Transcriptome profiling of Fraxinus excelsior genotypes infested by emerald ash borer.

European ash, Fraxinus excelsior is facing the double threat of ongoing devastation by the invasive fungal pathogen, Hymenoscyphus fraxineus and the imminent arrival of the non-native emerald ash borer (EAB), Agrilus planipennis. The spread of EAB which is currently moving westwards from European Russia and Ukraine into central Europe, poses an additional substantial threat to European ash, F. excelsior. While the molecular basis for resistance or variation in resistance among European ash genotypes is heavily investigated, comparatively little is known about the molecular ash traits involved in resistance against EAB. In this study we have gathered transcriptomic data from EAB inoculated genotypes of F. excelsior that have previously shown different levels of susceptibility to EAB. Resultant datasets show differential gene expression in susceptible and resistant genotypes in response to EAB infestation. This data will provide important information on the molecular basis of resistance to the EAB and allow the development of management plans to combat a pending threat of a culturally and ecologically important European tree species.

Fungal Endophytes Isolated from Elymus repens, a Wild Relative of Barley, Have Potential for Biological Control of Fusarium culmorum and Pyrenophora teres in Barley.

Twenty-four fungal endophytes, isolated from a wild relative of barley, Elymus repens, were screened in barley against an isolate of Fusarium culmorum and an isolate of Pyrenophora teres under controlled conditions. In all experiments, the endophytes were applied individually as seed dressings. Five endophytes could significantly reduce symptoms of Fusarium culmorum (Periconia macrospinosa E1 and E2, Epicoccum nigrum E4, Leptodontidium sp. E7 and Slopeiomyces cylindrosporus E18). In particular, treatment with Periconia macrospinosa E1 significantly reduced Fusarium symptoms on roots by 29-63% in two out of four experiments. Using, a gfp transformed isolate of P. macrospinosa E1, it was possible to show that the fungus was present on roots 14 days after sowing, coinciding with the disease scoring. To test for a potential systemic effect of the seed treatment, eight endophyte isolates were tested against the leaf pathogen Pyrenophora teres. Three isolates could significantly reduce symptoms of P. teres (Lasiosphaeriaceae sp. E10, Lindgomycetaceae sp. E13 and Leptodontidium sp. E16). Seed treatment with Lasiosphaeriaceae sp. E10 reduced net blotch leaf lesion coverage by 89%, in one out of three experiments. In conclusion, specific endophyte isolates exerted varying degrees of protection in the different experiments. Nevertheless, data suggest that endophytic strains from E. repens in a few cases are antagonistic against F. culmorum and P. teres, but otherwise remain neutral when introduced to a barley host in a controlled environment.

The Bacillus cereus Strain EC9 Primes the Plant Immune System for Superior Biocontrol of Fusarium oxysporum.

Antibiosis is a key feature widely exploited to develop biofungicides based on the ability of biological control agents (BCAs) to produce fungitoxic compounds. A less recognised attribute of plant-associated beneficial microorganisms is their ability to stimulate the plant immune system, which may provide long-term, systemic self-protection against different types of pathogens. By using conventional antifungal in vitro screening coupled with in planta assays, we found antifungal and non-antifungal Bacillus strains that protected the ornamental plant Kalanchoe against the soil-borne pathogen Fusarium oxysporum in experimental and commercial production settings. Further examination of one antifungal and one non-antifungal strain indicated that high protection efficacy in planta did not correlate with antifungal activity in vitro. Whole-genome sequencing showed that the non-antifungal strain EC9 lacked the biosynthetic gene clusters associated with typical antimicrobial compounds. Instead, this bacterium triggers the expression of marker genes for the jasmonic and salicylic acid defence pathways, but only after pathogen challenge, indicating that this strain may protect Kalanchoe plants by priming immunity. We suggest that the stimulation of the plant immune system is a promising mode of action of BCAs for the development of novel biological crop protection products.

Site-specific, silicon-induced structural and molecular defence responses against powdery mildew infection in roses.

BACKGROUND: Silicon (Si) application to miniature potted roses can decrease severity of powdery mildew (Podosphaera pannosa) and this is associated with increased accumulation of callose and hydrogen peroxide (H2 O2 ) as well as hypersensitive (HR) cells. We used microscopy, gene expression and specific inhibitors of callose and H2 O2 to determine how effective these plant responses are in stopping infection. RESULTS: Pathogen arrest in Si-treated (Si+) plants was accompanied by increased accumulation of callose and H2 O2 in papillae and HR cells, respectively. These responses were reduced by application of specific inhibitors (2-deoxy-d-glucose for callose and catalase for H2 O2 ), which increased disease severity in Si+, but not in Si- plants. As markers for HR and callose, expression of the HR-specific gene hsr203J and the wound-related callose synthase GSL5, respectively, was studied. An up-regulation of expression was only seen after isolation of HR cells with laser capture microdissection. The up-regulation was higher in Si+ than in Si- plants and occurred concomitantly with more efficient photosynthesis in Si+ plants at high disease severity as compared to Si- plants. CONCLUSION: Silicon-mediated activation of callose and H2 O2 are decisive factors in the defence of rose against P. pannosa and these responses were accompanied with more efficient photosynthesis to strengthen the plant. Only by isolation of HR cells using laser capture microdissection as compared to analysis of whole leaf tissues allowed detection of elevated transcript levels of hsr203J and GSL5 at infection sites as markers for HR. © 2021 Society of Chemical Industry.

Succession of the fungal endophytic microbiome of wheat is dependent on tissue-specific interactions between host genotype and environment.

Fungi living inside plants affect many aspects of plant health, but little is known about how plant genotype influences the fungal endophytic microbiome. However, a deeper understanding of interactions between plant genotype and biotic and abiotic environment in shaping the plant microbiome is of significance for modern agriculture, with implications for disease management, breeding and the development of biocontrol agents. For this purpose, we analysed the fungal wheat microbiome from seed to plant to seeds and studied how different potential sources of inoculum contributed to shaping of the microbiome. We conducted a large-scale pot experiment with related wheat cultivars over one growth-season in two environments (indoors and outdoors) to disentangle the effects of host genotype, abiotic environment (temperature, humidity, precipitation) and fungi present in the seed stock, air and soil on the succession of the endophytic fungal communities in roots, flag leaves and seeds at harvest. The communities were studied with ITS1 metabarcoding and environmental climate factors were monitored during the experimental period. Host genotype, tissue type and abiotic factors influenced fungal communities significantly. The effect of host genotype was mostly limited to leaves and roots, and was location-independent. While there was a clear effect of plant genotype, the relatedness between cultivars was not reflected in the microbiome. For the phyllosphere microbiome, location-dependent weather conditions factors largely explained differences in abundance, diversity, and presence of genera containing pathogens, whereas the root communities were less affected by abiotic factors. Our findings suggest that airborne fungi are the primary inoculum source for fungal communities in aerial plant parts whereas vertical transmission is likely to be insignificant. In summary, our study demonstrates that host genotype, environment and presence of fungi in the environment shape the endophytic fungal community in wheat over a growing season.

Bioimaging structural signatures of the oomycete pathogen Sclerospora graminicola in pearl millet using different microscopic techniques.

In this case study, the mycelium growth of Sclerospora graminicola in the infected tissues of pearl millet and the process of sporulation and liberation of sporangia and zoospores were observed using four different microscopic techniques. The cotton blue-stained samples observed under light microscope revealed the formation of zoospores with germ tubes, appressoria and initiation of haustorium into the host cells, while the environmental scanning electron microscopy showed the rapid emergence of sporangiophores with dispersed sporangia around the stomata. For fluorescence microscopy, the infected leaf samples were stained with Fluorescent Brightener 28 and Calcofluor White, which react with ß-glucans present in the mycelial walls, sporangiophores and sporangia. Calcoflour White was found to be the most suitable for studying the structural morphology of the pathogen. Therefore, samples observed by confocal laser scanning microscopy (CLSM) were pre-treated with Calcofluor White, as well as with Syto-13 that can stain the cell nuclei. Among the four microscopic techniques, CLSM is ideal for observing live host-pathogen interaction and studying the developmental processes of the pathogen in the host tissues. The use of different microscopic bioimaging techniques to study pathogenesis will enhance our understanding of the morphological features and development of the infectious propagules in the host.

Prevalence of Soil-borne Diseases in Kalanchoe blossfeldiana Reveals a Complex of Pathogenic and Opportunistic Fungi.

Greenhouse cultivation of ornamentals is subjected to a high incidence of soil-borne fungal pathogens. In Kalanchoe, these pathogens are responsible for root and stem rot, and for infection of the vascular tissue. Well-known soil-borne pathogens are believed to cause these diseases. Yet, a systematized survey of these pathogens is lacking for Kalanchoe produced commercially. Here, we studied the occurrence of soil-borne fungal pathogens associated with cultivation of Kalanchoe in Denmark and production of cuttings and stock plants in greenhouse facilities located in Turkey and Vietnam. Molecular identification of pathogens complemented mycological identification and pathogenicity testing of the soil-borne fungal pathogens. This study revealed that the fungi Corynespora cassiicola, Thielaviopsis basicola, Fusarium solani, and F. oxysporum are the most prevalent pathogens associated with root and stem rotting and wilt of Kalanchoe under the conditions studied. Furthermore, the study showed that some of the pathogens are part of an infection complex comprising both primary and opportunistic fungal species. The fact that some of the pathogens were present in some regions, while absent in others, suggests how they move between greenhouse facilities on infected plant material. This study generated important information about the soil-borne fungal complex affecting Kalanchoe, which is useful for a better understanding of the biology of the disease and for designing control strategies.

The barley powdery mildew candidate secreted effector protein CSEP0105 inhibits the chaperone activity of a small heat shock protein.

Pathogens secrete effector proteins to establish a successful interaction with their host. Here, we describe two barley (Hordeum vulgare) powdery mildew candidate secreted effector proteins, CSEP0105 and CSEP0162, which contribute to pathogen success and appear to be required during or after haustorial formation. Silencing of either CSEP using host-induced gene silencing significantly reduced the fungal haustorial formation rate. Interestingly, both CSEPs interact with the barley small heat shock proteins, Hsp16.9 and Hsp17.5, in a yeast two-hybrid assay. Small heat shock proteins are known to stabilize several intracellular proteins, including defense-related signaling components, through their chaperone activity. CSEP0105 and CSEP0162 localized to the cytosol and the nucleus of barley epidermal cells, whereas Hsp16.9 and Hsp17.5 are cytosolic. Intriguingly, only those specific CSEPs changed localization and became restricted to the cytosol when coexpressed with Hsp16.9 and Hsp17.5, confirming the CSEP-small heat shock protein interaction. As predicted, Hsp16.9 showed chaperone activity, as it could prevent the aggregation of Escherichia coli proteins during thermal stress. Remarkably, CSEP0105 compromised this activity. These data suggest that CSEP0105 promotes virulence by interfering with the chaperone activity of a barley small heat shock protein essential for defense and stress responses.

Silicon-induced changes in antifungal phenolic acids, flavonoids, and key phenylpropanoid pathway genes during the interaction between miniature roses and the biotrophic pathogen Podosphaera pannosa.

Application of 3.6 mm silicon (Si+) to the rose (Rosa hybrida) cultivar Smart increased the concentration of antimicrobial phenolic acids and flavonoids in response to infection by rose powdery mildew (Podosphaera pannosa). Simultaneously, the expression of genes coding for key enzymes in the phenylpropanoid pathway (phenylalanine ammonia lyase, cinnamyl alcohol dehydrogenase, and chalcone synthase) was up-regulated. The increase in phenolic compounds correlated with a 46% reduction in disease severity compared with inoculated leaves without Si application (Si-). Furthermore, Si application without pathogen inoculation induced gene expression and primed the accumulation of several phenolics compared with the uninoculated Si- control. Chlorogenic acid was the phenolic acid detected in the highest concentration, with an increase of more than 80% in Si+ inoculated compared with Si- uninoculated plants. Among the quantified flavonoids, rutin and quercitrin were detected in the highest concentrations, and the rutin concentration increased more than 20-fold in Si+ inoculated compared with Si- uninoculated plants. Both rutin and chlorogenic acid had antimicrobial effects on P. pannosa, evidenced by reduced conidial germination and appressorium formation of the pathogen, both after spray application and infiltration into leaves. The application of rutin and chlorogenic acid reduced powdery mildew severity by 40% to 50%, and observation of an effect after leaf infiltration indicated that these two phenolics can be transported to the epidermal surface. In conclusion, we provide evidence that Si plays an active role in disease reduction in rose by inducing the production of antifungal phenolic metabolites as a response to powdery mildew infection.

Disease-reducing effect of Chromolaena odorata extract on sheath blight and other rice diseases.

Sheath blight caused by Rhizoctonia solani (teleomorph: Thanatephorus cucumeris) is a major cause of crop loss in intensive rice production systems. No economically viable control methods have been developed. We screened aqueous extracts of common herbal plants that could reduce sheath blight lesions and found that foliar spraying and seed soaking application of extracts of either fresh or dried leaves of Chromolaena odorata gave up to 68% reduction in sheath blight lesion lengths under controlled and semi-field conditions. The observed reductions were not dependent on growth conditions of C. odorata and rice cultivar. The effect was observed until 21 days after inoculation and was not dependent on microbial activity. Under semi-field conditions, extracts also reduced severity of other important rice diseases, i.e., blast (Pyricularia oryzae) using foliar spray (up to 45%), brown spot (Bipolaris oryzae) using seed treatment (up to 57%), and bacterial blight (Xanthomonas oryzae pv. oryzae) using both application methods (up to 50%).

Role of hydrogen peroxide during the interaction between the hemibiotrophic fungal pathogen Septoria tritici and wheat.

Hydrogen peroxide (H(2)O(2)) is reported to inhibit biotrophic but benefit necrotrophic pathogens. Infection by necrotrophs can result in a massive accumulation of H(2)O(2) in hosts. Little is known of how pathogens with both growth types are affected (hemibiotrophs). The hemibiotroph, Septoria tritici, infecting wheat (Triticum aestivum) is inhibited by H(2)O(2) during the biotrophic phase, but a large H(2)O(2) accumulation occurs in the host during reproduction. Here, we infiltrated catalase, H(2)O(2) or water into wheat during the biotrophic or the necrotrophic phase of S. tritici and studied the effect of infection on host physiology to get an understanding of the survival strategy of the pathogen. H(2)O(2) removal by catalase at both early and late stages made plants more susceptible, whereas H(2)O(2) made them more resistant. H(2)O(2) is harmful to S. tritici throughout its life cycle, but it can be tolerated. The late accumulation of H(2)O(2) is unlikely to result from down-regulation of photosynthesis, but probably originates from damage to the peroxisomes during the general tissue collapse, which is accompanied by release of soluble sugars in a susceptible cultivar.