Increased accumulation of phenolic metabolites in groundnut (Arachis hypogaea L.) genotypes contribute to defense against Sclerotium rolfsii infection

Abstract

Fungal infections cause several metabolic changes to the plants, which can affect its physiology and survival in various ways. In the present study, we have analysed various phenolic compounds and activity of oxidative enzymes in healthy and Sclerotium rolfsii-infected groundnut genotypes. Increased phenolics content and higher activity of oxidative enzymes was observed in the tolerant genotype (CS 19, GG 16) followed by susceptible genotype (GG 20, TG 37A). Among the phenolic compounds tested, chlorogenic acid content has increased greatly in leaf, stem and root of infected tolerant genotypes compared to the respective controls. In vitro growth of S. rolfsii showed significant inhibition at concentrations 500 and 1000 µg/mL of phenolic compounds in the radial growth inhibition assay. These results have strongly suggested that, higher accumulation of chlorogenic acid could be an important factor in imparting resistance and protecting groundnut against S. rolfsii infection in tolerant genotypes.     

Calcium Enhanced Zoospore Production of Pythium myriotylum in vitro

Pythium myriotylum is the causal organism of Cocoyam Root Rot Disease (CRRD). Significant numbers of zoospores were induced within 1.5 h in cultures in Petri dishes containing P. myriotylum soaked in 0.01 M Ca⁺⁺ and sterile deionized distilled water. Soaking solutions # 2 and # 3 inhibited the production of zoospores of P. myriotylum. This may be due to the delay in maturation of sporangia and the release of zoospores when the soaking solutions contain sucrose. Significant necrosis of detached cocoyam plantlet roots in 100 ml beakers confirmed the infection of zoospores of two `local white' cocoyam genotypes. Detached `yellow' cocoyam roots in 100 ml beakers of genotype RO3015 resisted infection of P. myriotylum with no necrosis of the inoculated roots, which may indicate resistance. This provides a quick and reliable pathogenicity test of P. myriotylum on susceptible cocoyam detached roots. Necrosis of inoculated detached cocoyam roots could be reliably used to screen cocoyam germplasm for resistance to P. myriotylum.     

Time-course analysis of the phenols in cucumber mosaic virus-resistant, -tolerant and -susceptible tomato genotypes

In this study, changes in quantity and quality of phenolic compounds were compared in cucumber mosaic virus (CMV)-inoculated and -un-inoculated plants of nine resistant, tolerant, susceptible and highly susceptible genotypes at three different time intervals. Total phenolic contents and the number of phenolic compounds were generally increased in CMV-inoculated plants. Maximum per cent increase in total phenolic contents over un-inoculated controls was observed as 77.55% in resistant genotype TMS-1, 84.17% in tolerant genotype L06238 and 82.88% in resistant genotype L02223 after 10, 20 and 30 days of inoculation, respectively. Thin layer chromatography of inoculated and un-inoculated plants indicates that in most of the tested genotypes, the number of phenolic compounds varied from cultivar to cultivar and within the same cultivar, depending upon the status of plants and growth stages. However, the trend of increase in quantity and quality of phenolic compounds in the tested units was not constant to draw a meaningful conclusion.     

Changes in Cell Wall-bound Phenolic Compounds and Lignin in Roots of Date Palm Cultivars Differing in Susceptibility to Fusarium oxysporum f. sp. albedinis

The roots of date palm contain four cell wall‐bound phenolic acids identified as p‐hydroxybenzoic, p‐coumaric, ferulic and sinapic acids. The ferulic acid represents the major phenolic compound since it constitutes 48.2–55.8% of cell wall‐bound phenolic acids. All these phenolic acids were present in the resistant cultivar (BSTN) and the susceptible cultivar (JHL). However, the pre‐infection contents of p‐coumaric, ferulic and sinapic acids were greater in the resistant cultivar than in the susceptible one. For the contents of p‐hydroxybenzoic acid, there was no significant difference between the resistant cultivar and the susceptible cultivar. Similarly, the pre‐infection contents of lignin were approximately equal for both cultivars. Inoculation of the date palm roots by Fusarium oxysporum f. sp. albedinis induced important modifications to the contents of the cell wall‐bound phenolic compounds and lignin, which made it possible to distinguish between resistant and susceptible cultivars. The post‐infection contents of cell wall‐bound phenolic compounds underwent a rapid and intense increase with a maximum accumulation on the tenth day for p‐hydroxybenzoic acid (1.54 μmol/g), p‐coumaric acid (2.77 μmol/g) and ferulic acid (2.64 μmol/g) and on the fifteenth day for sinapic acid (1.85 μmol/g). The maximum contents accumulated in the resistant cultivar were greater than those in the susceptible cultivar, namely, 11 times for p‐hydroxybenzoic acid, 2.6 times for p‐coumaric acid, 1.8 times for ferulic acid and 12.3 times for sinapic acid. In the susceptible cultivar, p‐coumaric acid and ferulic acid contents also increased after inoculation although they did not reach the pre‐infection contents of the resistant cultivar. The contents of p‐hydroxybenzoic acid in the susceptible cultivar roots did not present post‐infection modification and those of sinapic acid decreased instead. The lignin contents increased in both cultivars with a maximum accumulation on the fifteenth day. However, the maximum contents accumulated in the resistant cultivar roots were 1.5 times greater than those of the susceptible cultivar. These results showed clear differences between the resistant BSTN and the susceptible JHL cultivars. The implication of cell wall‐bound phenolic compounds and lignin in the resistance of date palm to F. oxysporum f. sp. albedinis appears to be dependent on the speed and intensity of their accumulation with greater contents in the first stage of infection.     

Combining transgenic and natural resistance to obtain broad resistance to tospovirus infection in tomato (Lycopersicon esculentum mill)

This study was undertaken to develop tomato plants with broad resistanceto tospoviruses which are a major limiting factor to tomato productionworldwide. A nontransgenic tomato line Stevens-Rodale (S-R), six transgenictomato lines expressing the nucleocapsid (N) protein gene of the lettuceisolate of tomato spotted wilt virus (TSWV-BL), and progeny of the crosses between S-Rand three of the transgenic lines homozygous for the N gene were evaluated fortheir resistance to tospovirus infection in greenhouse inoculation tests. S-Rhas the Sw-5 gene that confers resistance to several TSWVisolates. The six transgenic lines showed high levels of resistance wheninoculated with either TSWV-BL or a tomato isolate from Hawaii (TSWV-H).However, these same plants were highly susceptible to the Brazilian isolate ofgroundnut ringspot virus (GRSV-BR). Plants with the Sw-5gene were resistant to TSWV-BL and GRSV-BR, but were susceptible to TSWV-H.When inoculated with any of the three viruses, the F₁ progeny of thecrosses exhibited a susceptible, tolerant, or resistant phenotype with a higherproportion of the plants being either tolerant or resistant. When F₂progeny from F₁ resistant plants of each cross were inoculated withany of the three viruses, a higher proportion of tolerant and resistant plantswas observed compared to the F₁ progeny. Our results show thepotential to obtain broad resistance to tospoviruses by combining transgenicand natural resistance in a single plant.     

Phenolic compounds in date palm cultivars sensitive and resistant toFusarium oxysporum

In date palm (Phoenix dactylifera L.) leaves, the main compounds of the phenolic pool were quercetin and isorhamnetin heterosides, (+)-catechin and (-)-epicatechin. Although previously observed only in date palm fruits, 5-caffeoylshikimic acid (dactylifric acid) and its positional isomers (3-caffeoylshikimic acid and 4-caffeoylshikimic acid) were detected also in the leaves and roots. Quantitative, but not qualitative, differences between cultivars resistant and susceptible toFusarium oxysporum f. sp.albedinis during growth period were observed Acknowledgements: The authors are very grateful to Dr. Janati, Director of SCAS (INRA, Marrakech) for supplying the plant material used in this study. The study was funded by the Programme de Cooperation Franco-Marocaine (Al 473/90) between Cadi Ayyad University (Marrakech, Maroc) and Montpellier II University (France)     

不同抗性短枝木麻黄种源苗木接种青枯病菌后酚类物质含量的变化

为了解短枝木麻黄(Casuarina equisetifolia)抗青枯病的机理,对接种青枯病菌(Ralstonia solanacearum)后短枝木麻黄的单宁和黄酮含量变化进行了分析。结果表明,不同抗性短枝木麻黄种源小枝的总酚和单宁含量呈现不同的变化趋势,高抗、中抗种源均呈现先升高后降低的变化趋势,峰值均约为126 mg g–1,但中抗种源的峰值出现时间较晚,而易感种源则呈逐渐升高趋势。抗、感种源木麻黄接种青枯菌后,小枝中缩合单宁含量均呈现逐渐升高的趋势,但高抗种源的缩合单宁含量均显著高于易感种源,增加70.33%。抗性种源黄酮含量呈S型上升趋势,易感种源则持续缓慢升高。这表明接种青枯病菌后,抗、感短枝木麻黄种源表现出不同的防御特征,次生物质含量增幅越大,抑菌抗氧化能力越强,短枝木麻黄表现出的抗性越强。     

The relationship between glycoalkaloids and resistance to the white potato cyst nematode, Globodera pallida in potato clones derived from Solanum vernei

No relationship was found between the degree of resistance to Globodera pallida and total glycoalkaloid content of the roots or tubers of a number of potato clones derived from Solanum vernei × S. tuberosum. Nematode infestation of the roots did not lead to increases in the glycoalkaloid content of susceptible or resistant potatoes.     

Effect of epiphytes on the extent of necrotic injuries of resistant and susceptible poplar clones infected with Dothichiza populea

Poplar cuttings of a resistant clone, Populus ‘Grandis’, and susceptible clones, Populus nigra ‘Italica’ and Populus ‘Robusta’, were infected with the pathogenic fungus Dothichiza populea alone, or with the pathogen and one of five strains of epiphytes antagonistic towards it (in vitro), isolated from poplar bark. The extent of injury was examined for 28 days after infection by determining the length of necrotic patches and their area as expressed in per cent of the total area of a cutting or the area of necrotic injuries caused by the pathogen alone.All the poplar cuttings of both the resistant and susceptible clones became diseased when infected with the pathogen alone. Surprisingly enough, however, the least affected clone was the susceptible P. ‘Robusta’, in which necrotic injuries covered 28% of the total area, as against 40% and 70% in the resistant P. ‘Grandis’ and the susceptible P. nigra ‘Italica’, respectively.When the cuttings were infected simultaneously with Dothichiza populea and its antagonistic epiphytes, the diseased area in the resistant clone diminished by as much as two-thirds, and in the susceptible P. nigra ‘Italica’, by one-third in comparison with the area affected by the pathogen alone. In turn, in the susceptible P. ‘Robusta’ the introduction of three out of five epiphytes stimulated the growth of the pathogenic fungus producing on average a double increase in the necrotic area. The differences in the response of the pathogen to the presence of epiphytes recorded in the susceptible clones indicate a marked influence of the plant on the nature of interactions between its epiphytic microflora and the pathogen.     

The use of foliage assessment to improve the identification of tolerance to damage by nematodes (Globodera pallida) in potatoes

Six potato trials, two in each of three years, were conducted in collaboration with the Agricultural Development and Advisory Service (ADAS) at sites infested with potato cyst nematodes (G. pallida Pa 2/3). The trials were part of a selective screen to identify PCN tolerant and intolerant clones with each trial consisting of four blocks divided into nematicide treated and untreated sub-blocks. A range of partially resistant and susceptible material was assessed for yield losses due to PCN damage to the roots and for the effect on the foliage by comparison between the nematicide treated and untreated areas. The relationships between the foliage symptoms, untreated yields, treated yields, proportional yield loss, initial PCN population and the post-harvest PCN population levels are examined. Significant correlation coefficients were obtained between foliage symptoms and yield of clones in PCN infected soil and also between foliage symptoms with percentage yield loss due to PCN infestation. The conclusions were that the assessment of PCN damage to foliage vigour/development can contribute positively to a more accurate identification of tolerant or intolerant potato genotypes.     

Isolation of anti-fungal phenolic compounds from petioles of two Hevea brasiliensis (rubber) genotypes and their effect on Phytophthora meadii

Phenolic compounds were present in greater amounts in non‐infected petioles of genotypes of Hevea brasiliensis that are resistant to Phytophthora leaf disease than in genotypes that are susceptible. Phenolic compounds extracted from petioles of either susceptible (PB86) or resistant (RRIC100) genotypes, before or after infection with Phytophthora meadii, had anti‐fungal properties. Artificially infected petioles of PB86 had phenolic acids, triterpenoids or flavonoids, whereas healthy petioles contained only triterpenoids or flavonoids. However, healthy or infected petioles of RRIC100 contained only trace amounts of the above compounds and of vanillin (3‐methoxy‐4‐hydroxybenzaldehyde). Vanillin and umbelliferone (7‐hydroxycoumarin) were shown to suppress zoospore germination of P. meadii on glass slides and to inhibit its growth in pea broth and V‐8 juice agar. Vanillin was slightly more active than umbelliferone. Resistance of RRIC100 to Phytophthora was suspected as being related to the polymerisation of phenolic compounds to form lignin, which may suppress further spread of the pathogen's mycelium into healthy tissues. Formation of lignin from phenolic aldehydes as a barrier to disease spread may be a critical factor in resistance.     

Epiphytic microflora of poplar clones susceptible and resistant to infection by Dothichiza populea

One-year-old poplar shoots (nodes, internodes and lenticels) of clones susceptible to infection by the pathogenic fungus Dothichiza populea, viz. Populus nigra Italica and P. Robusta, resistant ones, viz. P. Grandis and P. Hybrida 275, as well as a hybrid of a susceptible and a resistant clone, viz. P. maximowiczii x P. nigra (P. Kórnik 42), were used. The plate method was employed to determine: 1. the abundance of the epiphytic microflora on a mineral medium with glucose; 2. the quantitative composition of epiphytic communities by determining the numbers of typical bacteria (including rod-shaped, spherical and sporulating forms), actinomycetes and yeasts in microscopic preparations from epiphyte colonies; 3. the abundance and level of activity of epiphytes antagonistic towards Dothichiza populea. In all poplar clones the epiphytic microflora was most abundant on nodes and least abundant on lenticels. In the resistant clones epiphytes were 7 (P. Grandis) to as many as 84 times (P. Hybrida 275) less numerous than in the susceptible ones. In the microflora communities of the susceptible poplars, rod-shaped bacteria were the most abundant, and in the resistant ones and the hybrid, yeasts, which made up from 60% to 70% of the strains tested. Spherical and sporulating bacteria as well as actinomycetes were found in numbers not exceeding 4% of the total number of epiphytes. The proportion of antagonistic microflora in whole epiphytic communities was higher in the resistant clones and the hybrid than in the susceptible clones, with the microflora having a more restrictive effect on the development of the pathogen.     

Defense response of resistant and susceptible genotypes of castor (Ricinus communis L.) to wilt disease

The biochemical basis of resistance in castor (Ricinus communis L.) to Fusarium wilt, caused by the pathogen Fusarium oxysporum f. sp. ricini, was investigated. Induction of plant defence against pathogen attack is regulated by a complex network of different signals. Thus changes in various biochemical defenses including antioxidant enzymes, phenolic compounds and pathogenesis related (PR) proteins were investigated in the roots of resistant and susceptible genotypes of castor at 0, 24, 48 and 72 h.a.i. Infection by F. oxysporum significantly increased the superoxide dismutase (SOD) and peroxidase (POX) activities in the roots of susceptible genotypes, while the catalase (CAT) activities were appreciably higher in the roots of resistant genotypes at different stages. Constitutive levels of ascorbate peroxidase (APX) and polyphenol oxidase (PPO) were higher in the resistant genotypes. Also, the activities of phenylalanine ammonia lyase (PAL) and β 1, 3 glucanase significantly increased in the roots of the resistant genotypes after infections. The rate of increment of thiobarbituric acid reactive substances (TBARS) was higher in resistant genotypes after infection. Analysis of isozyme banding pattern of SOD, POX, PPO and esterase on native PAGE electrophoresis revealed that interaction between plant and fungi invoked various isozymes at 48 h of infection. SOD 3 was observed only in resistant genotypes at 24 h.a.i. except Geeta. Similarly induction of POX 5 was observed only in resistant genotypes at 48 h of infection, though the intensity of POX 5 was very less.     

Release of phenols from Lupinus albus L. roots exposed to Cu and their possible role in Cu detoxification

The mechanisms enabling plants to tolerate high concentrations of available Cu in their rhizosphere are still poorly understood. To better understand the mechanisms involved, Lupinus albus L. (white lupin) was grown over 40 days in a hydroponic system compelling roots to develop under sterile conditions in the presence of a nutrient solution containing 0.5, 20 or 62 M Cu. The following parameters were investigated in detail: low molecular weight phenols in nutrient solution (colorimetric assay), high molecular weight phenols in roots and in solution (HPLC-MS, HPLC-UV), pH, redox potential in solution (electrochemistry) and Cu distribution in the plant (AAS) as well as in apical root sections (EDX microanalysis). Finally, in vitro adsorption studies using voltammetry were conducted to evaluate the Cu adsorption behaviour of different phenolic compounds. When exposed to 62 M Cu, biomass production of white lupin was strongly reduced. Plants grown in the presence of 20 M Cu had a similar dry matter production compared to the control plants grown in a 0.5 M Cu solution. However, an increased release of soluble and high molecular weight phenols into the solution was observed. The concentration of polyphenolic compounds in the roots (particularly isoflavonoids like genistein and genistein-(malonyl)-glucoside) was significantly higher for lupins grown in a 20 M Cu solution compared to the control plants. As shown by an in vitro adsorption study, these phenolic compounds can bind Cu ions. In addition, plants exposed to 20 and 62 M Cu cumulated high Cu amounts in root cell walls whereas only low amounts reached the symplasm. Therefore, it is proposed that the complexation of Cu²⁺ ions in the rhizosphere and in the roots apoplasm by phenolic compounds could alleviate Cu-mediated toxicity.     

Kinetin regulation of growth and secondary metabolism in waterlogging and salinity treated Vigna sinensis and Zea mays

Waterlogging mostly increased fresh weight and water content in shoots and roots of Vigna sinensis and Zea mays while salinity seemed to have a decreasing effect. There was a marked induction of proline in shoots and roots of both plants by salinity with lower values in logged plants. In addition, anthocyanin content was increased in Vigna sinensis by both treatments and in Zea mays only by salinity. Meanwhile the treatments significantly accumulated phenolic compounds in plant shoots. Also there were increased activities of phenylalanine ammonia lyase (PAL) and tyrosine ammonia lyase (TAL) in shoots and roots of both plants. Foliar application of kinetin equilibrated, if any, the effects of both treatments on contents of proline, anthocyanin and phenolic compounds as well as activities of PAL and TAL in shoots and roots of treated plants. These findings reveal that kinetin alleviates the stress symptoms and regulates the changes in phenolic metabolism of waterlogged or salinity treated Vigna sinensis and Zea mays.     

Inducible Proteins in Citrus Rootstocks with Different Tolerance Towards the Root Rot Pathogen Phytophthora palmivora

Activities of defence‐related proteins (β‐1,3‐glucanases, chitinases and peroxidases) and concentrations of total soluble phenolics were measured in roots and leaves of non‐infected and infected plants to investigate the response of different citrus rootstock genotypes to the root rot pathogen Phytophthora palmivora Butler. Infection with the pathogen increased concentrations of total proteins, total phenolics and β‐1,3‐glucanase activity in roots of all genotypes, and increases were associated with the extent of root mass reductions and thus susceptibility of the plants. Root chitinase and root peroxidase levels were slightly reduced or unaltered upon infection. β‐1,3‐Glucanase activity was also elevated in leaves of infected plants, but increases did not differ between tolerant and susceptible rootstocks. Effects of root infection on leaves were typically the reverse of effects on roots for chitinase‐ and peroxidase levels and more pronounced in susceptible rootstock genotypes. Although differences in enzyme expression were observed between susceptible and tolerant citrus seedlings, effects were usually associated with disease progression, and not with resistance to P. palmivora. It is suggested that increased activities of the proteins and soluble phenolics studied are not implicated in the primary defence to Phytophthora root diseases, but may contribute to the inhibition of the pathogen during infection in tolerant citrus.     

Selection of turmeric callus tolerant to culture filtrate of <Emphasis Type="Italic">Pythium Graminicolum</Emphasis> and regeneration of plants

Root rot disease tolerant clones of turmeric variety Suguna of Curcuma longa L. were isolated using continuous in vitro selection technique against pure culture filtrate of Pythium graminicolum. Large amount of profuse, compact, creamish white callus was obtained from in vivo vegetative bud when cultured on LSBM fortified with 2,4-D (3 mg l⁻¹) after 45 days of culture. Callus was challenged with pure culture filtrate of P. graminicolum to isolate viable callus within 30 days of culture, which was further subjected to pure culture filtrate treatment. After three cycles of treatment, four cell lines which are tolerant to culture filtrate was isolated through continuous in vitro selection and subcultured on regeneration medium LSBM fortified with BAP (4 mg l⁻¹) along with the control non-selected callus to obtain complete plantlets through discontinuous in vitro selection technique. Plants regenerated from tolerant and non-selected calli were screened for disease tolerance by adopting in vitro sick plot technique. The data obtained from this experiment revealed a ratio of 225:49 tolerant: susceptible in vitro clones retrieved from tolerant callus. However, plants regenerated from the CL1a₁ and non-selected calli were susceptible under in vitro sick plot technique. The root rot disease tolerant clones were hardened and established in soil with 90% survival frequency.     

An RFLP marker in tomato linked to the Fusarium oxysporum resistance gene I2

Summary The locus, I2, which in tomato confers resistance against Fusarium oxysporum f. sp. lycopersici race 2, was introgressed into Lycopersicon esculentum from the wild species L. pimpinellifolium (P.I. 126915). We searched for restriction fragment length polymorphisms (RFLPs) between nearly isogenic lines (NILs) in clones that map to the region introgressed from the wild species. Since I2 maps to chromosome 11, we used DNA clones from this chromosome as hybridization probes to Southern blots containing bound DNA of the NILs digested with 23 restriction enzymes. Of the 14 chromosome 11 clones, 9 exhibited polymorphism. These clones were further hybridized to verification filters that contained DNA from resistant and susceptible L. esculentum varieties digested with the enzymes that gave the polymorphism. One clone, TG105, was found to be associated with I2; 19 susceptible lines showed a different RFLP with this probe than 16 resistant lines, including the original L. pimpinellifolium accession used as a source for the resistance gene. These results together with our mapping analysis indicate that TG105 is closely linked to the resistance gene.     

cDNA cloning of mRNAs induced in resistant barley during infection by Erysiphe graminis f.sp. Hordei

Summary Near-isogenic cultivars of Hordeum vulgare which differ for the Mlp gene for resistance to Erysiphe graminis f.sp. hordei were inoculated with race 3 of this pathogen and in vitro translation products of mRNA populations compared by 2-dimensional gel electrophoresis and fluorography. This revealed the presence of new mRNA species in infected leaves compared to non-inoculated controls. These new mRNA species were more abundant in resistant leaves than susceptible leaves. A cDNA library was prepared from poly(A)⁺RNA isolated from infected leaves carrying the Mlp gene for resistance (cvMlp). The library was screened by differential hybridization using [³²P]-labelled cDNA prepared from poly(A)⁺RNA of both control and infected leaves. Six cDNA clones showing greater hybridization to cDNA prepared from infected leaves were selected. These six cDNA clones hybridized to DNA isolated from barley leaves but not to DNA from conidia of the fungus. In Northern blot analysis of RNA from infected leaves the six cDNA clones each hybridized to mRNA species of different size. Translation products for three of the cDNA clones corresponded to infection-related translation products identified on 2-dimensional fluorograms. The cDNA clones were used to study the kinetics of host mRNA induction during infection of the near-isogenic cultivars of barley. The host mRNA species corresponding to the cDNA clones were induced prior to 24 h after inoculation during the primary penetration processes. In addition the mRNAs corresponding to four of the cDNA clones increased to greater amounts in cvMlp than in the near-isogenic susceptible cultivar (cvmlp) over a 2-d period following inoculation. These results suggest that the Mlp gene has a regulatory role in host gene expression resulting in enhanced expression of several host mRNA species following infection by the powdery mildew fungus.