Root Bioassay Research Articles

Feeding the Microbes: A Strategy to Control Verticillium Wilt

Verticillium dahliae is one of the most devastating soilborne pathogens for horticulture production. The pathogen has a broad host range and currently there is no effective chemical disease management, therefore, novel sustainable integrated disease management strategies should be considered. In this respect, we questioned whether the soil application of common microbiological growth media can influence the plant—microbe interactions and, subsequently, offer protection against V. dahliae. Indeed, the addition of Nutrient Broth (NB) and Potato Dextrose (PD) in non-sterilized soil reduced Verticillium wilt symptoms in eggplants. The addition of NB in sterilized soil did not reduce the disease symptoms compared to controls, however the addition of PD enhanced plant protection against V. dahliae. Following the results of a split root bioassay in eggplants, the possibility that NB and PD triggered the plant defense mechanisms against V. dahliae was excluded, since NB and PD did not reduce wilt symptom. Therefore, PD may be used as an easy food source for V. dahliae, detouring the pathogen from the root system of plants, while NB may affect the soil microbiome by enhancing antagonism in rhizosphere, or antagonistic interaction between V. dahliae and rhizospheric microbiome. Subsequently, several bacterial strains were isolated from the NB-treated rhizosphere and examined for their biocontrol activity against V. dahliae. Among the examined strains, a Pseudomonas putida strain, Z13, significantly reduced Veticillium severity and incidence under greenhouse conditions.

Imazethapyr persistence in sandy loam detected using white mustard bioassay

ABSTRACTField experiments were conducted during two years at Srem region to investigate the influence of meteorological conditions, time and rate of application on soil persistence of imazethapyr in sandy loam type of soil. Imazethapyr was applied PRE- and POST-EM and in both cases in three application rates: 80, 120 and 160 g a.i./ha. Soil samples were collected from the day of herbicide application in predetermined intervals up to one year after application and residual concentrations were determined with a white mustard root bioassay. Imazetapyr persistence was significantly influenced by meteorological conditions with average half-life being 6 days longer in season with lower precipitation level. Time of application induced slower imazethapyr dissipation resulting in higher average t1/2 (seven and nine days in first and second year of examination, respectively). Application rates had no consistent effect on imazethapyr persistence. Imazethapyr residue level one year after application caused no visible injuries on white mustard shoots, while root growth reduction ranged from 4.6 to 27.7%. Obtained residue levels were further compared with known data on crop sensitivity in order to assess possibility of crop injuries one year after imazethapyr application.

Comparison of bioassays to biotype grape phylloxera ("Daktulosphaira vitifoliae" Fitch) on "Vitis" ssp.

Grape phylloxera biotypes exist throughout viticultural regions causing substantial economic losses. In the past different biotyping assays were employed to determine host adaption and potential harm of phylloxera strains or field populations. Standardised and efficient laboratory assays are required to define biotypes according to their aggressivity as well as to make accurate pest management and quarantine decisions. We aim to provide information on the consistency of the three most commonly used assays to accurately identify grape phylloxera biotype. Two phylloxera biotypes (A, C) were tested on two host plants (rootstock 'Teleki 5C' V. berlandieri x V. riparia and V. vinifera 'Riesling') using three assays: Simple isolation chamber, excised root bioassay and aseptic dual culture bioassay. Insect number, life table and plant-based response parameters (root galling) were compared. The simple isolation chamber and aseptic dual culture bioassay produced consistent results, whereas the excised root bioassay did not. We demonstrated that biotype results depend on whether the technique used is tuberosity- or nodosity-based. Pest management decision based on a single assay may inaccurately assess the phylloxera aggressivity potential. Thus, we recommend using two assay types which allows comparison of both root gall types.

Green Synthesized Zinc Oxide (ZnO) Nanoparticles Induce Oxidative Stress and DNA Damage in Lathyrus sativus L. Root Bioassay System.

Zinc oxide nanoparticles (ZnONP-GS) were synthesised from the precursor zinc acetate (Zn(CH3COO)2) through the green route using the milky latex from milk weed (Calotropis gigantea L. R. Br) by alkaline precipitation. Formation of the ZnONP-GS was monitored by UV-visible spectroscopy followed by characterization and confirmation by energy-dispersive X-ray spectroscopy (EDX), transmission electron microscopy (TEM), and X-ray diffraction (XRD). Both the ZnONP-GS and the commercially available ZnONP-S (Sigma-Aldrich) and cationic Zn2+ from Zn(CH3COO)2 were tested in a dose range of 0–100 mg·L−1 for their potency (i) to induce oxidative stress as measured by the generation reactive oxygen species (ROS: O2•−, H2O2 and •OH), cell death, and lipid peroxidation; (ii) to modulate the activities of antioxidant enzymes: catalase (CAT), superoxide dismutase (SOD), guaiacol peroxidase (GPX), and ascorbate peroxidase (APX); and (iii) to cause DNA damage as determined by Comet assay in Lathyrus sativus L. root bioassay system. Antioxidants such as Tiron and dimethylthiourea significantly attenuated the ZnONP-induced oxidative and DNA damage, suggesting the involvement of ROS therein. Our study demonstrated that both ZnONP-GS and ZnONP-S induced oxidative stress and DNA damage to a similar extent but were significantly less potent than Zn2+ alone.

Polyvinyl polypyrrolidone attenuates genotoxicity of silver nanoparticles synthesized via green route, tested in Lathyrus sativus L. root bioassay

The silver nanoparticles (AgNPs) were synthesized extracellularly from silver nitrate (AgNO3) using kernel extract from ripe mango Mengifera indica L. under four different reaction conditions of the synthesis media such as the (i) absence of the reducing agent, trisodium citrate (AgNPI), (ii) presence of the reducing agent (AgNPII), (iii) presence of the cleansing agent, polyvinyl polypyrrolidone, PVPP (AgNPIII), and (iv) presence of the capping agent, polyvinyl pyrrolidone, PVP (AgNPIV). The synthesis of the AgNPs was monitored by UV–vis spectrophotometry. The AgNPs were characterised by the energy-dispersive X-ray spectroscopy, transmission electron microscopy, X-ray diffraction, and small-angle X-ray scattering. Functional groups on the AgNPs were established by the Fourier transform infrared spectroscopy. The AgNPs (AgNPI, AgNPII, AgNPIII and AgNPIV) were spherical in shape with the diameters and size distribution-widths of 14.0±5.4, 19.2±6.6, 18.8±6.6 and 44.6±13.2nm, respectively. Genotoxicity of the AgNPs at concentrations ranging from 1 to 100mgL−1 was determined by the Lathyrus sativus L. root bioassay and several endpoint assays including the generation of reactive oxygen species and cell death, lipid peroxidation, mitotic index, chromosome aberrations (CA), micronucleus formation (MN), and DNA damage as determined by the Comet assay. The dose-dependent induction of genotoxicity of the silver ion (Ag+) and AgNPs was in the order Ag+>AgNPII>AgNPI>AgNPIV>AgNPIII that corresponded with their relative potencies of induction of DNA damage and oxidative stress. Furthermore, the findings underscored the CA and MN endpoint-based genotoxicity assay which demonstrated the genotoxicity of AgNPs at concentrations (≤10mgL−1) lower than that (≥10mgL−1) tested in the Comet assay. This study demonstrated the protective action of PVPP against the genotoxicity of AgNPIII which was independent of the size of the AgNPs in the L. sativus L. root bioassay system.

The Efficacy of Chinese Medicinal Herbs towards Grape Phylloxera (Daktulosphaira vitifoliae Fitch) (Hemiptera, Phylloxeridae).

Bio-insecticidal effects of seven Chinese medicinal herbs on mortality, fecundity, developmental periods and life table parameters of the grape phylloxera were investigated. In an excised root bioassay experiment aqueous extracts from seven Chinese medicinal herbs increased grape phylloxera first instar mortality (26.00–38.50%) compared to other instars. The intrinsic rate of increase (rm), finite rate of increase (λ), fecundity rate and net reproductive rate (R0) were significantly reduced by A. bidentata, A. tataricus, O. basilicum, P. frutescens and N. cataria. In a glasshouse pot trial, eggs, nymphs, adults and total population were significantly reduced before population establishment compared to those after its population established, by A. bidentata, A. tataricus and O. basilicum. Overall, A. bidentata, A. tataricus and O. basilicum can be used to suppress all life-stages of grape phylloxera.

Evaluation of Inhibitory Effect of Organic and Inorganic Salts Against Ilyonectria liriodendri, The Causal Agent of Root Rot Disease of Kiwifruit

AbstractThis study evaluated efficacy of 42 organic and inorganic salts as possible alternatives to synthetic fungicides for the control of Ilyonectria root rot of kiwifruit. Preliminary in vitro tests showed that ammonium bicarbonate, ammonium carbonate, potassium benzoate, potassium sorbate, sodium benzoate and sodium metabisulphite at 2% completely inhibited mycelial growth of the fungus. No significant differences were observed among these salts and disodium EDTA (P ≤ 0.05). However, the ED50, minimum inhibition concentration (MIC), and minimum fungicidal concentration (MFC) values indicated that sodium metabisulphite was more toxic to Ilyonectria liriodendri than these other six salts. Soil bioassays showed that sodium metabisulphite, sodium benzoate and potassium sorbate at 0.25% completely inhibited mycelial growth of the fungus, whereas potassium benzoate reduced the mycelial growth of fungus by 90.30%; however, the differences in inhibitory effects were statistically insignificant (P ≤ 0.05). Moreover, there was no significant difference between 0.1% sodium metabisulphite and 0.5% ammonium carbonate, 0.75% ammonium bicarbonate and 1.5–2.0% disodium EDTA (P ≤ 0.05). Unlike disodium EDTA, complete inhibitory was observed with ammonium carbonate and ammonium bicarbonate at higher concentrations. However, in root bioassays, applications of 2% ammonium bicarbonate, 1.5% ammonium carbonate and 2% disodium EDTA were phytotoxic to kiwifruit seedlings, but 0.25% four other salts were neither phytotoxic to kiwifruit seedlings nor did it adversely affect root length, root fresh weight and root dry weight of seedling. This study also showed I. liriodendri to be capable of growth in both acidic and basic environments. However, while the fungus showed uninhibited growth at pH values of 5–11, growth decreased significantly at both higher and lower pH values (P ≤ 0.05) and was completely inhibited at pH 12.

Determination of Thiencarbazone in Soil by Oriental Mustard Root Length Bioassay

Using an oriental mustard root length bioassay, thiencarbazone bioavailability and dissipation in five Saskatchewan soils was investigated under laboratory conditions. Thiencarbazone bioavailability was assessed at 0 to 3.9 µg ai kg−1. Thiencarbazone concentrations corresponding to 50% inhibition (I50values) obtained from dose-response curves varied from 0.56 to 1.71 µg kg−1. Multiple regression analysis indicated that organic carbon content (P = 0.018) and soil pH (P = 0.017) predicted thiencarbazone bioavailability. Thiencarbazone dissipation was examined in soils incubated at 23 C and moisture content of 85% field capacity. Thiencarbazone half-lives estimated from dissipation curves were 9 to 50 d, and organic carbon content (P = 0.002) and soil pH (P = 0.008) were significant factors affecting thiencarbazone dissipation. Thiencarbazone bioavailability decreases and dissipation rate is slower in Canadian prairie soils of high organic matter content and low soil pH. Because root length of oriental mustard plants also was reduced by ammonium, therefore ammonium-containing or -producing fertilizers can cause false positive results for thiencarbazone soil residues. Canaryseed roots had the same sensitivity to ammonium as oriental mustard roots but were not inhibited by thiencarbazone. Therefore canaryseed root length bioassay was effective in identifying inhibition caused by ammonium toxicity. Use of oriental mustard root and canaryseed root bioassays together can aid in interpreting bioassay results for detection of thiencarbazone residues.

Screening of rootstock hybrids with Vitis cinerea Arnold for phylloxera resistance

AbstractThe resistance to phylloxera is one of the most important properties of grapevine rootstock. The breeding and selection of rootstock varieties represents a biological method of fight against phylloxera. In this study altogether 59 rootstock hybrids originating from nine different populations were evaluated. Pedigrees of all populations involved Vitis cinerea Arnold introduced through the hybrid Bruci/(Vitis berlandieri Planch. × Vitis rupestris Scheele) × Vitis cinerea Arnold/ and the variety Börner. Pot experiments were performed in a glasshouse while those with excised root pieces in root bioassay took place in a laboratory. This combination showed to be a very good selection tool for rootstock breeding. In seven rootstock hybrids, it was found that the resistance to phylloxera gallicoles and radicicoles was very high. The highest number of resistant hybrids was found in Binova × Börner. The variety Börner showed to be the best gene source of phylloxera resistance and even in case of a simple crossing produced sufficient number of resistant progeny.

Systemic Resistance in Tomato against Meloidogyne incognita Induced by Plant Growth Promoting Rhizobacterium, Pseudomonas fluorescens

Resistance-related enzymatic activities due to selected strains of Pseudomonas fluorescens were assayed by split root bioassay in tomato under glass house conditions. Application of P. fluorescens as seedling root dip increased the level of peroxidase (PO), polyphenol oxidase (PPO) and phenylalanine ammonia lyase (PAL) in tomato roots indicated the systemic resistance induced by the bacterial formulations against root knot nematode, Meloidogyne incognita. Among the four P.fluorescens isolates tested alone and in consortium with Bacillus subtilis, Bbv 57, consortial application of P. fluorescens, Pf 128 with B. subtilis, Bbv 57 recorded the highest enzymatic activity in tomato roots compared to other strains either alone or in consortium. Results indicated that P. fluorescens was capable of inducing systemic resistance against M. incognita in tomato by the accumulation of the defense enzymes like PO, PPO and PAL.

Effect of Removal of Garlic Mustard (Alliaria petiolata, Brassicaeae) on Arbuscular Mycorrhizal Fungi Inoculum Potential in Forest Soils

Garlic mustard (Alliaria petiolata), a biennial species, is considered to be among the most troublesome of the invasive plants in the Eastern Deciduous forest of North America. It has been shown to prevent or reduce mycorrhizal colonization of native herbaceous ground layer plants and trees in these forests. It is estimated that 70-90% or more of herbaceous native ground layer plant species form associations with arbuscular mycorrhizal fungi (AMF). Loss of the mycorrhizal association can reduce growth, reproductive success, and competitiveness of plant species. Using a corn root bioassay, we examined the effect of garlic mustard removal on the soil AMF mycorrhizal inoculum potential (MIP), in control plots and plots that had second-year garlic mustard removed annually for the past five years (2005-2009). Removal treatment plots had significantly (P = 0.0240, df = 28) greater MIP than control plots (25.72±2.26% and 18.29±2.04%, respectively). MIP was negatively correlated with cover of garlic mustard (r 2 = 0.1325, P < 0.05, df = 30), which accounted for 13.2% of the variation in MIP. Cover of native vegetation in removal treatment plots (104.50±2.6%) was greater than that of the control plots (95.14±3.66%), (P = 0.0236, df = 115). These results show that removal of garlic mustard results in an increase in soil MIP and cover of native species; however, there is not a complete loss of MIP associated with garlic mustard invasion. Following removal of garlic mustard, our data suggest that mycorrhizal plants recover more slowly than non-mycorrhizal species, apparently due to a delay in the establishment of a well-functioning mycorrhizal association. Our study is the first to demonstrate that the MIP of native soils and cover of native species increase following reduction in the cover of garlic mustard.

Evaluasi Pertumbuhan dan Hasil Beberapa Genotipe Cabai (Capsicum annuum L.) untuk Toleransi terhadap Cekaman Aluminium

The aim of this research was to evaluate growth and yield in response to aluminum (Al) stress in several genotypes of chilli ( Capsicum annuum L.). The research was conducted in University Farm of IPB, Cikabayan, Bogor from February to August 2006. Ultisol soils from Gajrug (Lebak, Banten) in polybag with pH 4.2, Al-dd 30.08 me/ 100 g and Al-saturated 83.81% was used in this experiment. Eight genotypes of chilli (4 tolerant and 4sensitive) were tested on two different conditions, in media without Al-stress condition (Al-saturated 0.77%) and wich Al-stress (Al-saturated 60.85%). Characters observed were plant height, shoot dry-weight, total number of fruits per plant, number of fruit harvested per plant, fruit dimension (length and diameter), fruit weight harvested per plant, and weight of each fruit. The results showed that there were differences of response to Al-stress among genotypes on the study. PBC 619 and Jatilaba were selected as tolerant genotypes, while Cilibangi 3 and Helm were sensitive. This evaluation of agronomic characters for tolerant and sensitive to Al-stress appeared very consistent with root bioassay results. Based on this result, the root bioassay method can be used to select chili genotypes for Al-stress tolerance. Key words: Growth and yield evaluation, aluminum stress, Capsicum annuum , ultisol soil.

Estimating the relative nutrient uptake from different soil depths in Quercus robur, Fagus sylvatica and Picea abies

The distribution of fine roots and external ectomycorrhizal mycelium of three species of trees was determined down to a soil depth of 55 cm to estimate the relative nutrient uptake capacity of the trees from different soil layers. In addition, a root bioassay was performed to estimate the nutrient uptake capacity of Rb+ and NH4+ by these fine roots under standardized conditions in the laboratory. The study was performed in monocultures of oak (Quercus robur L.), European beech (Fagus sylvatica L.) and Norway spruce [Picea abies (L.) Karst.] on sandy soil in a tree species trial in Denmark. The distribution of spruce roots was found to be more concentrated to the top layer (0-11 cm) than that of oak and beech roots, and the amount of external ectomycorrhizal mycelia was correlated to the distribution of the roots. The uptake rate of [86 Rb+] by oak roots declined with soil depth, while that of beech or spruce roots was not influenced by soil depth. In modelling the nutrient sustainability of forest soils, the utilization of nutrient resources in deep soil layers has been found to be a key factor. The present study shows that the more shallow-rooted spruce can have a similar capacity to take up nutrients from deeper soil layers than the more deeply rooted oak. The distribution of roots and mycelia may therefore not be a reliable parameter for describing nutrient uptake capacity by tree roots at different soil depths. (Less)

The response of ectomycorrhizal fungal inoculum to long-term increases in nitrogen supply

The inoculum of ectomycorrhizal (EM) fungi was examined in a 16 y long nitrogen fertilization experiment maintained in a temperate oak savanna. To measure EM fungal inoculum, bur oak seedlings were grown in three types of bioassays: (i) intact soil cores that measure inoculum such as spores, mycelia and mycorrhizal roots; (ii) resistant propagule bioassays that measure inoculum types resistant to soil drying; and (iii) previously mycorrhizal root bioassays that measure the ability of EM fungi to colonize new roots from mycorrhizal roots. Colonization of bur oak seedlings was characterized by morphotyping and where necessary by restriction analysis and internal transcribed spacer (ITS) sequencing. Fourteen morphotypes were found in intact soil core bioassays with species of Cortinarius, Cenococcum and Russula abundant. Five morphotypes were found in resistant propagule bioassays with Cenococcum, a thelephoroid morphotype and a Wilcoxina-like ascomycete abundant and frequent. In intact soil core bioassays total percent root colonization and number of morphotypes were not affected by N supply in 2000 and 2001. However the composition of EM fungi colonizing oak seedling roots was different with increased N supply such that Russula spp. (primarily Russula aff. amoenolens) were most abundant at the highest level of N supply. Dominant Russula spp. did not colonize any roots in resistant propagule bioassays but did colonize oak seedling roots from previously mycorrhizal roots. Results suggest that in this savanna N supply can influence the kinds of inoculum propagules present and thereby might affect the dynamics of ectomycorrhizal communities by differentially influencing reproductive and colonization strategies.

The response of ectomycorrhizal fungal inoculum to long-term increases in nitrogen supply

The inoculum of ectomycorrhizal (EM) fungi was examined in a 16 y long nitrogen fertilization experiment maintained in a temperate oak savanna. To measure EM fungal inoculum, bur oak seedlings were grown in three types of bioassays: (i) intact soil cores that measure inoculum such as spores, mycelia and mycorrhizal roots; (ii) resistant propagule bioassays that measure inoculum types resistant to soil drying; and (iii) previously mycorrhizal root bioassays that measure the ability of EM fungi to colonize new roots from mycorrhizal roots. Colonization of bur oak seedlings was characterized by morphotyping and where necessary by restriction analysis and internal transcribed spacer (ITS) sequencing. Fourteen morphotypes were found in intact soil core bioassays with species of Cortinarius, Cenococcum and Russula abundant. Five morphotypes were found in resistant propagule bioassays with Cenococcum, a thelephoroid morphotype and a Wilcoxina-like ascomycete abundant and frequent. In intact soil core bioassays total percent root colonization and number of morphotypes were not affected by N supply in 2000 and 2001. However the composition of EM fungi colonizing oak seedling roots was different with increased N supply such that Russula spp. (primarily Russula aff. amoenolens) were most abundant at the highest level of N supply. Dominant Russula spp. did not colonize any roots in resistant propagule bioassays but did colonize oak seedling roots from previously mycorrhizal roots. Results suggest that in this savanna N supply can influence the kinds of inoculum propagules present and thereby might affect the dynamics of ectomycorrhizal communities by differentially influencing reproductive and colonization strategies.

The development and reproduction of grape phylloxera (Daktulosphaira vitifoliae Fitch) strains of Hungarian vine-districts in root-bioassays

The root bioassay method allows for 4-6 week continuous observation of grape phylloxera feeding on the grape root. 10 Hungarian phylloxera strains were compared on susceptible Vitis vinifera cv. Chardonnay and the resistant rootstocks of V. berlandieri x V. riparia Teleki 5C and V. berlandieri x V. rupestris Georgikon 121 in in vitro observations. The strains originated from Villany and Eger (Figure 3) had higher reproduction on the root of V. berlandieri x V. riparia Teleki 5C (201 and 119 eggs) and were more aggressive than the others (average production 10 eggs). The continuous high level of survival, development and reproduction of the Eger strain on Teleki 5C (V. berlandieri x V. ripuria) until day 46 may be due to adaptation.

Changes in ectomycorrhizal communities and nutrient availability following prescribed burns in two upland pine–oak forests in the New Jersey pine barrens

Forests in the New Jersey pine barrens are frequently prescribed burned to reduce fuel loads and risk of wildfire. To acquire baseline data for effects of prescribed burns on ectomycorrhizal diversity and nutrient uptake, field studies were undertaken in two upland pine–oak forests in the New Jersey pine barrens subjected to different burn regimes. Ectomycorrhizal diversity was assessed by extraction of roots from soil cores and separation according to morphological characters. Nutrient availability to plant roots was measured using root bioassays. Relative to unburned plots, plots exposed to a fire at Greenwood, where burning was more frequent, had decreased total abundance of ectomycorrhizal tips, richness of ectomycorrhizal types, and Simpson's diversity in the L and F horizons, but increased Simpson's diversity in the deeper A horizon. At Lebanon, under a less frequent burn regime, richness was lower in whole cores and in the A horizon of burned versus unburned plots. Nitrogen, phosphorus, and potassium uptake by root bioassays indicated higher field availability of nutrients to roots in burned plots than roots in unburned plots, indicating a fertilization effect of the fire. Prescribed burning primarily impacted ectomycorrhizal community structure in the L and F horizons at these sites. Changes in function of ectomycorrhizae (nutrient uptake) in response to the burns was associated with decreased ectomycorrhizal diversity.

Ectomycorrhizal fungal biomass in roots and uptake of P from apatite by Pinus sylvestris seedlings growing in forest soil with and without wood ash amendment.

Forest soil from an experimental Norway spruce forest with four levels of wood ash addition (0, 1, 3 and 6 tonnes ha(-1)) was used to inoculate pine (Pinus sylvestris) seedlings with indigenous ectomycorrhizal (EM) fungi. Uptake of 32P and 86Rb in a root bioassay was used to estimate the demand for P and K by seedlings grown in the different soils. Utilisation of P from apatite was tested in a laboratory system where uptake by the ectomycorrhizal mycelium was separated from uptake by roots. The demand for P and K in the seedlings was similar regardless of the ash treatment. Variation in EM levels, estimated as fungal biomass (ergosterol) in roots, was large in the different soils, but not related to ash addition. Uptake of P from apatite was, on average, 23% of total seedling P and was not related to EM levels. It was concluded that the improved P uptake from apatite by EM fungi found in earlier studies is probably not a general phenomenon among EM fungi. The small effect of ash addition on EM levels and P uptake suggests that addition of granulated wood ash is a forest management treatment that will have only minor influence on ectomycorrhizal symbiosis.

Systemic Resistance Induced by Trichoderma hamatum 382 in Cucumber Against Phytophthora Crown Rot and Leaf Blight.

Phytophthora root rot, crown rot, leaf and stem blight, and fruit rot of cucumber can cause serious losses, and are difficult to control. Although composts can be used successfully for control of Phytophthora root rots, little is known about their effects on Phytophthora diseases of aboveground plant parts. This research shows that the severity of Phytophthora root and crown rot of cucumber caused by Phytophthora capsici was suppressed significantly in cucumber transplants produced in a composted cow manure-amended mix compared with those in a dark sphagnum peat mix. In split root bioassays, Trichoderma hamatum 382 (T382) inoculated into the compost-amended potting mix significantly reduced the severity of Phytophthora root and crown rot on paired roots in the peat mix. This effect did not differ significantly from that provided by a drench with benzothiadiazole (BTH) or mefenoxam (Subdue MAXX). Based on area under disease progress curves, T382 also significantly reduced the severity of Phytophthora leaf blight in transplants produced in the compost mix compared with controls not inoculated with T382. Efficacy of T382 did not differ significantly from that provided by a drench with BTH. T382 re-mained spatially separated from the pathogen in plants in both the split root and leaf blight bioassays, suggesting that these effects were systemic in nature.

Suppression of root-knot disease by Pseudomonas fluorescens CHA0 in tomato: importance of bacterial secondary metabolite, 2,4-diacetylpholoroglucinol

The antimicrobial metabolites 2,4-diacetylphloroglucinol (2,4-DAPG) and pyoluteorin contribute to the ability of Pseudomonas fluorescens strain CHA0 to control plant diseases caused by soil-borne pathogens. P. fluorescens strain CHA0 and its derivatives CHA89 (antibiotics-deficient) and CHA0/pME3424 (antibiotics overproducing) were investigated as potential biocontrol agents against Meloidogyne javanica the root-knot nematode. Exposure of root-knot nematode to culture filtrates of P. fluorescens under in vitro conditions significantly reduced egg hatch and caused substantial mortality of M. javanica juveniles. Nutrient broth yeast extract (NBY) medium amended with 2% (w/v) glucose or 1 mM EDTA markedly repressed hatch inhibition activity of the strain CHA0 but not that of CHA0/pME3424 or CHA89. On the other hand, NBY medium amended with glucose significantly enhanced nematicidal activity of the strain CHA0/pME3424. Neither glucose nor EDTA had an influence on the nematicidal activity of the strains CHA0 and CHA89. Under in vitro conditions, antibiotic overproducing strain CHA0/pME3424 and CHA0 expressed phl‘-’lacZ reporter gene but strain CHA89 did not. Expression of the reporter gene reflects actual production of DAPG. In general, CHA0/pME3424 expressed reporter gene to a greater extent compared to its wild type counterpart CHA0. Regardless of the bacterial strains, reporter gene expression was markedly enhanced when NBY medium was amended with glucose but EDTA had no such effect. A positive correlation between the degree of juvenile mortality and extent of phl‘-’lacZ reporter gene expression was also observed in vitro. Strain CHA0 produced zones of 4–6 mm on MM medium containing gelatin while strain CHA0/pME3424 and CHA89 did not. When MM medium containing gelatin was amended with 2% glucose of 1 mM EDTA size of haloes produced by the strain CHA0 reduced to 2 mm. Under glasshouse conditions aqueous cell suspension of the strains CHA0 or CHA0/pME3424 at various inoculum levels (10 7, 10 8 or 10 9 cfu ml −1) significantly reduced root-knot development. CHA89 caused significant reduction in galling when applied at 10 9 cfu ml −1. To better understand the mechanism of nematode suppression, split root bioassay was performed. Split-root experiments, that guarantee a spatial separation of inducing agent and a challenging pathogen, showed that soil treatment of one half of the root system with cell suspension of CHA0 or CHA0/pME3424 resulted in a significant systemic induced resistance leading to reduction of M. javanica infection of tomato roots in the non-baterized nematode treated half. The results clearly suggest that the antibiotic 2,4-DAPG from P. fluorescens CHA0 act as the inducing agents of systemic resistance in tomato roots. Populations of CHA0 and its derivatives declined progressively by 10-fold between first and fourth harvests (0–21 days after inoculation). However, bacterial populations increased at final harvest (28 days after application).