Effects of Daylength on Endogenous Gibberellins in Solanum andigena III. Gibberellin Production by the Leaves

Using an agar diffusion technique, it was found that leaves from potato plants growing under long days produced more gibberellin-like substances than did leaves from plants growing under short days. Short day plants irradiated with red light during the long dark period and harvested during the ensuing light period, contained levels of gibberellin-like substances approximating those found in long day grown plants. Red irradiation during the long dark period also resulted in an increase in gibberellin production in short day plants. Four zones of gibberellin-like activity (A, B, C, D) were separated by thin layer chromatography in extracts from potato leaves. Red light treatment reduced the levels of peak D and brought about a concomitant increase in the levels of peak A.     

Grapevine downy mildew control using reduced copper amounts in organic viticulture

Copper is an essential natural micronutrient. However, copper used as a plant protection product may have long-term consequences due to its accumulation in the soil. Limitations on copper use have therefore been defined in organic farming (Regulation EC 889/2008). In the light of new developments and evidence, the European Commission has planned to assess whether further restrictions are needed in the quantities of copper permitted. A two-year field trial was therefore set up with new copper formulations to evaluate the possibility of reducing the copper quantities applied with treatments and consequently to reduce copper soil residues. Plots were prepared, each containing 12 plants and repeated four times in randomized blocks. The test organism was Plasmopara viticola (Berk. and M.A. Curtis) Berl. and De Toni. Cupric formulations characterised by a low metallic content (Glutex CU 90 and Labicuper) were tested in comparison with a reference product (standard) and an untreated control. Evaluations of treatments were carried out periodically on 100 leaves and 100 bunches for each replicate. Data obtained were subjected to statistical analysis. Chemical analyses were performed to determine copper residues on leaves, grapes and soil. Samplings of leaves and grapes were carried out for each replicate. Soil samples were taken from 0-20 cm and 20-40 cm depth. Total copper was determined using spectrophotometry in atomic absorption by acetylene-air flame (FAAS at lambda = 324.8 microm). The results showed that the tested products were effective in controlling downy mildew with a lower copper dosage than with the cupric formulations used as a standard. Glutex CU 90 formulation led to an annual input of copper that was a little more than a third compared to the standard and Labicuper about a fifth or a sixth. At harvest, copper levels in grapes were much lower than RML (fixed at 50 mg/kg). With regard to the impact of cupric treatments on organic vineyard soil, no statistically significant differential increase in Cu residue was observed in soil between tested products versus untreated control. In conclusion, the environmental impact of copper in organic viticulture could be minimized through the new cupric formulations developed by agrochemical companies.     

Ribonuclease and proteinase activities in potato leaves immunized against Phytophthora infestans

Local treatment of potato plants with arachidonic acid caused a repeated increase of ribonuclease and proteinase activities in leaves that were directly sprayed with this inducer. Immunization caused no significant systemic changes in the enzyme activity of other potato leaves. However, leaves above the induction zone in the first days after the challenge were found to have a pronounced ribonuclease activity increase in comparison to that gradually rising with disease development in inoculated leaves from plants that were not induced. The proteolytic activity in the leaves of immunized plants after the challenge was decisively on a lower level than that in the leaves of noninduced plants subjected to inoculation.     

Arbuscular mycorrhiza‐mediated resistance in tomato against Cladosporium fulvum‐induced mould disease

Root colonization with arbuscular mycorrhizal fungi (AMF) enhances plant resistance particularly against soil‐borne pathogenic fungi. In this study, mycorrhizal inoculation with Glomus mosseae (Gm) significantly alleviated tomato mould disease caused by the air‐borne fungal pathogen, Cladosporium fulvum (Cf). The disease index (DI) in local leaves (receiving pathogen inoculation) and systemic leaves (just above the local leaf without pathogen inoculation) was 36.4% and 11.7% in mycorrhizal plants, respectively, whereas DI was 59.6% and 36.4% in the corresponding leaves of AMF non‐inoculated plants, after 50 days of Gm inoculation, corresponding to 15 days after Cf inoculation by leaf infiltration. Foliar spray inoculation with Cf also revealed that AMF pre‐inoculated plants had a higher resistance against subsequent pathogen infection, where the DI was 41.3% in mycorrhizal plants vs. 64.4% in AMF non‐inoculated plants. AMF‐inoculated plants showed significantly higher fresh and dry weight than non‐inoculated plants under both control (without pathogen) and pathogen treatments. AMF‐inoculated plants exhibited significant increases in activities of superoxide dismutase and peroxidase, along with decreases in levels of H₂O₂ and malondialdehyde, compared with non‐inoculated plants after pathogen inoculation. AMF inoculation led to increases in total chlorophyll contents and net photosynthesis rate as compared with non‐inoculated plants under control and pathogen infection. Pathogen infection on AMF non‐inoculated plants led to decreases in chlorophyll fluorescence parameters. However, pathogen infection did not affect these parameters in mycorrhizal plants. Taken together, these results indicate that AMF colonization may play an important role in plant resistance against air‐borne pathogen infection by maintaining redox poise and photosynthetic activity.     

Release of lipoxygenase products and monoterpenes by tomato plants as an indicator of Botrytis cinerea-induced stress

Changes in emission of volatile organic compounds (VOCs) from tomato induced by the fungus Botrytis cinerea were studied in plants inoculated by spraying with suspensions containing B. cinerea spores. VOC emissions were analysed using on-line gas chromatography–mass spectrometry, with a time resolution of about 1 h, for up to 2 days after spraying. Four phases were delimited according to the starting point and the applied day/night rhythm of the experiments. These phases were used to demonstrate changes in VOC flux caused by B. cinerea infestation. Tomato plants inoculated with B. cinerea emitted a different number and amount of VOCs after inoculation compared to control plants that had been sprayed with a suspension without B. cinerea spores. The changes in emissions were dependent on time after inoculation as well as on the severity of infection. The predominant VOCs emitted after inoculation were volatile products from the lipoxygenase pathway (LOX products). The increased emission of LOX products proved to be a strong indicator of a stress response, indicating that VOC emissions can be used to detect plant stress at an early stage. Besides emission of LOX products, there were also increases in monoterpene emissions. However, neither increased emission of LOX products nor of monoterpenes is specific for B. cinerea attack. The emission of LOX products is also induced by other stresses, and increased emission of monoterpenes seems to be the result of mechanical damage induced by secondary stress impacts on leaves.     

Inhibitory effects of some invasive alien species leaf extracts against tomato (Lycopersicon esculentum Mill.) bacterial wilt (Ralstonia solanacearum)

The inhibitory activity of selected invasive alien species leaf extracts against tomato bacterial wilt was studied in vitro and greenhouse. In vitro inhibitory activity was undertaken in a disc diffusion sensitivity test. Furthermore, aqueous extracts of Eichhorina crassipes, Mimosa diplotricha and Lantana camara and methanolic extract of Prosopis juliflora which showed better inhibitory effect in vitro were evaluated against R. solanacearum in greenhouse on tomato. The plant extracts were applied and evaluated at three different times of application (at the time of inoculation; two days before pathogen inoculation; and two days after pathogen inoculation). The result showed that most of the treatment combinations significantly reduce disease incidence and area under disease progress curve and increase biomass of tomato plants, but the effectiveness of tested plant species depends on the type of plant species and application time. The application of plant extracts at the time of pathogen inoculation resulted in highest reduction of disease development on tomato plants. Aqueous extract of E. crassipes was found to be the most effective plant extract in disease suppression and increase above-ground biomass compared to inoculated control. The study revealed that the tested plant species have a potential of inhibiting the development of bacterial wilt on tomato.     

Effects of Plant Nutrition on Susceptibility of Potato Leaves to Phytophthora infestans

A complete NPK fertilizer applied at planting, but not at later times, influenced the resistance of potato leaves to infection by Phytophthora infestans and the subsequent growth of lesions. Lesion size (7 days after inoculation) and lesion growth rate (between 7 and 9 days after inoculation) increased linearly with increasing fertilizer rates. Increasing rates of phosphate or potash, applied separately, did not significantly affect the development of blight lesions but a linear increase in lesion size was associated with increasing application rates of nitrogen.     

Induction of Growth Increase and Systemic Resistance to Exserohilum turcicum in Maize by Foliar Spray of Phosphates

A single foliar spray of 0.1 M solution of phosphate salts on the upper side of maize (cv. ‘jubilee') leaves 1, 2 and 3 at the 5–6 fully–expanded leaf-stage, 2-4 h before inoculation induced systemic resistance to northern leaf blight caused by Exserohilum turcicum. Nine days after challenge, protection was demonstrated by 91% reduction in the size and 69% in the number of E. turcicum lesions developing on leaves 4, 5, 6 and 7. Appropriate mixing of phosphate solution with KOH revealed that the level of protection was not necessarily dependent on the pH of the solution. The size and number of lesions decreased to 62% and 56%, respectively, 12 days after challenge. There was no damage or chlorotic stipling on the induced leaves (1, 2, and 3) as a result of the phosphate spray. There were no significant differences in the reduction in the number or size of the lesions obtained when the foliar spray was applied 2-4 h or 1, 3, 6, 8, or 10 days before inoculation. One foliar spray ofK₂HPO₄ on leaves 1, 2 and 3 in these intervals before inoculation, remarkably stimulated growth of inoculated plants, which was expressed by several parameters. The possible dual use of phosphate salts as foliar fertilizers and as resistance-inducing agents is discussed     

Methylobacterium sp. resides in unculturable state in potato tissues in vitro and becomes culturable after induction by Pseudomonas fluorescens IMGB163

Aims:  To induce growth of endophytic bacteria residing in an unculturable state in tissues of in vitro -grown potato plantlets. To isolate and identify the induced bacteria and to localize the strains in tissues of in vitro -grown potato plantlets.
Methods and Results:  The inoculation of in vitro -grown potato plants with Pseudomonas fluorescens IMBG163 led to induction of another bacterium, a pink-pigmented facultative methylotroph that was identified as Methylobacterium sp . using phylogenetic 16S rDNA approach . Two molecular methods were used for localizing methylobacteria in potato plantlets: PCR and in situ hybridization (ISH/FISH). A PCR product specific for the Methylobacterium genus was found in DNA isolated from the surface-sterilized plantlet leaves. Presence of Methylobacterium rRNA was detected by ISH/FISH in leaves and stems of inoculated as well as axenic potato plantlets although the bacterium cannot be isolated from the axenic plants.
Conclusion:  Methylobacterium sp. resides in unculturable state within tissues of in vitro -grown potato plants and becomes culturable after inoculation with P. fluorescens IMBG163.
Significance and Impact of the Study:  In order to develop endophytic biofertilizers and biocontrol agents, a detailed knowledge of the life-style of endophytes is essential. To our knowledge, this is the first report on increase of the culturability of endophytes in response to inoculation by nonpathogenic bacteria.     

Evaluation of four whole-plant inoculation methods to analyze the pathogenicity of Erwinia amylovora under quarantine conditions

Four methods were tested to assess the fire-blight disease response on grafted pear plants. The leaves of the plants were inoculated with Erwinia amylovora suspensions by pricking with clamps, cutting with scissors, local infiltration, and painting a bacterial suspension onto the leaves with a paintbrush. The effects of the inoculation methods were studied in dose-time-response experiments carried out in climate chambers under quarantine conditions. A modified Gompertz model was used to analyze the disease-time relatiobbnships and provided information on the rate of infection progression (rg) and time delay to the start of symptoms (t0). The disease-pathogen-dose relationships were analyzed according to a hyperbolic saturation model in which the median effective dose (ED50) of the pathogen and maximum disease level (ymax) were determined. Localized infiltration into the leaf mesophile resulted in the early (short t0) but slow (low rg) development of infection whereas in leaves pricked with clamps disease symptoms developed late (long t0) but rapidly (high rg). Paintbrush inoculation of the plants resulted in an incubation period of medium length, a moderate rate of infection progression, and low ymax values. In leaves inoculated with scissors, fire-blight symptoms developed early (short t0) and rapidly (high rg), and with the lowest ED50 and the highest ymax.     

Salicylic acid induced systemic resistant on peanut against Alternaria alternata

Abstract

The effect of Salicylic Acid (SA) in inducing resistance in groundnut plants against Alternaria alternata was investigated. Foliar application of SA at the concentration of 1 mM significantly reduced the leaf blight disease intensity and increased the pod yield under glasshouse conditions. Changes in the activities of phenylalanine ammonium lyase, chitinase β-1,3 glucanase and in phenolic content on groundnut after application of SA and inoculation with A. alternate were studied. In SA-treated leaves (plants) an increase in phenolic content was observed five days after challenge inoculation with A. alternata in groundnut plants pretreated with SA. There was a marked increase in chitinase and pathogen inoculation in SA-treated leaves. In chitinase, β-1,3 glucanase activities were observed in response to plants with an increase in SA treated leaves. Foliar applications of SA-induced in peroxidase and polyphenol oxidase activities were observed upon challenge inoculation with pathogen.     

Factors affecting the detection of potato leafroll virus in potato foliage by enzyme-linked immunosorbent assay

Using antiserum globulins that reacted only weakly with plant materials, potato leafroll virus (PLRV) at 10 ng/ml was detected consistently by enzyme-linked immunosorbent assay (ELISA). The reaction with PLRV particles was slightly impaired in potato leaf extracts that were diluted less than 10^-1 but not at greater dilutions. Antiserum globulins that reacted more strongly with plant materials could be used satisfactorily for coating microtitre plates but were unsuitable for conjugating with enzyme. The detection end-point of PLRV, in leaf sap of potato cv. Cara plants grown from infected tubers in the glasshouse, was about 10^-2 and the virus was reliably detected in extracts of composite samples of one infected and 15 virus-free leaves. PLRV concentration was much less in extracts of roots or stolons than in leaf extracts. The virus was detected in infected leaves of all 27 cultivars tested. PLRV was readily detectable 2 wk before symptoms of secondary infection developed in field-grown plants of cv. Cara and Maris Piper and remained so for at least 5 wk. Its concentration was slightly greater in old than in young leaves and was similar to that in glasshouse-grown plants. In field-grown plants of cv. Maris Piper with primary infection, PLRV was detected in tip leaves 21–42 days after lower leaves were inoculated by aphids; in some shoots it later reached a concentration, in tip leaves, similar to that in leaves with secondary infection. Symptoms of primary infection developed in the young leaves of some infected shoots but were inconspicuous and were not observed until at least a week after PLRV was detected by ELISA.     

Effects of Extracellular Extracts from Leaves on the Tuberization of Cuttings of Potato (Solanum tuberosum L.)

Extracellular extracts from leaves were applied to moderately induced cuttings of potato. The more inductive cycles the plants from which the leaf extracts were obtained had experienced, the lower the percentages of tuberization in thcuttings and the lower the tuber yields on the central buds.     

Induced Systemic Resistance to Cercospora Leaf Spot of Carrot by Inoculation with Cercospora carotae

The possibility of inducing systemic disease resistance in carrot leaves was investigated with the pathogen Cercospora carotae as the agent for both induction and challenge. The inoculation of plants with C. carotae induced resistance in newly emerged leaves challenged 23 days later. There was an optimum inoculum concentration of 3 × 10⁴ conidia ml^-1 for inducing resistance, the response decreasing with higher concentrations. Infection of carrot leaves by C. carotae could thus reduce susceptibility to subsequent infections by foliar pathogens later in the growing season. However, the inoculation of leaves with C. carotae did not increase the resistance of roots after harvest when they were wounded and challenged with Botrytis cinerea .     

Revealing the importance of meristems and roots for the development of hypersensitive responses and full foliar resistance to Phytophthora infestans in the resistant potato cultivar Sarpo Mira

The defence responses of potato against Phytophthora infestans were studied using the highly resistant Sarpo Mira cultivar. The effects of plant integrity, meristems, and roots on the hypersensitive response (HR), plant resistance, and the regulation of PR genes were analysed. Sarpo Mira shoots and roots grafted with the susceptible Bintje cultivar as well as non-grafted different parts of Sarpo Mira plants were inoculated with P. infestans. The progress of the infection and the number of HR lesions were monitored, and the regulation of PR genes was compared in detached and attached leaves. Additionally, the antimicrobial activity of plant extracts was assessed. The presented data show that roots are needed to achieve full pathogen resistance, that the removal of meristems in detached leaves inhibits the formation of HR lesions, that PR genes are differentially regulated in detached leaves compared with leaves of whole plants, and that antimicrobial compounds accumulate in leaves and roots of Sarpo Mira plants challenged with P. infestans. While meristems are necessary for the formation of HR lesions, the roots of Sarpo Mira plants participate in the production of defence-associated compounds that increase systemic resistance. Based on the literature and on the presented results, a model is proposed for mechanisms involved in Sarpo Mira resistance that may apply to other resistant potato cultivars.     

A novel indicator plant to test the hypersensitivity of phytopathogenic bacteria

Hypersensitive response is an important, definitive test to separate plant pathogenic bacteria from saprophytes. The current standard protocol uses tobacco and four o'clock plants as indicators for Gram negative and Gram positive phytopathogenic bacteria, respectively, and inoculation is accomplished by infiltrating bacterial suspensions into intact leaves. Both plants, especially the four o'clock, have thin leaves which make inoculation difficult, sometimes leading to inaccurate tests. Here we propose the use of Sedum hybridum as an alternative indicator plant. Sedum plants are readily available, easy to propagate and fast growing. Their leaves are much thicker, thus infiltration of a bacterial suspension is easier compared to tobacco and four o'clock. Additionally, sedum plants can be used universally to test both Gram negative and Gram positive phytopathogenic bacteria.