The Induced Resistance Response of Carrot Root Slices to Heat-killed Conidia and Cell-free Germination Fluid of Botrytis cinerea Pers. ex Pers.: 2. Nuclear Migration, Nucleolar Volume and Uptake of Tritiated Uracil

Sixty-eight per cent of nuclei in the cells of the upper fourlayers of carrot slices treated with heat-killed conidia ofBotrytis cinerea for 6 h followed by inoculation with live sporesfor 18 h, migrated to the cell face nearest to the treated surface,compared with 46 per cent in cells of control slices showinga wound-healing response only. Nucleolar volumes in the surfacecell layers of control slices increased from a mean of 1.0 µm³to 3.8 µm³ over 24 h, and in ‘induced’ slicesto 7.28 µm³. Using a 40 min pulse of [5–³H]uracil,there was an increase within 15 h of slicing in the number oflabelled nuclei in cells from control slices undergoing healing.Within 8 h after treatment of slice surfaces with heat-killedconidia, there was an accelerated incorporation of label into‘nuclear’ RNA. Slices from roots cold-stored for12 months failed to show an induction response and nucleolarvolumes did not increase more than in control slices. Theseresults are discussed in relation to active defence mechanismsin plant tissue. Botrytis cinerea, carrot, induced resistance, nuclear migration, nucleolar volume, RNA incorporation     

Activité Fongicide du N-cetyl N,N,N-triméthylammonium Contre Botrytis cinerea Pers.

Fungicidal activity of N-cetyl N,N,N-trimethyl ammonium chloride to Botrytis cinerea Pers. Grey mould of grapes caused by Botrytis cinerea is the most important phytopatho-logical problem in swiss vineyards, since the appearance of resistant strains to dicarboxi-mides. Research for new chemicals is necessary. One compound, a cationic surfactant, the N-cetyl N,N,N-trimethyl ammonium chloride (CTAC), is very active in vitro against Botrytis cinerea. This product disturbs lethally the plasma membrane of conidia.     

Endogene Hyphen bei Botrytis cinerea Pers.

Endohyphae in Botrytis cinerea Pers. Genesis, following differentiation and significance for pathogenesis The formation of endohyphae (secondary hyphae) in different cell types of Botrytis cinerea Pers. (6 isolates) is described. The following differentiation possibilities of these endohyphae within and—after egress—outside their cells of origin are presented. The influence of external factors (substrate, temperature, humidity, O₂-concentration) on these processes is considered. The significance of the endohyphae for pathogenesis is mainly due to their direct infectivity as well as the ability to develop normal mycelium.     

Endogenous Elicitor Activity from Damaged Carrot Root Tissue and Induced Resistance to Botrytis cinerea

A peptide-containing fraction (mol. wt. c 5000 Da) extractedfrom carrot root tissues damaged by slicing or freeze-thawing,induced active defence mechanisms in carrot slices against Botrytiscinerea. Endogenous elicitor activity was present in homogenatesof fresh and freeze-thawed tissues and those treated with germinationfluid, but was absent in autoclaved tissue. The detection ofthe elicitor in homogenates within 2h of treatment, suggeststhat it is released or activated during the early stages ofcell damage. Botrytis cinerea, carrot, induced resistance, endogenous elicitor     

Resistenz verschiedener Kultur- und Wildtomatenpflanzen (Lycopersicon spp.) gegenüber Botrytis cinerea Pers.

Resistance of different cultivated and wild tomato plants (Lycopersicon spp.) to Botrytis cinerea Pers. 20 provenances of different cultivated and wild tomato plants (Lycopersicon spp.) were screened for resistance to Botrytis cinerea Pers. using an in vitro-leaf necrosis test. The Botrytis resistance decreased with increasing age of the leaves corresponding to their insertion height (relative youth resistance respectively senescence susceptibility). The 6 B. cinerea-isolates used for inoculation differed significantly in virulence. With increasing inoculum age a virulence reduction of the various B. cinerea-isolates occurred. Within the investigated test plant collection 2 wild species –L. columbianum and L. hirsutum– proved to be resistant in each stage of development to all B. cinerea-isolates and additionally showed field resistance.     

Evidence for a constitutive cytoplasmic cutinase in ungerminated conidia of Botrytis cinerea Pers.: Fr.

Cytoplasmic soluble proteins from ungerminated conidia of Botrytis cinerea exhibited cutinase activity, while cell wall binding proteins lacked this activity. Cutinase activity in proteins extracted from cell walls and cytoplasm of ungerminated conidia of Botrytis cinerea was determined using p-nitrophenyl butyrate (PNB) and TLC analysis of products derived from hydrolysis of [³H]cutin. Treatment of conidia with indoxyl acetate, a substrate indicative of non-specific esterase and cutinase activity, also gave a positive reaction in the cytoplasm of ungerminated conidia. The possible role of a putative constitutive cutinase in the cytoplasm of conidia in the early stages of infection of plants by B. cinerea is discussed.     

The Freezing Response of an Arctic Cushion Plant, Saxifraga caespitosa L.: Acclimation, Freezing Tolerance and Ice Nucleation

Cold hardiness in actively growing plants of Saxifraga caespitosaL., an arctic and subarctic cushion plant, was examined. Plantscollected from subarctic and arctic sites were cultivated ina phytotron at temperatures of 3, 9, 12 and 21 °C undera 24-h photoperiod, and examined for freezing tolerance usingcontrolled freezing at a cooling rate of 3–4 °C eitherin air or in moist sand. Post-freezing injury was assessed byvisual inspection and with chlorophyll fluorescence, which appearedto be well suited for the evaluation of injury in Saxifragaleaves. Freezing of excised leaves in moist sand distinguishedwell among the various treatments, but the differences werepartly masked by significant supercooling when the tissue wasfrozen in air. Excised leaves, meristems, stem tissue and flowerssupercooled to –9 to –15 °C, but in rosettesand in intact plants ice nucleation was initiated at –4to –7 °C. The arctic plants tended to be more coldhardy than the subarctic plants, but in plants from both locationscold hardiness increased significantly with decreasing growthtemperature. Plants grown at 12 °C or less developed resistanceto freezing, and excised leaves of arctic Saxifraga grown at3 °C survived temperatures down to about –20 °C.Exposure to –3 °C temperature for up to 5 d did notsignificantly enhance the hardiness obtained at 3 °C. Whenwhole plants of arctic Saxifraga were frozen, with roots protectedfrom freezing, they survived –15 °C and –25°C when cultivated at 12 and 3 °C, respectively, althougha high percentage of the leaves were killed. The basal levelof freezing tolerance maintained in these plants throughoutperiods of active growth may have adaptive significance in subarcticand arctic environments. Saxifraga caespitosa L., arctic, chlorophyll fluorescence, cold acclimation, cushion plant, freezing stress, freezing tolerance, ice nucleation, supercooling     

Screening of wild accessions resistant to gray mold (Botrytis cinerea Pers.) in Lycopersicon

Tomato gray mold (Botrytis cinerea Pers.) is a common disease worldwide, and often causes serious production loss by infecting leaves, stems, flowers and fruits. Presently, no resistant cultivars are available. To find new breeding materials for gray mold resistance, assessment for resistance of the leaflet and stem in six tomato cultivars, 44 wild tomato accessions and a Solanum lycopersicoides accession was performed. Although no correlation was observed (r=−0.127^ns) between resistance of the leaflet and the stem, L. peruvianum LA2745, L. hirsutum LA2314 and L. pimpinellifolium LA1246 showed high resistance both in the leaflet and in the stem. Particularly, in the leaves of LA2745, no lesions were observed even more than two weeks after the inoculation with conidia, and F1s between a cultivated tomato and LA2745 also showed high resistance as observed in LA2745. From these results, LA2745 is thought to be a promising material for breeding gray-mold resistant cultivars.     

A Laboratory Leaf Assay of Carrot Susceptibility to Botrytis cinerea

Laboratory experiments were conducted to develop an assay for evaluation of carrot (Daucus carota L.) leaf reaction to Botrytis cinerea Pers. ex Pers. The detached carrot leaflets were inoculated with the colonized agar plugs attached to the cut surface of the midrib or by spraying conidial suspension. The estimation of the infected leaflet area expressed as the area under the disease progress curve enabled the discrimination between carrot genotypes differing in their susceptibility to the pathogen. Evaluation based on the measurement of the length of the midrib lesion did not allow such differentiation. The inoculation of leaf surface with suspension of conidia was less reliable and the development of the disease symptoms was not reproducible. The established assay using colonized plugs enables a fast assessment of carrot leaf susceptibility to grey mould and can be particularly useful for a non‐destructive, preliminary evaluation of precious and limited source materials.     

Cold Acclimation, Freezing Resistance and Protein Synthesis in Alfalfa (Medicago sativaL. cv. Saranac)

Mohapatra, S. S., Poole, R. J. and Dhindsa, R. S. 1987. Coldacclimation, freezing resistance and protein synthesis in alfalfa(Medicago sativa L. cv. Saranac).—J. exp. Bot. 38: 1697–1703. Changes in freezing resistance (percent survival at —10°C), pattern of protein synthesis and translatable mRNApopulation during cold acclimation of alfalfa (Medicago sativaL. cv. Saranac) have been examined. Two days of cold acclimationat 4 °C increased freezing resistance from about 6% to 40%,protein content by 200% and total RNA content by 100%. Acclimationfor longer periods did not cause further increases in freezingresistance, protein content or RNA content. Examination of proteinchanges by sodium dodecyl sulphate-polyacrylamide gel electrophoresis(SDS-PAGE) coupled with protein staining, and by fluorographyof in vivo labelled proteins separated by SDS-PAGE, showed thatseveral proteins are increasingly or newly synthesized duringcold acclimation. Analysis of in vitro translation productsby SDS-PAGE and fluorography shows changes in the populationof translatable mRNAs. It is concluded that in this varietyof alfalfa cold acclimation for only 2 d is sufficient to confermaximum freezing resistance, and that changes in proteins duringcold acclimation are regulated most probably at the transcnptionallevel. Key words: Freezing resistance, protein synthesis, cold acclimation, SDS-PAGE, Medicago sativa L.     

Kugelige, umhüllte Mikrokonidien-Aggregate als Überdauerungs- und Verbreitungseinheiten von Botrytis cinerea Pers.

Spherical, coated microconidia-aggregates as survival and dispersal units of Botrytis cinerea Pers. As a reaction to longer lasting unfavourable growth conditions Botrytis cinerea is able to form spherical microconidia-aggregates enclosed in a protective coat. Genesis, structure and occurrence of these so far unknown microconidia-aggregates are described and their function as survival and dispersal units of the fungus is discussed.     

Local and systemic protection of poinsettia (Euphorbia pulcherrima Willd.) against Botrytis cinerea Pers. induced by benzothiadiazole

Benzothiadiazole (BTH) was found to be highly effective in increasing resistance of two poinsettia cultivars — ‘Coco White’ and ‘Malibu Red’, moderately susceptible to the fungus Botrytis cinerea. BTH applied at a concentration of 0.3 mM on the discs cut out from the leaves of these poinsettia cultivars reduced disease symptoms by more than 60 % in comparison to the control discs treated with water and exposed to infection. It was also observed that the applied inducer at a concentration of 0.03 and 0.3 mM had a favourable influence on the increase of poinsettia systemic resistance of SAR type (systemic acquired resistance). The effectiveness of BTH was much less when disease development was examined on detached leaves (a 20 % reduction of lesion area) in comparison with a pronounced inhibition of grey mould development on intact leaves of previously induced plants (a 80 % protection of intact plants). Benzothiadiazole in the concentration range from 0.03 to 1.4 mM added to in vitro agar medium was not found to have an inhibitory influence on Botrytis cinerea mycelium growth and sporulation.     

Action Spectrum for Photosporogenesis in Botrytis cinerea Pers. ex Fr

An action spectrum for photoinduced conidium formation in the fungus Botrytis cinerea Pers. ex Fr. was determined by exposing colonies to monochromatic radiation obtained from a diffraction grating monochromator. Wavelengths longer than 355 nm were ineffective even after exposures of 8 hours at intensities of 4,068 to 8,276 erg per cm² per second. Colonies were exposed at 22.5 C from 226 to 355 nm at about 5 nm intervals. Three prominent peaks of effectiveness occurred at about 231, 268, and 283 nm, and there was also a minor peak at about 303 nm. The most effective wavelength for inducing sporulation was 231 nm; it was 25% more effective than the second most effective wavelength (283 nm).     

In vivo-Untersuchungen zur Entstehung und Funktion der Chlamydosporen von Botrytis cinerea Pers. am Wirt-Parasit-System Fuchsia hybrida – B. cinerea

In vivo-investigations on the formation and function of chlamydospores of Botrytis cinerea Pers. in the host-parasite-system Fuchsia hybrida–B. cinerea On naturally with Botrytis cinerea Pers. infected and artificially inoculated outdoor- and greenhouse-plants of Fuchsia hybrida the extra- and intramatrical formation of the B. cinerea- chlamydospores was investigated. The chlamydospores served 1. as structures of survival, which were tested with regard to their tolerance of drought, nutrient- and oxygen-deficiency, attack by bacteria and pH-requirements. 2. The chlamydospores represented dispersal units, which were capable of germination. 3. The chlamydospores could function as structures of infection, because after chlamydospore germination the outgrowing mycelium – either directly or after production of macroconidia – could serve as secondary inoculum and start new infections.     

Changes in Tissue Freezing in Senecio vulgaris infected by Rust (Puccinia lagenophorae)

Freezing of healthy and rust (Puccinia lagenophorae) infectedleaves of Senecio vulgaris was compared calorimetrically bythermal analysis. In fully expanded leaves the threshold freezingtemperature was in the range –6.8 to –8.4 °Cin controls but –3.0 to –5.1 °C in leaves withsporulating rust sori. Comparable values in expanding leaveswere –5.0 to –8.9 °C and –3.9 to –6.7°C for healthy and rusted tissues, respectively. The bulktissue freezing point was between –1.0 and –4.0°C in both fully expanded and expanding healthy leaves,and was increased by infection by between +0.2 and 2.5 °C.Whereas healthy leaves supercooled by 3.1–5.8 °C,rusted leaves supercooled by only 1.8–4.9 °C Supercoolingof control leaves was reduced by dusting with aeciospores, particularlywhen leaves were wounded to simulate the rupture of the surfacecaused by sporulation, but wounding alone had no significanteffect. Supercooling of distilled water was also significantlyreduced by aeciospores, suspended at a concentration of 10⁵spores ml^–1. It is concluded that rust-induced changes in leaf freezing inS. vulgaris grown in controlled environments were due to anincrease in the number of sites for ice nucleation, caused bythe presence of the aeciospores, and increased penetration ofice into internal tissues, resulting from damage to the cuticleand epidermis. Although data for frost resistance obtained inthe growth-room are similar to previous field observations,the role of the above mechanisms under field conditions remainsunproven. Senecio vulgaris (groundsel), Puccinia lagenophorae (rust), low temperature, freezing resistance     

A Protein from Immature Raspberry Fruits which Inhibits Endopolygalacturonases from Botrytis cinerea and other Micro-organisms

A polygalacturonase-inhibiting protein (PGIP) was purified fromimmature raspberry fruits using ion exchange chromatography.The protein was composed of a single polypeptide chain withM_r of 38·5 kDa and a pI residing above pH 10. Kineticstudies suggested that the inhibition was of a non-competitivenature. The PGIP inhibited two endopolygalacturonases (endo-PG)purified from Botrytis cinerea and an endo-PG produced by Aspergillusniger to varying degrees but did not inhibit two exo-PGs purifiedfrom B. cinerea, bacterial endopectate lyases and bacterialendo-PGs. The concentration of PGIP at various stages of flowerand fruit development was determined. The inhibitor was notdetected in the flower, but reached a maximum of 69 units g^–1in the immature green fruit decreasing to 9 units g^–1as fruits matured. The N-terminal amino-acid sequence was determined. Key words: Polygalacturonase-inhibiting protein, Rubus idaeus, red raspberry, Botrytis cinerea, pectinases     

Supercooling ability of Rhododendron flower buds in relation to cooling rate and cold hardiness

The freezing process and supercooling ability in flower budsof 11 native Rhododendron species were examined with referenceto the cooling rate and cold hardiness by differential thermalanalysis. The freezing patterns of the excised whole buds variedwith the season: in autumn, buds froze as whole units, whilein winter, freezing was initiated in the scales and propagatedto each floret. The supercooling ability of florets was enhancedduring winter. The freezing patterns in winter buds were stronglyinfluenced by the cooling rate (1 to 30°C/hr). Althoughthe first exotherm in scales occurred at –5 to –10°Gand was rate-independent, the occurrence of several floret exothermsshifted considerably to lower subzero temperatures at slowerrates. The most reliable cooling rate for testing maximum supercoolingability was l°C/hr. The exotherm in florets of hardier speciesoccurred at –20 to –25°C and at –7 to–20°C for less hardy ones, and were well correlatedwith their killing temperatures. Water relations within budtissues in response to freezing are briefly discussed. (Received June 26, 1980; )     

Supercooling ability of Rhododendron flower buds in relation to cooling rate and cold hardiness

The freezing process and supercooling ability in flower budsof 11 native Rhododendron species were examined with referenceto the cooling rate and cold hardiness by differential thermalanalysis. The freezing patterns of the excised whole buds variedwith the season: in autumn, buds froze as whole units, whilein winter, freezing was initiated in the scales and propagatedto each floret. The supercooling ability of florets was enhancedduring winter. The freezing patterns in winter buds were stronglyinfluenced by the cooling rate (1 to 30°C/hr). Althoughthe first exotherm in scales occurred at –5 to –10°Gand was rate-independent, the occurrence of several floret exothermsshifted considerably to lower subzero temperatures at slowerrates. The most reliable cooling rate for testing maximum supercoolingability was l°C/hr. The exotherm in florets of hardier speciesoccurred at –20 to –25°C and at –7 to–20°C for less hardy ones, and were well correlatedwith their killing temperatures. Water relations within budtissues in response to freezing are briefly discussed. (Received June 26, 1980; )     

Myo-inositol Biosynthesis and Galactose Utilization by Wild Carrot (Daucus carota L. var. carota) Suspension Cultures

Wild carrot (Daucus carota var. carota) cell suspensions (63–120µm in diameter) were grown on a mineral salt medium containingdifferent carbon sources in the presence (10 mM) and absenceof myo-inositol. The data obtained after 14 and 21 days of growthshow that an external supply of myo-inositol is not essentialfor growth and development of wild carrot embryos. A linearrelationship was found between growth (d. wt) and embryo numberin the presence and absence of myo-inositol. Standard stock cell suspensions never exposed to exogenous myo-inositoland grown in the absence of 2, 4-D with glucose or galactoseas the carbon source synthesized radioactive myo-inositol whenexposed to D-[1–¹⁴C]glucose or D-[1–¹⁴C]galactose.Gas chromatographic analyses revealed the presence of myo-inositolin the bulk tissue grown in the presence of 2.25 µM 2,4-D with glucose, galactose, fructose or mannose as the solecarbohydrate. We could not detect any component indicating anisomer or a methylated derivative of an inositol in the tissueextracts. Stock cultures were maintained (with 2, 4-D) successfully forat least three successive sub-cultures on D-galactose as thesole carbohydrate. The growth achieved over this culture periodshowed that wild carrot cells used by us could quickly adaptto grow on D-galactose as rapidly as they grow on sucrose. Daucus carota L., wild carrot, suspension cultures, myo-inositol, galactose     

Polyderm, a Barrier to Infection of Red Raspberry Buds by Didymella applanata and Botrytis cinerea

A histological study was made of the axillary region of raspberrycanes infected naturally by Didymella applanata (Niessl) Sacc.and Botrytis cinerea Pers.: Fr. The outer suberized phellemlayer of the polyderm and a primary protective layer of suberizedand lignified cells across the adaxial cortex of the petioleprecluded infection of the axillary buds by hyphae growing froma saprophytic base in the leaf. No protective layer formed throughthe abaxial cortex at the petiole base; consequently both fungicolonized the epidermis, primary cortex and outermost non-suberizedphelloid cells of the polyderm beneath the node. Red raspberry, Rubus idaeus, Didymella applanta, Botrytis cinerea, polyderm, periderm, suberin, lignin