Endophytic fungi are a taxonomically and ecologically heterogenous group of organisms, mainly belonging to the Ascomycotina and Deuteromycotina. The isolation methods affect the species composition of the endophyte assemblage in a given host. The number of endophyte taxa isolated from a host species is usually large; however, only few, normally host specific species or strains are dominant. Endophyte assemblages are specific at the host species level, but species composition and frequencies are significantly affected by site-specific conditions. Moreover, the relative importance and number of endophytic species vary among individuals within sites. In some cases, each individual could be considered a separate ecosystem. In general, however, 40 individuals with 30 to 40 sampling units per organ and individual should be enough to detect 80% of taxa present in a given host at one site. Endophytes usually produce the enzymes necessary for the colonization of plant tissues. Substrate utilization studies and isozyme analysis have demonstrated that most endophytes are able to utilize most plant cell components. The production of growth promoting factors and of metabolites useful in the pharmaceutical and agricultural industry is widespread among endophytic fungi. The usefulness of endophytes in agricultural and pharmaceutical research is briefly discussed.
Delta-viniferin is a resveratrol dehydrodimer, an isomer of epsilon-viniferin. This compound has been reported as a molecule produced in vitro by the oxidative dimerization of resveratrol by plant peroxidases or fungal laccases. It was also recently identified in wines and in grape cell cultures. We have now identified this dimer by NMR, high-performance liquid chromatography-diode array detection (HPLC-DAD), and HPLC-mass spectrometry (MS) in grapevine leaves infected by Plasmopara viticola (downy mildew) or UV-C irradiated. Its concentration was higher than that of epsilon-viniferin and constitutes one of the most important phytoalexins derived from resveratrol.
Esca disease, which attacks the wood of grapevine, has become increasingly devastating during the past three decades and represents today a major concern in all wine-producing countries. This disease is attributed to a group of systematically diverse fungi that are considered to be latent pathogens, however, this has not been conclusively established. This study presents the first in-depth comparison between the mycota of healthy and diseased plants taken from the same vineyard to determine which fungi become invasive when foliar symptoms of esca appear. An unprecedented high fungal diversity, 158 species, is here reported exclusively from grapevine wood in a single Swiss vineyard plot. An identical mycota inhabits wood of healthy and diseased adult plants and presumed esca pathogens were widespread and occurred in similar frequencies in both plant types. Pioneer esca-associated fungi are not transmitted from adult to nursery plants through the grafting process. Consequently the presumed esca-associated fungal pathogens are most likely saprobes decaying already senescent or dead wood resulting from intensive pruning, frost or other mecanical injuries as grafting. The cause of esca disease therefore remains elusive and requires well executive scientific study. These results question the assumed pathogenicity of fungi in other diseases of plants or animals where identical mycota are retrieved from both diseased and healthy individuals.
The impact of water deficit on stomatal conductance (gs), petiole hydraulic conductance (Kpetiole), and vulnerability to cavitation (PLC, percentage loss of hydraulic conductivity) in leaf petioles has been observed on field-grown vines (Vitis vinifera L. cv. Chasselas). Petioles were highly vulnerable to cavitation, with a 50% loss of hydraulic conductivity at a stem xylem water potential (Ψx) of –0.95 MPa, and up to 90% loss of conductivity at a Ψx of –1.5 MPa. Kpetiole described a daily cycle, decreasing during the day as water stress and evapotranspiration increased, then rising again in the early evening up to the previous morning's Kpetiole levels. In water-stressed vines, PLC increased sharply during the daytime and reached maximum values (70–90%) in the middle of the afternoon. Embolism repair occurred in petioles from the end of the day through the night. Indeed, PLC decreased in darkness in water-stressed vines. PLC variation in irrigated plants showed the same tendency, but with a smaller amplitude. The Chasselas cultivar appears to develop hydraulic segmentation, in which petiole cavitation plays an important role as a ‘hydraulic fuse’, thereby limiting leaf transpiration and the propagation of embolism and preserving the integrity of other organs (shoots and roots) during water stress. In the present study, progressive stomatal closure responded to a decrease in Kpetiole and an increase in cavitation events. Almost total closure of stomata (90%) was measured when PLC in petioles reached >90%.
Inflorescences of field-grown grapevines (Vitis vinifera L. cv. Gamay) were inoculated with a Botrytis cinerea conidia suspension or dried conidia at different stages during bloom in moist weather. Approximately 10% of the conidia germinated within 72 h, resulting in two to three times more latent infections than uninoculated controls in pea-size (7 mm in diameter) berries. In surface-sterilized pea-size berries, latent B. cinerea was present predominantly in the receptacle area. After veraison, latent B. cinerea also was found in the style and, in mature berries, latent colonies were distributed throughout the pulp. Inoculation at full bloom led to the highest disease severity (66%) at harvest, compared with 38% in controls. Stilbene stress metabolites in the flowers were measured by high-performance liquid chromatography. Resveratrol accumulated mainly after pre-bloom and full-bloom inoculation, but did not prevent infection. Piceid levels did not change following inoculation, while epsilon-viniferin was found in necrotic tissues only, and pterostilbene and alphaviniferin were not detected at all. B. cinerea conidia suspensions also were applied to various locations on flowers of pot-grown cvs. Pinot Noir and Chardonnay. Inoculation of the receptacle area, but not that of the stigma and ovary, resulted in latent infections. Stilbene synthesis was similar to the field results, with resveratrol accumulating mainly in the calyptra and receptacle area. Constitutive soluble phenolic compounds (mainly derivatives of quercetin and hydroxy-cinnamic acid) were present at high concentrations in the calyptra but at low levels in the receptacle area. These experiments confirmed bloom as a critical time for B. cinerea infection in grapes and suggest that the most likely site of infection is the receptacle area or cap scar exposed at anthesis. Stilbenes may have a limited role in inhibition of flower infection and latency in susceptible grape cultivars, and epsilon-viniferin may be a by-product rather than a deterrent of infection.
Leaves of different Vitis vinifera L. cultivars, susceptible or resistant to downy mildew, Chasselas, Solaris, IRAC 2091 (cvs. Gamaret x Bronner) and Muscadinia rotundifolia were inoculated with Plasmopara viticola. Samples were then examined by scanning and transmission electron microscopy, by light microscopy and for their ability to synthesise stilbenes. These phytoalexins were strictly analysed at infection sites. In the susceptible Chasselas, P. viticola colonises, at 72h post-infection (hpi), all of the spongy mesophyll with functional haustoria and produces mainly the non toxic piceide. No necrotic zone was observed on Chasselas leaves. The ultrastructural response to downy mildew infection is different in each of the other three resistant grape cultivars. In Solaris, where leaf necrosis are rapidly induced, the infection is restricted to the upper part of the loose spongy mesophyll, and associated with a rapid cell wall disruption and the dispersion of cytoplasmic content along with the production of viniferins. In IRAC 2091, leaf necrosis are quite similar to those observed on Solaris but the infected plant cell, as well as the haustoria, show high electron dense cellular particles without any recognisable organelles, probably related to the effect of the toxic compound pterostilbene, which is synthesised in this grape cultivar. In M. rotundifolia leaf necrosis are much more scarce and smaller than in other cultivars, but pathogen and plant cells are both strongly affected, with concomitant expulsion of cytoplasmic materials through the stomata after P. viticola penetration. In this cultivar, the concentration of all identified stilbenes exceeds 1×10³ μmol mg(-1) FW. The critical role of stilbenes in the resistance of Vitis spp. is discussed.
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