Cell-cell recognition of host surfaces by pathogens. The adsorption of maize (Zea mays) root mucilage by surfaces of pathogenic fungi.

The adsorption of radioactive mucilage by pathogenic fungi was shown to be dependent upon time, the composition of mucilage, the type of fungal surface (conidia, hyphae, hyphal apices), fungal species, pH and bivalent cations. All fungal adhesins were inactivated by either proteinase or polysaccharase treatments. Adsorption was not inhibited by the numberous mono-, di- and oligo-saccharides that were tested individually, but it was inhibited absolutely by several polysaccharides. This suggested that adsorption of mucilage by pathogens involved conformational and ionic interactions between plant and fungal polymers but not fungal lectins bound to sugar residues of mucilage. Several fractionation schemes showed that pathogens bound only the most acidic of the variety of polymers that comprise mucilage. There was not any absolute distinction between ability to bind radioactive mucilage and type of pathogen or non-pathogen. However, there were notable differences in characteristics of adsorption between two types of pathogen. Differences were revealed by comparison of the adsorption capacities of conidia and germinant conidia and chromatography of radioactive mucilage on germinant conidia. An ectotrophic root-infecting fungus (a highly specialized pathogen) bound a greater proportion of mucilage than did a vascular-wilt fungus (of catholic host and tissue range) with more than one class of site for adsorption. In contrast with the vascular-wilt fungus, sites for adsorption on the specialized pathogen were present solely on surfaces formed by germination.     

Mucilages and polysaccharides in Ziziphus species (Rhamnaceae): localization, composition and physiological roles during drought-stress.

The drought-tolerant tree species Ziziphus mauritiana Lamk. and Z. rotundifolia Lamk. were shown to have similar high mucilage concentrations (7-10% dry weight) in their leaves, with large numbers of mucilage-containing cells in the upper epidermis and extracellular mucilage-containing cavities in the leaf veins and stem cortex. The main sugar constituents of the water-soluble mucilage extract were rhamnose, glucose and galactose. During drought-stress in two independent studies, foliar mucilage content was unaffected in both species, but glucose and starch contents declined significantly in crude mucilage extracts from droughted leaves. Enzymatic hydrolysis of the mucilage extract using alpha-amylase and amyloglucosidase released glucose, indicating that a mucilage-associated water-soluble glucan, with alpha-1,4- and alpha-1,6-linkages, may exist which was extracted together with the mucilage. From the current data, it is not possible to localize the glucan to determine whether or not it is associated with mucilage-containing cells. Data from pressure-volume analyses of drought-stressed and control leaves showed that, in line with their similar mucilage contents, the relative leaf capacitance isotherm (change in relative water content per unit change in water potential) was similar in both species. During drought-stress, reduced relative capacitance resulted from osmotic adjustment and decreased wall elasticity. Data suggest that in Ziziphus leaves, intracellular mucilages play no part in buffering leaf water status during progressive drought. In Ziziphus species, growing in environments with erratic rainfall, the primary role of foliar mucilage and glucans, rather than as hydraulic capacitors, may be as sources for the remobilization of solutes for osmotic adjustment, thus enabling more effective water uptake and assimilate redistribution into roots and stems prior to defoliation as the drought-stress intensified.     

Wheat cells accumulate a syringyl-rich lignin during the hypersensitive resistance response

The stem rust fungus Puccinia graminis f.sp. tritici is an obligately biotrophic pathogen attacking wheat (Triticum aestivum). In compatible host/pathogen-interactions, the fungus participates in the host's metabolism by establishing functional haustoria in the susceptible plant cells. In highly resistant wheat cultivars, fungal attack is stopped by a hypersensitive response of penetrated host cells. This mechanism of programmed cell death of single plant cells is accompanied by the intracellular accumulation of material with UV-fluorescence typical of phenolic compounds. A similar reaction can be induced in healthy wheat leaves by the application of a rust-derived elicitor. We analysed the biochemical composition of this defense-induced phenolic material. Contents of total soluble and cell wall esterified and etherified phenolic acids were determined in rust-inoculated and elicitor-treated leaves of the fully susceptible wheat cultivar Prelude and its highly resistant, near-isogenic line Prelude-Sr5. While no resistance-related changes occured in any of these fractions, the lignin content as determined by the thioglycolic acid and the acetyl bromide methods increased after elicitor treatment. Nitrobenzene oxidation revealed that the entire increase can be explained by an increase in syringyl units only. These biochemical data were confirmed by fluorescence emission spectra analyses which indicated a defense-induced enrichment of syringyl lignin for cell wall samples both from elicitor-treated wheat leaves and single host cells undergoing a hypersensitive response upon fungal penetration.     

A highly differentiated glomeromycotean association with the mucilage-secreting, primitive antipodean liverwort Treubia (Treubiaceae): clues to the origins of mycorrhizas

Thallus anatomy in three species of the primitive liverwort genus Treubia (Metzgeriidae, Treubiales) was studied by light and electron microscopy. The thallus exudes copious mucilage, a feature shared elsewhere in liverworts only with the mycotrophic subterranean axes of the allied genus Haplomitrium. The central strand in the thallus midrib has a unique histological organization and harbors an intra- and intercellular infection by a glomeromycotean fungus that is far more highly differentiated than most of the glomeromycotean associations described to date. The fungus enters the thallus via clefts in the ventral epidermis along the midrib and colonizes the parenchyma above, forming intracellular coils and prominent, relatively short-lived, hyphal swellings. Above the zone with intracellular colonization is a tissue area containing mucilage-filled intercellular spaces; here the fungus is entirely intercellular and forms abundant pseudoparenchymatous structures and, in more mature parts of the thalli, large hyphae with thick multistratose walls. Mucilage in Treubia differs in histochemistry and origin from that produced by apical papillae, via hypertrophied Golgi, in all other bryophytes. Remarkable parallels between fungal associations in Treubia, Haplomitrium, and Lycopodium, all members of very ancient lineages, suggest that these associations epitomize very early stages in the evolution of glomeromycotean symbioses.     

Microbial impacts on plant-herbivore interactions: the indirect effects of a birch pathogen on a birch aphid

The role of indirect interactions in structuring communities is becoming increasingly recognised. Plant fungi can bring about changes in plant chemistry which may affect insect herbivores that share the same plant, and hence the two may interact indirectly. This study investigated the indirect effects of a fungal pathogen ( Marssonina betulae) of silver birch ( Betula pendula) on an aphid ( Euceraphis betulae), and the processes underpinning the interaction. There was a strong positive association between natural populations of the aphid and leaves bearing high fungal infection. In choice tests, significantly more aphids settled on leaves inoculated with the fungus than on asymptomatic leaves. Individual aphids reared on inoculated leaves were heavier, possessed longer hind tibiae and displayed enhanced embryo development compared with aphids reared on asymptomatic leaves; population growth rate was also positively correlated with fungal infection when groups of aphids were reared on inoculated branches. Changes in leaf chemistry were associated with fungal infection with inoculated leaves containing higher concentrations of free-amino acids. This may reflect a plant-initiated response to fungal attack in which free amino acids from the degradation of mesophyll cells are translocated out of infected leaves via the phloem. These changes in plant chemistry are similar to those occurring during leaf senescence, and are proposed as the mechanistic basis for the positive interaction between the fungus and aphid.     

Reversible cytoplasmic rearrangements precede wall apposition,hypersensitive cell death and defense-related gene activation in potato/Phytophthora infestans interactions

We used video microscopy techniques as a tool for live examination of the dynamic aspects of plant/fungus interactions. Early, dynamic responses of epidermal midrib cells of leaves from a potato cultivar (Solanum tuberosum L. cv. Datura) carrying resistance gene R1 to Phytophthora infestans (race 1: compatible interaction, race 4: incompatible interaction) were monitored. Similar responses were observed in both types of interaction, ranging from no visible reaction of invaded plant cells to hypersensitive cell death. The overall defense response of each individual cell exhibited a highly dynamic behavior that appeared to be tightly coordinated with the growth of the fungus. Initial localized reactions, including major rearrangements within the cytoplasm, occurred directly at the fungal penetration site, where rapid apposition of autofluorescent material and callose took place. If fungal invasion stopped at this stage, the host cell restored its normal cytoplasmic activity and survived. Hypersensitive cell death occurred only when fungal growth had proceeded to the formation of a clearly identifiable haustorium. In such cases, cytoplasm and nucleus conglomerated around the intracellular fungal structure, followed by a sudden collapse of the whole conglomerate and an instantaneous collapse of the fungal haustorium. Only small quantitative differences between the compatible and incompatible interactions of the two fungal races were observed for these early responses of epidermal cells. In the incompatible interaction, a slightly larger number of epidermal cells responded to fungal attack. More pronounced quantitative differences between compatible and incompatible interactions occurred upon fungal invasion of the mesophyll. These differences in the number of responding cells were not reflected at the level of gene expression: the spatial and temporal activation patterns of two defense-related genes, encoding phenylalanine ammonia-lyase and pathogenesis-related protein 1, were similar in both types of interaction.Dedicated to Professor Peter Sitte, Freiburg, Germany, on the occasion of his 65th birthday     

Use of green fluorescent protein to quantify the growth of Colletotrichum during infection of tobacco

To develop a quantitative assay of fungal growth inside plant tissues, strains of Colletotrichum destructivum and Colletotrichum orbiculare were transformed with a modified green fluorescent protein (GFP) gene fused with a glyceraldehyde-3-phosphate dehydrogenase promoter from Aspergillus nidulans. Transformants expressed GFP in culture and had the same growth rate and general appearance as the wild type. GFP was observed in all fungal structures during infection of leaves of Nicotiana benthamiana, except for the melanized appressoria and setae. The timing and appearance of the fungal structures in the host appeared to be identical to that of the wild type. GFP accumulation in inoculated leaves of N. benthamiana was quantified in leaf extracts using a fluorescence microplate reader, and the quantity of fluorescence was strongly correlated with the growth of the fungus as measured by the amount of fungal actin gene expression using Northern blot hybridizations. These results demonstrated that assaying green fluorescence levels from a GFP-transformed fungus is an accurate, fast and easy means of quantifying fungal growth inside host plant cells.     

Antifungal activity of the ribosome‐inactivating protein BE27 from sugar beet (Beta vulgaris L.) against the green mould Penicillium digitatum

The ribosome‐inactivating protein BE27 from sugar beet (Beta vulgaris L.) leaves is an apoplastic protein induced by signalling compounds, such as hydrogen peroxide and salicylic acid, which has been reported to be involved in defence against viruses. Here, we report that, at a concentration much lower than that present in the apoplast, BE27 displays antifungal activity against the green mould Penicillium digitatum, a necrotrophic fungus that colonizes wounds and grows in the inter‐ and intracellular spaces of the tissues of several edible plants. BE27 is able to enter into the cytosol and kill fungal cells, thus arresting the growth of the fungus. The mechanism of action seems to involve ribosomal RNA (rRNA) N‐glycosylase activity on the sarcin–ricin loop of the major rRNA which inactivates irreversibly the fungal ribosomes, thus inhibiting protein synthesis. We compared the C‐terminus of the BE27 structure with antifungal plant defensins and hypothesize that a structural motif composed of an α‐helix and a β‐hairpin, similar to the γ‐core motif of defensins, might contribute to the specific interaction with the fungal plasma membranes, allowing the protein to enter into the cell.     

Effects of population density on alienicolae of Aphis fabae Scop.: The expression of migratory urge among alatae in the field

Inspection of naturally or artificially infected Hevea roots showed that Forms lignosus can penetrate undamaged roots directly, but does so more readily through wounds or natural openings like lenticels, or through the bases of lateral roots and bark scales. Therefore, Pomes-infected trees should be identified by leaf symptoms rather than by uncovering and inspecting roots, as this generally leads to root injury, which facilitates fungal penetration. Initial fungal entry into host tissue appears to be by mechanical pressure alone, but deeper penetration is through the action of extracellular enzymes. The fungus remains intercellular in the cortex but is intracellular in the woody tissue. Ray cell walls are penetrated mechanically, but the xylem through pits. The time taken for various stages of infection to occur is assessed. The amount of damage done by the fungus to roots and the blocking of xylem vessels by tyloses suggest that yellowing, curling and buckling of leaves on infected trees are drought symptoms and not a reaction to fungal toxins. The host reacts to the invasion of the cortex by forming a cork cambium and to the invasion of the woody tissue by blocking individual cells with phenols and resins, which could be important when breeding disease resistant Hevea root stocks.     

A Suberized Layer in the Cell Wall of Mucilage Cells of Cinnamomum

The ultrastructure of developing and mature mucilage idioblastsin the shoot apex and leaves of Cinnamomum burmanni and Cinnamomumverum (Lauraceae) is described. In all mucilage cells a suberizedlayer is present in the outer cellulosic cell wall from a veryearly stage in development onwards. This represents the firstultrastructurally documented record of a suberized wall layerin mucilage cells. Oil cells, the other type of secretory idioblastsin Lauraceae, commonly have subenzed wall layers, and the presentresults support suggestions of a possible homology of oil andmucilage cells in the so-called primitive angiosperms Suberized layer, mucilage idioblasts, Cinnamomum burmanni     

Detection of an elicitor on infection structures of Puccinia graminis using monoclonal antibodies

The basidiomycetous fungus Puccinia graminis f. sp. tritici causes the stem rust disease of wheat. Resistance of wheat to the fungus is often associated with the hypersensitive reaction of infected host cells. A glycoprotein isolated from germ tube cell walls of the pathogen elicits a hypersensitive-like response when injected into wheat leaves. Infection structures morphologically identical to those grown on wheat were induced in the absence of the host plant, and indirect immunofluorescence together with specific monoclonal antibodies to the elicitor was employed to locate the antigen at fungal infection structures. No binding occurred to germ tubes or appressoria. The antibodies located the antigen only at that part of the fungal infection structure that develops endophytically in nature and, moreover, only at the youngest part of this structure. In rust-infected wheat leaves, the immunolabel appeared only at haustoria, the structures thought to be involved in specific recognition between host and parasite.     

Fungal endophytes in Astromyelon-type (Sphenophyta,Equisetales, Calamitaceae) roots from the Upper Pennsylvanian of France

A distinctive fungal endophyte, Cashhickia acuminata nov. gen. et sp., is described from permineralized calamite roots from the Upper Pennsylvanian Grand-Croix cherts of France. Heavily infected roots contain numerous intracellular hyphae in the outer cortex that arise from a meshwork-like mycelium extending between cortical cells. All intracellular hyphae are oriented toward the root center; none occur on the inner periclinal host cell walls. Other roots of the same type show localized infection by this fungus in which isolated cortical cells contain or give rise to intracellular fungal growth. Within the cortical cells are host responses in the form of callosities that indicate the roots were alive at the time of infection. Other endophytes are present in the same host tissue but are less frequent. The discovery of this association provides the first detailed account on the morphology of a Carboniferous fungal root endophyte, as well as the spatial distribution within the host, and infection pathways within the cortical tissues.     

Actin versus tubulin configuration in arbuscule-containing cells from mycorrhizal tobacco roots

The involvement of the cytoskeleton in symbiotic interactions such as arbuscular mycorrhizas has received little attention. In this paper, we examine the organization of actin in tobacco mycorrhizal roots and compare actin and tubulin patterns within arbuscule-containing cells.
Our results show drastic reorganization of microfilaments and microtubules upon fungal infection and how those new cytoskeletal patterns relate to the host cytoplasm rearrangement and the intracellular fungal structures. Whereas in uninfected cells a network of cortical and perinuclear actin filaments was observed, in infected cells actin filaments closely follow the fungal branches and envelop the whole arbuscule in a dense coating network. Microtubules are less closely connected with the fungus surface. They run across the whole arbuscule mass, linking branches to each other and to the host cell cortex and nucleus.
These major differences between the two cytoskeletal components are used to advance some suggestions concerning their contribution to structural functions in the plant–fungus interactions during the mycorrhizal symbiosis.     

Taxol Determination from Pestalotiopsis pauciseta, a Fungal Endophyte of a Medicinal Plant

Taxol is the most effective antitumor agent developed in the past three decades. It has been used for effective treatment of a variety of cancers. A taxol-producing endophytic fungus Pestalotiopsis pauciseta (strain CHP-11) was isolated from the leaves of Cardiospermum helicacabum and screened for taxol production. The fungus was identified based on the morphology of the fungal culture and the characteristics of the spores and screened for taxol production. The amount of taxol produced by this endophytic fungus was quantified by HPLC and it produced 113.3 mg/L, thus the fungus can serve as a potential material for fungus engineering to improve taxol production. This fungal taxol also had strong anticancer activity against some cancer cells viz., BT 220, H116, Int 407, HL 251 and HLK 210 tested by Apoptotic assay and it is indicated that with the increase of taxol concentration from 0.005–0.05 mmol/L, taxol induced increased cell death through apoptosis.     

Comparative anatomy of oil cells and mucilage cells in the leaves of the Lauraceae in China

The distribution density of oil cells, the morphology and structure of both oil and mucilage cells, and their localization in the mesophyll of 112 species, 5 varieties and 2 forms in 21 genera of the Lauraceae are comparatively studied with the methods of tissue clearing and paraffin sectioning. The results show that there exist obvious differences of the distribution density of oil cells among the species in the Lauraceae. The presence of oil cells and mucilage cells is found to be a marked anatomical feature of the leaves in most of the plants in the Lauraceae. Their distribution in the mesophyll can be divided into 4 types: type Ⅰ , in which only oil cells are present; type Ⅱ, in which both the oil and mucilage cells are present; type Ⅲ, in which only mucilage cells are present; type Ⅳ, in which neither oil cells nor mucilage cells are present. The distribution density of oil cells, the distribution types of oil cells and mucilage cells and their localization in the mesophyll are of some taxonomic value at the specific level in the Lauraceae. In the whole Lauraceae or in some large genera, the evolutionary trend of the distribution types of oil cells and mucilage cells might be as follows: type Ⅰ → type Ⅱ →type Ⅲ →type Ⅳ. The characteristics of the 4 distribution types of oilcells and mucilage cells support the division of two subfamilies in the Lauraceae.     

菱的腺毛发育及分泌活动的超微结构研究

菱的腺毛由单列圆筒状细胞组成,具有短暂的分泌功能。它们起源于叶片远轴面、叶柄的表皮细胞以及苗端茎轴、花柄的表皮细胞。处于分泌期的腺毛细胞其胞质浓厚,液泡化程度小,细胞具丰富的线粒体、高尔基体。腺毛丧失了分泌功能后即发育成为表皮毛。粘液物质由高尔基体分泌小泡携带至腺毛细胞侧壁,经胞吐与渗透结合的方式分泌至细胞外,粘液的化学成分主要为多糖。     

Histological and Ultrastructural Changes in Leaves and Stems of Resistant and Susceptible Chickpea Cultivars to Ascochyta rabiei

The histo- and cytopathological effects in resistant (ILC-195) and susceptible (Canitez-87) chickpea cultivars were examined by light, transmission and scanning electron microscopy 3, 5 and 7 days after inoculation (d.a.i) of seedlings with Ascochyta rabiei. The fungus produced typical appressoria that penetrated both cuticle and stomata. The resistant plants had physical barriers and a cuticle layer against fungal penetration 3 d.a.i. The fungus spread intercellularly and subepidermally in the leaves and stems of susceptible plants 3 d.a.i., and was followed 5 d.a.i. by cell plasmolysis, degeneration of organelles and of cellulose, but not lignified, walls. Pycnidia formation occurred between 5 and 7 d.a.i. 7 d.a.i., organelle degeneration, pycnidia formation and symptom severity increased. Tracheidal elements, including lignified elements, were almost intact in both resistant and susceptible cultivars. In the susceptible plants, lignin cell walls were slightly degraded after 7 days. There was less cell degeneration and pycnidia formation in resistant plants. Some electron-dense large bodies and lipid granules were observed within intracellular fungal hyphae in infected cells of resistant plants 7 d.a.i.     

Asymmetric plant-mediated cross-effects between a herbivorous insect and a phytopathogenic fungus

• 1 Cross‐effects between a herbivorous insect and a phytopathogenic fungus on their common host plant were examined. Specifically, we addressed the questions whether (i) infection of Chinese cabbage leaves by the fungus Alternaria brassicae affects the development and host selection behaviour of the leaf beetle Phaedon cochleariae and (ii) whether herbivory influences host suitability of Chinese cabbage for A. brassicae.
• 2 Feeding on fungus‐infected leaves prolonged larval development and reduced pupal weight of P. cochleariae. Adult beetles avoided feeding and egg deposition on fungus‐infected leaves. In contrast to these local effects, no systemic effect of phytopathogenic infection on the herbivore was detected.
• 3 Herbivory did not influence fungal growth neither locally nor systemically.
• 4 Thus, our results demonstrate an asymmetric relationship between herbivore and fungus. Whereas herbivory had no visible impact on fungal growth, fungal infection of the plant induced local resistance against P. cochleariae.