Effects of three esca-associated fungi on Vitis vinifera L.: II. Characterization of biomolecules in xylem sap and leaves of healthy and diseased vines

Secondary metabolites and host defense compounds were shown to occur in xylem sap, and leaves of Vitis vinifera cv. Italia and cv. Matilde naturally infected by the esca-associated fungi Phaeomoniella chlamydospora (Pch), Togninia minima (Tmi) and Fomitiporia mediterranea (Fme). Samples of xylem sap and leaves were collected from healthy vines and from vines showing severe symptoms of brown wood-streaking caused by Pch and Tmi, or from vines with symptoms of both brown wood-streaking and white rot caused by Fme. Xylem sap collection was carried out during the early spring of 2003 and 2004, corresponding to the phenological phases: (A) cotton bud; (B) green tip; (C) leaves out; (D) stretched out leaves; and (E) visible clusters. In the present work we have studied the accumulation of biomolecules (pentaketides and α-glucans), host defense compounds (benzaldehydes, benzoic acid and cinnamic acid derivatives, flavonols, flavanols, flavan-3-ol derivatives and stilbenes) at different stages of grapevine development. Accumulation and changes in total phenolics and recurring phenolics, and of three phytotoxic secondary metabolites (scytalone, isosclerone and pullulan) were analyzed by HPLC. On comparing results for cv. Italia and cv. Matilde, it can be seen that phenolic concentrations are strongly related to the cv.     

Effects of three esca-associated fungi on Vitis vinifera L.: III. Enzymes produced by the pathogens and their role in fungus-to-plant or in fungus-to-fungus interactions

When the esca-associated fungi Phaeomoniella chlamydospora (Pch), Togninia minima (Tmi) and Fomitiporia mediterranea (Fme) were grown in liquid stationary cultures, it was seen that they were able to live in media containing resveratrol (RES) or tannic acid (TA) as the sole carbon source and that the fungi were able to convert both compounds. Particular attention is paid here to detecting RES and TA conversion. Pch, Tmi and Fme were partially inhibited by RES or TA. Pch, Tmi and Fme produced extracellular tannase, laccase and peroxidase enzymes in liquid or agarized cultures, whether glucose was present or not. When colonies of Pch, Tmi and Fme were confronted, they showed spatially and temporally heterogeneous patterns of laccase and peroxidase activity. The results indicate the non-synergistic, competitive association of Pch and Tmi and the inhibition of Fme growth. Muconic acid, a well-known intermediate in a large number of lignin and phenol oxidative processes, can partly or completely inhibit the lignolytic agent Fme, but is tolerated by Pch and Tmi. An explanation for wood pigmentation patterns by Pch, Tmi and Fme is given.     

Pisatin demethylation by fungal pathogens and nonpathogens of pea: association with pisatin tolerance and virulence

Previous studies have indicated that detoxification of their hosts' phytoalexins is a tolerance mechanism for some true fungi, but not the fungus-like Oomycota, and may be involved in determining the virulence of a pathogen. In the present study, the associations between demethylation of the pea phytoalexin pisatin, tolerance to pisatin, and virulence on pea were examined for 50 fungal isolates which represent 17 species of pathogens and nonpathogens of pea. All isolates ofPythium coloratum and P. irregulare failed to metabolize and were sensitive to pisatin, consistent with previous observations that members of the Oomycota generally lack the ability to metabolize and are sensitive to their hosts' phytoalexins. Among true fungi tested, the ability to demethylate pisatin was common, regardless of whether the particular isolate was pathogenic on pea or not. However, when the rate of pisatin demethylation was compared to virulence, all but one of the moderate to highly virulent isolates rapidly demethylated pisatin. In addition, the more rapidly demethylating isolates were generally more tolerant of pisatin. These results suggest that a specialized enzyme system for quickly detoxifying pisatin might be present in most pea pathogens. In previous studies a specific cytochrome P450 enzyme for demethylating pisatin was identified in the pea pathogen Nectria haematococca mating population VI, and genes (PDA genes) encoding that enzyme have been cloned from this fungus. When DNA specific for these genes was used to probe genomic DNA from other fungi that demethylate pisatin, significant hybridization was detected with only one fungus, the pea pathogen Fusarium oxysporum f. sp. pisi. If the other pea pathogens possess a specific cytochrome P450 system for detoxification of pisatin, the genes encoding these enzymes apparently share limited nucleotide similarity with N. haematococca PDA genes.     

Effects of three esca-associated fungi on Vitis vinifera L.: I. Characterization of secondary metabolites in culture media and host responses to the pathogens in calli

Phaeomoniella chlamydospora (Pch) and Togninia minima (Tmi) produced scytalone, isosclerone and pullulans in liquid cultures, as well as in calli. Secondary metabolites and host defense compounds were shown to occur in calli of Vitis vinifera cv. Italia and cv. Matilde infected by the esca-associated fungi Pch, Tmi and Fomitiporia mediterranea (Fme). Calli of both cvs. were grown as dual cultures with Pch, Tmi and Fme. The fungi grew well in the presence of calli of both cvs., but callus growth was reduced. Accumulation and changes of total phenolics and recurring phenolics, and of two phytotoxic pentaketides and pullulans were analyzed by HPLC. On comparing results for cv. Italia and cv. Matilde, it can be seen that concentrations of phenolics are strongly related to the cv. The paper discusses the possible relationship between melanin biosynthesis in Pch and Tmi, which utilize pentaketide metabolites as intermediates and their pathogenicity related to phytotoxity of scytalone and isosclerone.     

瓜枝孢弱致病菌诱导黄瓜植保素的积累及抑菌活性

瓜枝孢(Cladosporium cucumernium ) 弱致病菌株2-2A₂诱导黄瓜产生对黄瓜黑星病的抗病性反应。对经2-2A₂诱导后黄瓜叶片中提取的物质进行生物活性试验、硅胶板薄层层析、高效液相色谱分析,证明2-2A₂可以有效诱导黄瓜中类黄酮类植保素的积累。     

Competitiveness of Rhizobium leguminosarum bv. phaseoli strains in relation to environmental stress and plant defense mechanisms

Summary Six Rhizobium leguminosarum bv. phaseoli strains (Ciat 151, Ciat 895, Ciat 899, CE3, H2C, Kim5s) were tested for nodule occupancy in different bean cultivars at two field sites (one fertile, one acid tropical soil) and in the greenhouse. The effects of several environmental factors such as low pH, high temperature, Al and Mn toxicity, iron deficiency, bean tannins, and bean phytoalexins were tested in vitro. Strain Kim5s was competitive under all tested conditions while strains CE3 and H2C had consistently low nodule occupancy levels. Strain Ciat 151 was superior to the other inoculant strains in the acid soil but competed poorly in the fertile soil. Strain Ciat 895 was more competitive in the fertile soil. There was a decline in nodule occupancy for all strains tested from the first trifoliate leaf stage to the pod-filling stage. No plant genotype effect on nodule occupancy was observed. There were significant (P<0.05) plant genotype and location effects, but no significant strain effect on acetylene reduction activity, plant dry weight, and nodule number. The greenhouse experiments confirmed, at least partially, the results from the field trials. In Leonard jars with an acid soil, strains Ciat 151 and Kim5s were amongst the most competitive strains. In coinoculation experiments, Kim5s was the most competitive strain, followed by Ciat 899 and Ciat 895. The competitiveness of a given strain was affected by the coinoculant strain. Tolerance in vitro to low pH, high growth temperature, Al or Mn toxicity, or Fe limitation was not related to competitiveness of the inoculum strains. The sensitivity of the strains towards bean tannins or bean phytoalexins also was not correlated with their competitiveness.     

Signals that stop the rot: Regulation of secondary metabolite defences in cereals

Plants accumulate a vast arsenal of chemically diverse secondary metabolites for defence against pathogens. This review will focus on the signal transduction and regulation of defence secondary metabolite production in five food security cereal crops: maize, rice, wheat, sorghum and oats. Recent research advances in this field have revealed novel processes and chemistry in these monocots that make this a rich field for future research.     

Role of stilbenes in the resistance of grapevine to powdery mildew

Stilbene phytoalexins are identified as defence response in pathogen–grapevine interactions, but little information is available on the role of stilbenes on Erysiphe necator, causal agent of grapevine powdery mildew. Analysis of stilbenes in artificially infected leaf discs from susceptible to highly resistant cultivars was performed and compared to the development of the pathogen. Results indicate that stilbene synthesis is confined in infected cells, penetrated by an appressorium–peg. Stilbene amounts expressed by infection site allow discriminating susceptible and resistant cultivars. Highest viniferins concentrations on resistant cultivars are in correspondence with the observed inhibition of the pathogen growth. The analysis of stilbenes at the infection site and viniferins accumulation in grapevine defence reaction is discussed.     

Glycosylation and oxidative dimerization of resveratrol are respectively associated to sensitivity and resistance of grapevine cultivars to downy mildew

Levels of resistance to Plasmopara viticola, from susceptible to highly resistant, of different grapevine cultivars were observed in vineyards and confirmed by the symptoms developed after inoculations. On the abaxial surface of infected leaves, P. viticola developed abundant sporangiophores on susceptible cultivars (Chasselas, Gamay, Gamaret and Pinot Noir), whereas on less susceptible cultivars (Seyval Blanc and Johanniter) the parasite produced few sporangiophores and some necrotic spots at the site of infection. On resistant cultivars (Bronner, Solaris, IRAC 2091), P. viticola induced a hypersensitive response and only necrotic spots were visible and the disease ceased to develop. Stilbenes were analyzed 4, 7, 24, 48 h post-infection (hpi) on small leaf samples cut from the site of infection. Large differences were observed between the cultivars at 24 and 48 hpi. Susceptible grapevines produced resveratrol and its glycoside, piceide. In contrast, resistant plants produced high concentration of ε- and δ-viniferin. Resveratrol and piceide have little or no toxicity activity against P. viticola, whereas δ-viniferin is highly toxic and can be considered an important marker for resistance of grapevine to downy mildew. The importance of oxidative dimerization of resveratrol in comparison to the extent of its glycosylation in defense reaction of grapevines against P. viticola is discussed.     

Effects of resveratrol on the ultrastructure of Botrytis cinerea conidia and biological significance in plant/pathogen interactions

Many roles have been ascribed to stilbenes, namely as antimicrobial, deterrent or repellent compounds in plants, protecting them from attacks by fungi, bacteria, nematodes or herbivores, acting both as constitutive and active defense (phytoalexin) compounds. More recently, stilbenes (especially resveratrol and its derivatives) were acclaimed for their wondrous effects and wide range of purported healing and preventive powers as cardioprotective, antitumor, neuroprotective and antioxidant agents. Although there is a huge number of works concerning the role of resveratrol in human health, reports on the antifungal activity of this compound are still scarce. This study was thus conducted in order to investigate the toxicity of resveratrol at an ultra- structural level to dormant conidia of Botrytis cinerea, the causal microorganism for gray mold. In grapevine particularly, this disease can affect all the green organs but is particularly damaging for ripening berries. Observations using transmission electron microscopy showed the occurrence of damages on conidia treated with sub-lethal doses, that is, 60 μg/mL (2.6 × 10^− 4 M) of resveratrol, a concentration usually reached in grapevine leaves and grape berries challenged by this pathogen. These results provide further data about the overall mode of action of this phytoalexin and its role in the B. cinerea/grapevine interaction.