Constitutively activated barley ROPs modulate epidermal cell size, defense reactions and interactions with fungal leaf pathogens

RHO-like monomeric G-proteins of plants (ROPs, also called RACs), are involved in plant development and interaction with the environment. The barley (Hordeum vulgare) ROP protein HvRACB has been shown to be required for entry of the biotrophic powdery mildew fungus Blumeria graminis f.sp. hordei (Bgh) into living host cells. To get a deeper insight into evolutionarily conserved functions of ROPs in cell polarity and pathogen responses, we stably expressed constitutively activated (CA) mutant variants of different barley ROPs (HvRACB, HvRAC1, HvRAC3) in barley. CA HvROPs induced epidermal cell expansion and/or abolished polarity in tip growing root hairs. All three CA HvROPs enhanced susceptibility of barley to penetration by Bgh whereas only CA HvRAC1 supported whole cell H₂O₂ production in non-penetrated cells. Despite increasing penetration by Bgh, CA HvRAC1 promoted callose deposition at sites of fungal attack and resistance to penetration by Magnaporthe oryzae. The data show an involvement of ROPs in polar growth processes of the monocot barley and in responses to fungal pathogens with different life style.     

Hyperparasitic Attack of Trichothecium roseum on Conidia of Pestalotiopsis funerea

Trichothecium roseum (Tr) has been shown to be a highly effective hyperparasite on conidia of Pestalotiopsis funerea (Pf) in vivo and in vitro. The stages of this spore parasitism are: positive tropism of Tr towards Pf conidia, contact between Tr and Pf, formation of simple or lobed appressoria of Tr on the host conidial surface, penetration of the attacked host cells from the base of the appressoria, development of host-internal, mostly branching parasitic hyphae by Tr, desintegration, lysis and death of the parasitized host cells, exit of Tr from the destroyed host cells and its intensive sporulation over Pf remnants. Pf did not show any defence reactions against the attack by Tr. In addition to the antagonistic activities of Tr against Pf reported previously, which are due to extracellular toxins released by Tr, direct hyperparasitism is a second mechanism of antagonism, which contributes to the successful competitive ability of Tr in this fungal interaction.     

Mycoparasitic relationships

The mode of attack and the infection structures of the necrotrophic mycoparasite, Pythium acanthicum, as well as the responses of various fungal hosts to parasitism were studied using both electron and light microscopy. Many taxonomically distinct fungal hosts were used, though Phycomyces blakesleeanus, Pythium aphanidermatum, Rhizoctonia solani and a basidiomycete identified as Corticium sensu lato were studied in greatest detail. Parasitism was by direct penetration of the fungal host without appressorium formation by the parasite. The host's cells responded to contact by P. acanthicum by forming papillae. The morphological features of the papillae varied with the particular host. In P. blakesleeanus they were comprised of vesicles and segments of cytoplasm entrapped in a fibrillo-granular matrix, while in R. solani and the Corticium basidiomycete they contained considerable amounts of electron-opaque and electron-translucent material. Evidence for both mechanical and enzymatic penetration of the host fungi by the parasite are presented. Details of host wall and septum penetration by the parasite are presented using time-lapse light microscopy with in vivo systems. Many of these stages of parasitism were examined ultrastructurally. Some comparisons of these mycoparasitic relationships are discussed in relation to what is known from the literature about phytoparasitic interactions.     

The Effect of the Common Bunt on the Growth of Wheat Seedlings and Calluses

The growth of wheat (Triticum aestivum L., cv. Zhnitsa) seedlings and calluses infected with spores of common bunt causal agent Tilletia caries (DC) Tul. was studied. Inoculation with the pathogenic fungus enhanced both in vivo and in vitro growth due to cell division activation and cell expansion. These growth characteristics were also retained in wheat calluses infected with T. caries and grown on the hormone-free Murashige and Skoog nutrient medium. This implies the production of hormone-like substances by the fungus itself or by the infected plant. At the phase of fungal penetration into the plants, infection resulted in IAA accumulation in seedlings. Later, the IAA content reduced to a control level and the phytohormone balance was shifted toward cytokinins. Similar changes in the IAA and cytokinin levels were observed during early fungus development on wheat callus tissues. Such a sequence of events is supposed to be required for successful fungal penetration and localization in plant tissues and for the establishment of compatible interactions between the pathogen and the host plant.     

Cellular interactions of Candida albicans with human oral epithelial cells and enterocytes

The human pathogenic fungus Candida albicans can cause systemic infections by invading epithelial barriers to gain access to the bloodstream. One of the main reservoirs of C. albicans is the gastrointestinal tract and systemic infections predominantly originate from this niche. In this study, we used scanning electron and fluorescence microscopy, adhesion, invasion and damage assays, fungal mutants and a set of fungal and host cell inhibitors to investigate the interactions of C. albicans with oral epithelial cells and enterocytes. Our data demonstrate that adhesion, invasion and damage by C. albicans depend not only on fungal morphology and activity, but also on the epithelial cell type and the differentiation stage of the epithelial cells, indicating that epithelial cells differ in their susceptibility to the fungus. C. albicans can invade epithelial cells by induced endocytosis and/or active penetration. However, depending on the host cell faced by the fungus, these routes are exploited to a different extent. While invasion into oral cells occurs via both routes, invasion into intestinal cells occurs only via active penetration.     

Gene genealogies reveal cryptic species and host preferences for the pine fungal pathogen Grosmannia clavigera

Grosmannia clavigera is a fungal pathogen of pine forests in western North America and a symbiotic associate of two sister bark beetles: Dendroctonus ponderosae and D. jeffreyi. This fungus and its beetle associate D. ponderosae are expanding in large epidemics in western North America. Using the fungal genome sequence and gene annotations, we assessed whether fungal isolates from the two beetles inhabiting different species of pine in epidemic regions of western Canada and the USA, as well as in localized populations outside of the current epidemic, represent different genetic lineages. We characterized nucleotide variations in 67 genomic regions and selected 15 for the phylogenetic analysis. Using concordance of gene genealogies and distinct ecological characteristics, we identified two sibling phylogenetic species: Gc and Gs. Where the closely related Pinus ponderosa and P. jeffreyi are infested by localized populations of their respective beetles, Gc is present. In contrast, Gs is an exclusive associate of D. ponderosae mainly present on its primary host‐tree P. contorta; however, in the current epidemic areas, it is also found in other pine species. These results suggest that the host‐tree species and the beetle population dynamics may be important factors associated with the genetic divergence and diversity of fungal partners in the beetle‐tree ecosystems. Gc represents the original G. clavigera holotype, and Gs should be described as a new species.     

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.     

Biosynthesis of galactomannans found in filamentous fungi belonging to Pezizomycotina

The galactomannans (GMs) that are produced by filamentous fungi belonging to Pezizomycotina, many of which are pathogenic for animals and plants, are polysaccharides consisting of α-(1→2)-/α-(1→6)-mannosyl and β-(1→5)-/β-(1→6)-galactofuranosyl residues. GMs are located at the outermost layer of the cell wall. When a pathogenic fungus infects a host, its cell surface must be in contact with the host. The GMs on the cell surface may be involved in the infection mechanism of a pathogenic fungus or the defense mechanism of a host. There are two types of GMs in filamentous fungi, fungal-type galactomannans and O-mannose type galactomannans. Recent biochemical and genetic advances have facilitated a better understanding of the biosynthesis of both types. This review summarizes our current information on their biosynthesis.     

Estimates of biomechanical forces in Magnaporthe grisea

The mechanical actions of the fungus Magnaporthe grisea raise many intriguing questions concerning the forces involved. These include: (1) the material properties of the appressorial wall; (2) the strength of the adhesive that keeps the appressorium anchored to the rice leaf surface; and (3) the forces involved in the penetration process whereby a peg is driven through the host cell wall. In this paper we give order of magnitude estimates for all three of these quantities. A simple Young-Laplace law type argument is used to show that the appressorial wall elastic modulus is of order 10–100 MPa; and an adaptation of standard adhesion theory indicates a lower bound on the strength of the appressorial adhesive to be of the order 500 J/m². Drawing on ideas from plasticity theory and ballistics, estimates of the penetration force raise interesting questions about experiments performed on the penetration of inert substrates by the fungus.     

Natural Occurrence of Seiridium cardinale on Thuja in Germany

Seiridium cardinale (Wagener) Sutton & Gibson (syn. Coryneum cardinale Wagener) is the most dangerous parasitic fungus of cypress and other Gupressaceae, already appearing in epidemic proportions, especially in the mediterranean area. Recently (in 1989, 1990 and 1991) this pathogen has been detected in the northern part of Germany (Hamburg and Schleswig-Holstein) naturally infecting Thuja. The geographical distribution and the progressive spread of S. cardinale is reviewed. Diagnostic features of the fungus, modes of host penetration, principal symptoms of the disease, ways of dispersal and control methods are described. Possible causal factors for the recently observed occurrence of 5. cardinale in the north of Germany are discussed.     

Superoxide Generation in Chemically Activated Resistance of Barley in Response to Inoculation with the Powdery Mildew Fungus

In a previous work, a phenotype-specific accumulation of superoxide radical anions (O^??₂) after attack of the powdery mildew fungus (Blumeria [syn. Erysiphe] graminis f.sp. hordei) in near-isogenic barley (Hordeum vulgare L.) lines bearing different Mlx genes for resistance was described (Hückelhoven and Kogel, 1998). We have now a histochemical study of the pathogenesis-related O^??₂ generation in the systemic activated resistance (SAR) response induced in barley cv Pallas by the plant activator 2,6-dichloroisonicotinic acid (DCINA). SAR-specific defence was conducted prevalently characterized by penetration resistance. Fungal arrest was observed before haustorium formation by a highly localized cell wall reinforcement (effective papillae) and, in most cases, by a subsequent hypersensitive cell death (HR). No O^??₂ generation was found in association with these plant defence responses. However, a strong O^??₂ burst in the attacked epidermal cells was detected in the control plants which were not activated by DCINA. This burst coincided with cell wall penetration and subsequent contact of the pathogen with the host plasma membrane. A strong SAR-related O^??₂ burst was induced in the mesophyll tissue beneath the attacked and hypersensitively reacting epidermal cells in plants treated with DCINA. The accumulation of O^??₂ was confined to chloroplasts. The remarkable burst in mesophyll tissue was not followed by mesophyll-HR indicating that chloroplastic O^??₂ generation is not sufficient for the hypersensitive cell death. Since the same pattern of pathogenesis-related O^??₂ accumulation was identified for race-specific response mediated by the Mlg gene for powdery mildew resistance, the present data are consistent with the hypothesis that the SAR phenotype is a phenocopy of the Mlg-type resistance (Kogel et al., 1994).     

The effect of stem density on the probability of attack by the ambrosia beetle Platypus quercivorus varies with spatial scale

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Estimation of air-borne fungus spores; a comparison of slide and culture methods

Summary A survey of the atmospheric incidence of fungus spores in Sydney was carried out by daily exposure of both plates and slides. The predominating fungi on the plates were in order of frequency,Cladosporium, Alternaria, Epicoccum, Pleospora, Penicillium, Pullularia. A total of 3663 colonies were counted.On the slides, the total number of the fungus spores found for the year was 2043. The predominating fungi wereAlternaria, Cladosporium and rusts and smuts.Working under a grant from the National Health and Medical Research Council of Australia.     

Transformation of Gaeumannomyces graminis with β‐Glucuronidase Reporter and Hygromycin Resistance Genes

Take‐all disease is caused by Gaeumannomyces graminis, (Sacc.) Arx & D. Olivier, a soil‐borne fungus, which colonizes the root and crown tissue of many members of the Poaceae plant family. This fungus is able to grow along the surface of roots as darkly pigmented runner hyphae, which has the ability to penetrate the root. Here, we describe a genetic transformation of G. graminis var. graminis by using polyethylene glycol (PEG)‐based protoplast transformation. Fungus cells were transformed with a plasmid, pHPG, containing the gusA reporter gene that codes for β‐glucuronidase (GUS) and the hph gene for hygromycin resistance as the selectable marker. A de novo transformant selection assay was developed to identify the putative transformants that were expressing the hph gene. In addition, the transformed cells maintained the ability to infect the plant tissues. The GUS‐expressing fungus can be used to study fungal infection processes including fungal penetration, colonization and the role(s) of melanin during pathogenesis. Thus, this study is the first report of G. graminis var. graminis transformed with a visibly detectable reporter gene that provides a useful tool to a better understanding of host–Gaeumannomyces interactions.     

Biotrophic mycoparasitic interactions between Sphaerodes mycoparasitica and phytopathogenic Fusarium species

The objective of this study was to investigate the relationships between the biotrophic mycoparasite Sphaerodes mycoparasitica and pathogenic Fusarium strains. To study the interactions between S. mycoparasitica and four different phytopathogenic Fusarium strains, macroscopic observations were performed using dual-culture assays and microscopic examinations in combination with light and fluorescent microscopy. Both macroscopic and microscopic techniques were also vital in determining the host specificity of S. mycoparasitica with F. avenaceum, F. oxysporum, F. proliferatum, and F. sporotrichioides. Our results suggest that S. mycoparasitica established haustorial contact with F. avenaceum and F. oxysporum. Data obtained from the dual-culture assay and parasitism interactions revealed that this newly described contact biotrophic mycoparasitic fungus was capable of reducing F. avenaceum and F. oxysporum linear growth and size of hyphal cells through infection and penetration.     

Die Ultrastruktur des Penetrationsvorganges von Olpidium brassicae an Kohlrabi-Wurzeln

Zusammenfassung Olpidium brassicae durchdringt mit einem schlauchförmingen, umwandeten Penetrationsfortsatz die Wirtszellwand und durchwächst damit auch das Lumen toter Wirtszellen. Beim Eindringen in das Wirtscytoplasma treten zwei verschiedene Typen von Wirt-Parasit-Reaktionen auf: in plasmareichen Zellen bildet der Wirt eine Callosity in einer unregelmäßigen Einstülpung des Plasmalemmas in das Cytoplasma, in plasmaarmen Calyptrazellen entsteht dagegen ein Rohr aus unbekanntem stark osmophilen Material. Der Pilzprotoplast durchbricht auf unbekannte Weise das Wirtsplasmalemma und tritt in beiden Fällen als von einer einfachen Plasmamembran umgebenes Plasmodium ins Cytoplasma des Wirtes ein.

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Ultrastructure of the penetration process into roots of Brassica oleracea by Olpidium brassicae

Summary Olpidium brassicae penetrates the host cell wall with a wall-coated penetration tube, which too is able to grow through the lumen of dead host cells. Two host-parasite reactions are observed: plasmarich host cells are forming a callosity at the point of penetration, calyptra cells containing only little cytoplasm are forming a strongly osmiophilous tube. The host plasmalemma is penetrated by the fungus protoplast, which in each case finally is located in the host cytoplasm as a plasmodium coated by a single membrane only.

     

Host range of the European leaf sheath mining midge,Lasioptera donacis Coutin (Diptera: Cecidomyiidae), a biological control of giant reed,Arundo donax L.

The fundamental host range of the arundo leafminer, Lasioptera donacis a candidate agent for the invasive weed, Arundo donax was evaluated. L. donacis collects and inserts spores of a saprophytic fungus, Arthrinium arundinis, during oviposition. Larvae feed and develop in the decomposing leaf sheath channel tissue. Thirty-six closely related and economic grass species along with several key habitat associates were evaluated in no-choice tests. L. donacis and its associated saprophyte completed development only on A. donax, in concurrence with published reports from its native range in Mediterranean Europe. The arundo leafminer feeding leads to premature defoliation, constituting a different mode of attack on the host plant as compared to two previously released insects, the arundo wasp and arundo scale, which feed on shoot tips and rhizomes, respectively. Defoliation of A. donax is expected to increase light penetration into stands of A. donax which increases visibility for law enforcement, reduces the survival of cattle fever ticks, and enhance recovery of the native riparian vegetation along the Rio Grande and other habitats where this weed is invasive.     

Melanization of appressoria is critical for the pathogenicity of <Emphasis Type="Italic">Diplocarpon rosae</Emphasis>

Previous studies have shown the role of melanized appressoria in the pathogenicity of various fungi. Diplocarpon rosae is a worldwide outdoor fungal pathogen of rose plants causing black spot disease of rose leaves. To fully understand how this fungus colonizes its host, which is critical for the development of an efficient and sustainable disease management program, we studied the fungal (especially the appressoria) structures of D. rosae in detail at an early stage of infection. Using both microscopic and biochemical analyses, we observed strong melanized appressoria formation localized at the point of D. rosae penetration, which forms the pathogen–plant interface. Treatment of infected plants with melanin biosynthesis inhibitors (MBIs) prevented melanization of D. rosae appressoria and positively correlated with significant reductions in black spot disease symptoms, suggesting that melanization of appressoria might be a critical factor for the pathogenicity of D. rosae. Our findings were confirmed and validated by the lack of melanized appressorial ring formation on an artificial surface and on a D. rosae-non host plant system, Arabidopsis thaliana. Our findings suggest that localized melanization of appressoria is a crucial factor for the pathogenicity of D. rosae and treatment of the fungus with MBIs seems to be a promising disease management alternative for black spot disease of roses.     

Phymatotrichum (cotton) root rot caused by Phymatotrichopsis omnivora: retrospects and prospects

Phymatotrichum (cotton or Texas) root rot is caused by the soil‐borne fungus Phymatotrichopsis omnivora (Duggar) Hennebert. The broad host range of the fungus includes numerous crop plants, such as alfalfa and cotton. Together with an overview of existing knowledge, this review is aimed at discussing the recent molecular and genomic approaches that have been undertaken to better understand the disease development at the molecular level with the ultimate goal of developing resistant germplasm. Taxonomy: Phymatotrichopsis omnivora (Duggar) Hennebert [synonym Phymatotrichum omnivorum (Shear) Duggar] is an asexual fungus with no known sexual stage. Mitosporic botryoblastospores occasionally form on epigeous spore mats in nature, but perform no known function and do not contribute to the disease cycle. The fungus has been affiliated erroneously with the polypore basidiomycete Sistotrema brinkmannii (Bres.) J. Erikss. Recent phylogenetic studies have placed this fungus in the ascomycete order Pezizales. Host range and disease symptoms: The fungus infects most dicotyledonous field crops, causing significant losses to cotton, alfalfa, grape, fruit and nut trees and ornamental shrubs in the south‐western USA, northern Mexico and possibly parts of central Asia. However, this fungus does not cause disease in monocotyledonous plants. Symptoms include an expanding tissue collapse (rot) of infected taproots. In above‐ground tissues, the root rot results in vascular discoloration of the stem and rapid wilting of the leaves without abscission, and eventually the death of the plant. Characteristic mycelial strands of the pathogen are typically present on the root's surface, aiding diagnosis. Pathogenicity: Confocal imaging of P. omnivora interactions with Medicago truncatula roots revealed that infecting hyphae do not form any specialized structures for penetration and mainly colonize cortical cells and eventually form a mycelial mantle covering the root's surfaces. Cell wall‐degrading enzymes have been implicated in penetration and symptom development. Global gene expression profiling of infected M. truncatula revealed roles for jasmonic acid, ethylene and the flavonoid pathway during disease development. Phymatotrichopsis omnivora apparently evades induced host defences and may suppress the host's phytochemical defences at later stages of infection to favour pathogenesis. Disease control: No consistently effective control measures are known. The long‐lived sclerotia and facultative saprotrophism of P. omnivora make crop rotation ineffective. Chemical fumigation methods are not cost‐effective for most crops. Interestingly, no genetic resistance has been reported in any of the susceptible crop species.