Klebahn is the causal agent of tomato wilt disease. Isolates of V. dahliae can be classified based on pathogenicity to tomato, but the pathotypes are indistinguishable in morphology. We designed PCR primers for specific detection of isolates pathogenic to tomato (tomato pathotype) from the sequences of a pathotype-specific gene, vdt1. With the primer pair Tg5/Tc3, a PCR product (approximately 3.2 kb) specific to tomato pathotype was amplified from the genomic DNA of isolates. Using the primer pair, a tomato pathotype isolate was specifically detected from hypocotyls of inoculated tomato and eggplant. On the other hand, no amplification was observed from non-tomato pathotype isolates of V. dahliae, some other wilt pathogens of tomato and a healthy host plant. Therefore, the primer pair can be useful for pathotype-specific detection of V. dahliae as well as for diagnosis of wilt disease of tomato plant. Received 7 September 2001/ Accepted in revised form 3 December 2001     

Involvement of gacA Gene in the Suppression of Tomato Bacterial Wilt by Pseudomonas fluorescens FPT9601

Pseudomonas fluorescens FPT9601, a plant growth-promoting rhizobacterium (PGPR) isolated from tomato rhizosphere, can protect tomato (Lycopersicon esculentum Mill) from bacterial wilt disease caused by Ralstonia solanacearum. This strain produces antibiotics 2,4-diacetylphloroglucinol (2,4-DAPG) and hydrogen cyanide (HCN). It also produces proteases and uncharacterized siderophores (Sid). A mutant strain SM2214, obtained by Tn5 insertion, did not produce 2,4-DAPG, HCN or proteases, but overproduced Sid. Marker-exchange mutagenesis confirmed that a single transposon insertion caused the multiple phenotypic changes of this mutant. Complementation of the mutant with a 1.3-kb DNA fragment that was amplified from genomic DNA of the wild-type P. fluorescens strain by PCR could restore the lost functions of the mutant strain. Nucleotide sequencing revealed that the fragment contained a 642-bp open reading frame (ORF) highly homologous to the regulator responser gene gacA. The in vitro anti-bacterium test and plant protection experiment under greenhouse conditions indicated that the gacA gene played an important role in the suppression of tomato bacterial wilt disease. Received 20 November 2000/ Accepted in revised form 19 January 2001     

植物疫苗鄂鲁冷特对番茄青枯病的田间防治效果

为了明确植物疫苗鄂鲁冷特对番茄育苗及其田间青枯病防治效果的影响,在番茄育苗基质中添加植物疫苗,测定处理后种苗株高、根系长度及出苗率;在田间种植共设疫苗袋装、沟施、浇灌及生防菌剂、化学农药和清水对照6个处理,对根系土壤的养分含量、植株的生物学性状及青枯病发病率进行研究。结果表明,植物疫苗处理分别能使番茄植株株高、根系长度和出苗率提高13.71%、68.20%和56.66%;疫苗袋装和沟施处理的番茄根系土壤有机质、全氮、全磷、全钾和交换性钙含量均显著高于其它处理;疫苗袋装、沟施和浇灌均能显著提高番茄植株的株高、花数和产量,其中疫苗袋装处理效果最好,番茄产量最高为99.55 t/hm~2,对不同生育期的平均防治效果最高为93.47%。表明植物疫苗鄂鲁冷特的应用能育出壮苗,降低青枯发病率,提高产量。