Approaching risk assessment of complex disease development in horse chestnut trees: a chemical ecologist's perspective

Abstract The chemo‐ecological predispositions were investigated for the development of a complex disease on the basis of an insect–fungus mutualism using the system of horse chestnuts (Aesculus hippocastanum and Aesculus x carnea), the horse chestnut leaf miner (Cameraria ohridella) and the biotrophic powdery mildew (Erysiphe flexuosa). Both C. ohridella and E. flexuosa can appear on the same horse chestnut leaf tissue simultaneously. The olfactory detection of fungal infection by the insect, its ability to discriminate the potentially mutualistic fungus from other fungi and the impact of fungal infection on insect oviposition were examined. Gas chromatography coupled with mass spectroscopic and electroantennographic detection by C. ohridella (GC‐MS/EAD) was used to assess the olfactory detection of fungal‐infected A. hippocastanum and A. x carnea leaves by C. ohridella. Infection‐related compounds, such as benzyl alcohol, dodecane, tridecane and methyl salicylate as well as fungus‐related C8 compounds, are perceived by C. ohridella. The discrimination of E. flexuosa from another phytopathogenic fungus, such as Guignardia aesculi, is based primarily on the differing pattern of C8 compounds of these fungi. Oviposition on fungal‐infected leaves of A. hippocastanum and leaves treated with fungal‐related compounds showed that C. ohridella is able to respond to the modifications in the leaf volatile profiles of horse chestnuts caused by the different fungal infections. Thus, from the perception point of view, the necessary predispositions for the development of a close insect–fungus relation between the biotrophic fungus E. flexuosa and the leaf‐mining insect C. ohridella are fulfilled. However, decreased oviposition on infected leaves does not enhance the selective contact between the species. As a consequence, an important predisposition for forming an insect–fungus mutualism is not fulfilled by these two species and, according to this approach, the risk of forming a complex disease can be assessed as low.     

Epidemiology,identification and disease management of grape black rot and potentially useful metabolites of black rot pathogens for industrial applications – a review

Phyllosticta ampelicida (teleomorph: Guignardia bidwellii) is the fungal plant pathogen that causes black rot on grapevine. It is able to infect all green, expanding grape tissues. Black rot poses a threat to both yield and wine quality; a severe attack can virtually cause a complete crop loss. The fungus is native to North America, and was spread to Europe at the end of the 19th century. With the beginning of the 21st century, an increasing importance of the disease was observed in several European winegrowing regions. Successful black rot control strategies combine sanitary measures, cultural techniques, growing cultivars with reduced susceptibility and the use of effective fungicides. Berries are most susceptible to infections between flowering and bunch closure and consequently, fungicide applications against black rot need to focus on this period. In this paper, forecast models and decision support systems that help to achieve satisfactory control with a minimum of fungicide input are presented. Black rot in grapevine was reported to be well controlled in field experiments by a broad range of fungicide classes including quinone outside inhibitors (98 ± 3% efficacy), demethylation inhibitors (98 ± 3% efficacy) and dithiocarbamates (92 ± 7% efficacy). Average efficacies of other fungicide classes tested so far ranged from 32 to 69%; meta data on the efficacy of fungicides in numerous field trials are given. At present, black rot causes major problems primarily if vineyard management and fungicide use were reduced or abandoned. Organically managed vineyards are highly affected. Hence, site and cultivar selection as well as cultural measures are of the highest importance especially in organic viticulture. Recent studies showed that some strains formerly classified as G. bidwellii in fact belong to a distinct species (P. parthenocissi). Black rot pathogens produce some phytotoxic secondary metabolites, such as phenguignardic acid, guignardic acid, alaguignardic acid and the guignardianones A, E and F, which could stimulate the development of new herbicides of natural origin.     

Extensive host range of an endophytic fungus, <Emphasis Type="Italic">Guignardia endophyllicola</Emphasis> (anamorph: <Emphasis Type="Italic">Phyllosticta capitalensis</Emphasis>)

Isolation of endophytic species of Guignardia (anamorph: Phyllosticta) from healthy leaves of 94 plants (91 species and 3 varieties) in 69 genera, 42 families, was carried out in a test site (Kyoto Herbal Garden) to investigate the host range of Guignardia endophyllicola (anamorph: Phyllosticta capitalensis). Species of Guignardia and Phyllosticta were isolated from the leaves of 67 plants (66 species and 1 variety) belonging to 54 genera, 38 families. Among them, 53 isolates from different plants belonging to 43 genera in 36 families were similar in morphology, and sequence analysis of internal transcribed spacer (ITS) regions of ribosome DNA revealed these isolates to be conspecific with G. endophyllicola. In addition, this fungus was isolated from leaves of various plants collected in different places in Japan and Thailand. Thus, this endophytic fungus has been revealed to live within various vascular plants, angiosperms, gymnosperms, and pteridophytes.     

Studies on the Japanese species belonging to the genus <Emphasis Type="Italic">Phyllosticta</Emphasis> (1)

As the first report of monographic studies of the genus Phyllosticta in Japan, four new species, Phyllosticta disanthi on Disanthus cercidifolius, P. hoveniicola on Hovenia dulcis, P. ligustricola on Ligustrum obtusifolium, and Phyllosticta alliacea on Allium fistulosum, are described and illustrated. A new teleomorphic state of P. alliorum, Guignardia alliacea, is confirmed.     

Engineering d‐limonene synthase down‐regulation in orange fruit induces resistance against the fungus Phyllosticta citricarpa through enhanced accumulation of monoterpene alcohols and activation of defence

Terpene volatiles play an important role in the interactions between specialized pathogens and fruits. Citrus black spot (CBS), caused by the fungus Phyllosticta citricarpa, is associated with crop losses in different citrus‐growing areas worldwide. The pathogen may infect the fruit for 20–24 weeks after petal fall, but the typical hard spot symptoms appear when the fruit have almost reached maturity, caused by fungal colonization and the induction of cell lysis around essential oil cavities. d ‐Limonene represents approximately 95% of the total oil gland content in mature orange fruit. Herein, we investigated whether orange fruit with reduced d ‐limonene content in peel oil glands via an antisense (AS) approach may affect fruit interaction with P. citricarpa relative to empty vector (EV) controls. AS fruit showed enhanced resistance to the fungus relative to EV fruit. Because of the reduced d ‐limonene content, an over‐accumulation of linalool and other monoterpene alcohols was found in AS relative to EV fruit. A global gene expression analysis at 2 h and 8 days after inoculation with P. citricarpa revealed the activation of defence responses in AS fruit via the up‐regulation of different pathogenesis‐related (PR) protein genes, probably as a result of enhanced constitutive accumulation of linalool and other alcohols. When assayed in vitro and in vivo, monoterpene alcohols at the concentrations present in AS fruit showed strong antifungal activity. We show here that terpene engineering in fruit peels could be a promising method for the development of new strategies to obtain resistance to fruit diseases.     

Pustulan and branched β-galactofuranan from the phytopathogenic fungus Guignardia citricarpa, excreted from media containing glucose and sucrose

Guignardia citricarpa is a phytopathogenic fungus and the causal agent of citrus black spot. Incubation in a semi-defined media resulted in formation of exopolysaccharides [EPS(s)]. A medium containing glucose gave rise to a (1→6)-linked β-glucan (200 kD), pustulan, which was characterized by NMR and methylation analysis. A sucrose-containing medium provided a homogalactan (376 kD) and methylation analysis showed nonreducing end- (20%), 6-O- (53%) and 5,6-di-O-substituted Galf units (27%). An HMQC spectrum of the homogalactan showed C-1/H-1 signals at δ 108.2/4.820, 108.3/4.820 and 107.1/5.079, corresponding to three types of β- -Galf units. A DEPT analysis showed inverted signals (CH₂) at δ 67.8 and 67.2, corresponding to 6-O-substituted β- -Galf units, whereas a C-5 signal at δ 77.0 suggests 5-O-substitution, confirming a novel structure for a β-galactofuranan.     

山苍子叶内生真菌的纯化与鉴定

本研究选用PDA培养基,通过组织块分离法从山苍子叶中分离得到两株内生真菌。对照真菌鉴定手册,根据菌株和菌丝体形态学特征,并给合ITS区段的碱基序列分析,鉴定两株内生分别属于顶孢霉属和芒果球座菌属。     

Prepenetration Stages of Guignardia psidii in Guava: Effects of Temperature, Wetness Duration and Fruit Age

This study examined the effects of temperature and wetness duration in vitro and in vivo as well as the effects of fruit age on germination and appressoria formation by conidia of Guignardia psidii , the causal agent of black spot disease in guava fruit. The temperatures tested for in vitro and in vivo experiments were 10, 15, 20, 25, 30, 35 and 40°C. The wetness periods studied were 6, 12, 24, 36 and 48 h in vitro and 6, 12 and 24 h in vivo . Fruit 10, 35, 60, 85 and 110-days old were inoculated and maintained at 25°C, with a wetness period of 24 h. Temperature and wetness duration affected the variables evaluated in vitro and in vivo . All variables reached their maximum values at between 25 and 30°C with a wetness duration of 24 h in vivo and 48 h in vitro . These conditions resulted in 31.3% conidia germination, 33.6% appressoria formation and 32.5% appressoria melanization in vitro , and 50.4% conidia germination and 9.5% appressoria formation in vivo . Fruit age also influenced these factors. As fruit age increased, conidia germination and appressoria formation gradually increased. Conidia germination and appressoria formation were 10.8% and 2.3%, respectively, in 10-day-old fruits. In 110-day-old fruits, conidia germination and appressoria formation were 42.5% and 23.2% respectively.     

Diverse non-mycorrhizal fungal endophytes inhabiting an epiphytic,medicinal orchid (<Emphasis Type="Italic">Dendrobium nobile</Emphasis>): estimation and characterization

Although the terrestrial and temperate orchids–fungal biology have been largely explored, knowledge of tropical epiphytic orchids–fungus relationships, especially on the ecological roles imparted by non-mycorrhizal fungal endophytes, is less known. Exploitation of the endophytic fungal mycobiota residing in epiphytic orchid plants may be of great importance to further elucidate the fungal ecology in this special habitat as well as developing new approaches for orchid conversations. The composition of fungal endophytes associated with leaves, stems and roots of an epiphytic orchid (Dendrobium nobile), a famous Chinese traditional medicinal plant, was investigated. Microscopic imaging, culture-dependant method and molecular phylogeny were used to estimate their entity and diversity. Totally, there were 172 isolates, at least 14 fungal genera and 33 different morphospecies recovered from 288 samples. Ascomycetes, coelomycetes and hyphomycetes were three major fungal groups. There were higher overall colonization and isolation rates of endophytic fungi from leaves than from other tissues. Guignardia mangiferae was the dominant fungal species within leaves; while the endophytic Xylariaceae were frequently observed in all plant tissues; Colletotrichum, Phomopsis and Fusarium were also frequently observed. Phylogenetic analysis based on ITS gene revealed the high diversity of Xylariacea fungi and relatively diverse of non-Xylariacea fungi. Some potentially promising beneficial fungi such as Clonostachys rosea and Trichoderma chlorosporum were found in roots. This is the first report concerning above-ground and below-ground endophytic fungi community of an epiphytic medicinal orchid, suggesting the ubiquitous distribution of non-mycorrhizal fungal endophytes in orchid plants together with heterogeneity and tissue specificity of the endophyte assemblage. Possible physiological functions played by these fungal endophytes and their potential applications are also discussed briefly. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.