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This study combined phytosanitary surveys, laboratory analyses, and mathematical modeling to show how hail-induced wounds can foster the infections of the blight pathogen Cryphonectria parasitica, locally associated with extensive dieback of chestnut (Castanea sativa). Orchards and coppices located within and outside the assessed dieback area in a single location in the North West of Italy were inspected to appraise the abundance of hail-induced wounds and C. parasitica infections. The incidence of C. parasitica was significantly higher within the dieback area compared with outside (92% versus 60%; P < 0.05). Hail-induced wounds were observed on small branches and shoots of all trees sampled within the dieback area, whereas they were less abundant outside (20% of trees), suggesting either that the dieback was directly associated with the injuries caused by the hailstorms or that those injuries may have facilitated infections of C. parasitica. Isolations conducted on 359 branches and shoots showed that hail-induced wounds served as infection courts for C. parasitica and that infections depended on the size rather than on the number of hail wounds. We fitted a logistic model showing that hail-induced wounds whose perimeter was larger than 66 mm were at particular risk of C. parasitica infection. A newly designed geometrical-based model is proposed to relate hailstones size, hail wound perimeter, and the risk of infection. We established that hail-induced wounds are entry points for virulent and hypovirulent strains of C. parasitica, since 6.5% of isolates were infected by Cryphonectria hypovirus-1.

Stumps play a pivotal role in the epidemiology of the fungal forest pathogens Heterobasidion spp. because they are the main courts of primary airborne infections. The aims of this study were (i) to determine the susceptibility of seven tree species (i.e., Larix sibirica, Picea abies, Picea sitchensis, Pinus contorta, Pinus strobus, Pinus sylvestris and Pseudotsuga menziesii) to primary infection by H. annosum and H. parviporum through comparative inoculation experiments of conidia on wood discs in controlled conditions, (ii) to compare the susceptibility of wood discs of the same tree species to natural airborne infections in two Latvian Norway spruce forest stands infested either by H. annosum or H. parviporum, (iii) to explore the rates of infection of wood discs at increasing distances from spore sources in these two forests to make inferences on the range of spores dispersal. Results obtained by spraying wood discs with conidial suspensions in controlled conditions are in agreement with those obtained by exposing wood discs to the natural airborne inoculum in the forests, as clearly supported by the significant correlation (r = 0.79, p &lt, 0.05) between the two sets of data. Susceptibility was highest in Pinus species, followed by P. abies and P. sitchensis. Susceptibility was lowest for L. sibirica and P. menziesii. The area colonized by Heterobasidion spp. in the sapwood of wood discs was much greater than that colonized in the heartwood. A sharp decrease in the rate of infection of wood discs with distance from spore sources (i.e., fruiting bodies) was observed, further confirming the importance of local spore sources in the epidemiology of Heterobasidion spp. Taken together, these findings could help designing tactics to manage these fungal forest pathogens.

The Heterobasidion annosum species complex includes major fungal pathogens of conifers worldwide. State-of-the-art preventative stump treatments with urea or with commercial formulations of the fungal biological control agent Phlebiopsis gigantea (i.e., Rotstop®) may become no longer available or are not approved for use in many areas of Europe infested by the three native Heterobasidion species and by the North American invasive H. irregulare, making the development of new treatments timely. The efficacy of Proradix® (based on Pseudomonas protegens strain DSMZ 13134), the cell-free filtrate (CFF) of the same bacterium, a strain of P. gigantea (MUT 6212) collected in the invasion area of H. irregulare in Italy, Rotstop®, and urea was comparatively investigated on a total of 542 stumps of Abies alba, Picea abies, Pinus pinea, and P. sylvestris in forest stands infested by the host-associated Heterobasidion species. Additionally, 139 logs of P. pinea were also treated. Results support the good performances of Rotstop®, and especially of urea against the native Heterobasidion species on stumps of their preferential hosts and, for the first time, towards the invasive North American H. irregulare on stumps of P. pinea. In some experiments, the effectiveness of Proradix® and of the strain of P. gigantea was weak, whereas the CFF of P. protegens strain DSMZ 13134 performed as a valid alternative to urea and Rotstop®. The mechanism of action of this treatment hinges on antibiosis; therefore, further improvements could be possible by identifying the active molecules and/or by optimizing their production. Generally, the performance of the tested treatments is not correlated with the stump size.

Two chestnut (Castanea sativa) orchards of north-western Italy were sampled with passive spore traps 35 times over 24 months. Samples were analysed through a newly developed quantitative PCR assay to quantify propagule loads of the emerging fungal pathogen Gnomoniopsis castaneae. Average propagule deposition patterns were assessed along with temporal and climatic variables, including sampling month and season, temperatures, relative humidity, precipitations, and wind. Machine learning algorithms combining information theory, fractal analysis, unbiased recursive partitioning, ordinary least squares and logistic regressions, were used to model propagule deposition patterns. The trained models were validated on independent data gathered from 24 samplings conducted in a third chestnut orchard during the same timeframe. Results showed that propagule deposition rate (DR) was variable within and among sites, with a site average ranging from 173 to 765 spores ?m ?h. Propagule deposition was observed across all seasons, although the DR dropped substantially during wintertime (p

International audience; Gnomoniopsis castaneae is an emerging fungal pathogen currently scored as the major nut rot agent on chestnut, although it is also associated with cankers on both chestnut and hazelnut, as well as with necrosis on chestnut galls and leaves. Described for the first time in 2012, G. castaneae has been reported in several countries across Europe, Asia and Australasia, often in relation to severe outbreaks. The goal of this review is to provide a comprehensive summary of the state of the art about G. castaneae, highlighting the main results achieved by the research and stressing the most relevant knowledge gaps that still need to be filled. This overview includes topics encompassing the taxonomy of the fungal pathogen, its host range and geographic distribution, the symptomatology and the diagnostic methods available for its detection, its impact, biology, ecology and epidemiology. The main interactions between G. castaneae and other organisms are also discussed, as well as the possible control strategies. In these past few years, relevant progresses in the knowledge of G. castaneae have been achieved, yet the complexity of the challenges that this pathogen poses to chestnut growers and to the scientific community advocates for further advances.

The role of ectomycorrhizal fungi in modulating host susceptibility to fungal pathogens is poorly known. Pinus sylvestris susceptibility to two congeneric fungal pathogens was compared between mycorrhizal and non-mycorrhizal plants. The ectomycorrhizal fungus was Suillus luteus, while the pathogens were Heterobasidion irregulare and H. annosum. H. irregulare is native to North-America and invasive to Europe, whereas S. luteus and H. annosum are native to Eurasia. Non-mycorrhizal plants were equally susceptible to both pathogens. Mycorrhizal plants were significantly less susceptible to H. annosum than non-mycorrhizal plants, whereas there were no differences when considering H. irregulare. The abundance of ectomycorrhizas was negatively and significantly correlated to the level of host susceptibility only to H. annosum. This study shows that the protective role of ectomycorrhizal fungi may differ when the host is challenged by congeneric fungal pathogens. Results add a further clue to explain and predict the invasiveness of H. irregulare in Europe. (C) 2018 Elsevier Ltd and British Mycological Society. All rights reserved. This work was supported by the Italian Ministry of Education, University and Research, within the FIRB program (grant number RBFR1280NN). Authors are grateful to Francois Rineau for assistance with mycorrhiza inoculations and to Matteo Garbelotto for helpful comments on the manuscript and for revising the English. The authors wish to thank the Editor and the anonymous Reviewers for insightful suggestions on how to improve the manuscript.

The biocontrol potential of a commercial product based on the bacterium Pseudomonas protegens (strain DSMZ 13134) and of its cell-free filtrate (CFF) was tested under controlled conditions in vitro and on wood discs against genotypes of the three native European Heterobasidion species and on the non-native H. irregulare, all reported as destructive forest pathogens in Europe. In vitro experiments through traditional dual culture assays and by incorporating CFF into the media showed that treatments were effective in reducing significantly and regardless of the culture medium and incubation temperature both mycelial growth and rate of conidial germination of Heterobasidion spp., and that antibiosis could be the main mechanism involved in the inhibition of Heterobasidion spp. Outcomes of dual culture assays performed on two-divided Petri plates further suggest that antibiosis operates through the production of diffusible rather than volatile compounds. Based on comparative experiments on wood discs of preferential hosts of Heterobasidion spp., CFF performed significantly better than the commercial product against most of Heterobasidion spp., further confirming that the antagonistic activity hinges on antibiosis rather than on other mechanisms. While on wood discs the tested treatments seem poorly effective against the non-native H. irregulare, CFF significantly and substantially reduced infections of the native European Heterobasidion species compared to controls mostly to the level of the state of the art chemical treatment urea and of the biological treatment based on Phlebiopsis gigantea.

In plant pathosystems in which some hosts are transmissive and some are dead-ends, infection is mediated by multiple factors including the susceptibility of both hosts, the sporulation potential of transmissive hosts, the mobility of infectious propagules, the presence of environmental factors conducive to infection, and the variability in distribution of both host types. The factors above were studied for the California forest disease sudden oak death caused by the pathogen Phytophthora ramorum. This pathogen is exotic to California, and while it sporulates at significant levels on the leaves of California bay laurels, four susceptible oak species appear to be non-infectious dead-end hosts. Here, we report, for the first time, on inoculum levels necessary to successfully infect adult oaks and on the distribution of such inoculum levels through time and space thanks to a seven-year-long monitoring effort across a network of 128 monitoring points. Through a series of geostatistical and statistical analyses, we show that the presence of high inoculum loads is positively correlated with close proximity to bay laurels, with high rainfall levels, and with warmer temperatures. Data are consistent with splash dispersal of the pathogen and show that increased presence of tanoak corresponds to a reduced presence of bay laurels and to a lower frequency of high inoculum events. Removal of bay laurels resulted in a substantial decrease of number of events in which spore loads were high enough to infect oaks. This effect was significant when bays were removed 10 m around sampling points, thus indicating that removal of bays 10 m around oaks is a valid approach to reduce infections of oaks.

The effects of plant symbionts on host defence responses against pathogens have been extensively documented, but little is known about the impact of pathogens on the symbiosis and if such an impact may differ for nonnative and native pathogens. Here, this issue was addressed in a study of the model system comprising Pinus pinea, its ectomycorrhizal symbiont Tuber borchii, and the nonnative and native pathogens Heterobasidion irregulare and Heterobasidion annosum, respectively. In a 6-month inoculation experiment and using both in planta and gene expression analyses, we tested the hypothesis that H. irregulare has greater effects on the symbiosis than H. annosum. Although the two pathogens induced the same morphological reaction in the plant-symbiont complex, with mycorrhizal density increasing exponentially with pathogen colonization of the host, the number of target genes regulated in T. borchii in plants inoculated with the native pathogen (i.e. 67% of tested genes) was more than twice that in plants inoculated with the nonnative pathogen (i.e. 27% of genes). Although the two fungal pathogens did not differentially affect the amount of ectomycorrhizas, the fungal symbiont perceived their presence differently. The results may suggest that the symbiont has the ability to recognize a self/native and a nonself/nonnative pathogen, probably through host plant-mediated signal transduction.

An emerging nut rot of chestnut caused by the fungus Gnomoniopsis castaneae was reported soon after the invasion of the exotic gall wasp Dryocosmus kuriphilus in Italy. The goal of this work was to assess the association between the spread of the fungal pathogen and the infestation of the pest by testing if: I) viable inoculum of G. castaneae can be carried by adults of D. kuriphilus ; II) the fungal colonization is related to the number of adults inhabiting the galls; III) the fungal colonization of chestnut buds and the oviposition are associated. Fungal isolations and PCR-based molecular assays were performed on 323 chestnut galls and on their emerging D. kuriphilus adults, whose number was compared between galls colonized and not colonized by G. castaneae . To test the association between fungal colonization and oviposition, Monte Carlo simulations assuming different scenarios of ecological interactions were carried out and validated through isolation trials performed on 597 and 688 chestnut buds collected before and after oviposition, respectively. Although DNA of G. castaneae was detected in a sample of 40% of the adults developed in colonized galls, the fungus could never be isolated from insects, suggesting that the pest is an unlikely vector of viable inoculum. On average, the emerging adults were significantly more abundant from galls colonized by G. castaneae than from not colonized ones (3.76 vs. 2.54, P The simulations implying no interaction between G. castaneae and D. kuriphilus after fungal colonization were confirmed as the most likely. In fact, G. castaneae was present in 33.8% of the buds before oviposition, while no association was detected between fungal colonization and oviposition (odds ratio 0.98, 0.71–1.33 95% CI). These findings suggest that the fungus/pest synergy is asymmetrically favorable to the pest and occurs after oviposition.

Based on a comprehensive literature analysis, we present a critical review of those optical remote sensing techniques operating with the visible (VIS) and near infrared (NIR) bands for the assessment of health in forest trees. Physical, biological and physio-pathological issues of VIS-NIR reflectance of leaves are described pointing out that a decrease of NIR reflectance is highly influenced by stress conditions on tree caused by abiotic and biotic factors. In many cases the NIR spectral band is more sensitive than the VIS one, allowing to detect plant stress long before the appearance of visible symptoms. A description of the main remote sensing methods is provided, including radiometric measurements and multispectral imaging approaches. False colour infrared (FCIR) images collection and their photointerpretation and processing are shown as they represent the most relevant means to acquire information of canopy from its reflectance properties. The amount and the quality of the obtainable data depend on: (i) field conditions; (ii) the type of the adopted instrument (camera, radiometer); (iii) the recording system position (ground platforms, aircraft, satellite); (iv) the format of the data (analogical, digitalised or digital); and (v) the photointerpretation technique. Results from literature are discussed stressing the limits of remote sensing methods. Remote sensing in VIS and NIR spectral bands is generally a powerful classification tool to detect and score tree stress. Nevertheless, it is not a diagnostic tool in that it does not provide information on the cause of stress. Moreover, the method should be adequately tested at single tree level for many important pathogens, in particular root rot, butt rot and stem rot fungi. In perspective, new high spatial resolution satellite images and their GIS software elaboration might be suitable to improve remote sensing analysis.

Summary A total of 2257 recently felled stumps were sampled from 22 mixed naturally regenerated forest stands in the Aosta Valley, western Italian Alps, and examined for Heterobasidion annosum butt rots. Disease incidence ranged from 6% to 71% depending on sites, and H. annosum accounted for 92% of the total number of diseased stumps. H. annosum incidence was significantly higher (Chi-squared multiple comparison tests, p

The effects of biological invasions by non-native species have been widely studied in terms of environmental, economic, and human health impacts. However, little is known about the consequences that non-native plant pathogens may determine on host plant symbionts, such as ectomycorrhizal (ECM) fungi. In this study, interactions between Suillus luteus, an ECM fungus of pine trees, and the allopatrically differentiated fungal pathogens of pines Heterobasidion irregulare and H. annosum were investigated in dual culture by morphological and gene expression analyses. Growth of S. luteus was inhibited by both Heterobasidion species, but based on statistical analysis, growth inhibition was due to the isolate rather than to the species. The expression analysis on genes related to cell wall hydrolytic enzymes and hydrophobins, putatively involved in the fungus-fungus interaction, allowed to identify significantly up- and down-regulated genes both in the symbiont and in the pathogens. Based on the transcript analysis, it was not possible to distinguish the impact of the two pathogenic species on the ECM fungus. The only exception was a S. luteus gene coding for a putative chitinase (SlGH18_8356) that was found to be differentially regulated during interaction with H. irregulare compared to H. annosum.

Gnomoniopsis castanea is an emerging fungal pathogen causing nut rot of sweet chestnut, Castanea sativa. This study was aimed at testing and modelling the effects of climate on disease incidence. Up to 120 ripe nuts were collected in 2011 from trees in each of 12 sites located in the northwest of Italy. The incidence of G. castanea in each site was expressed as the number of infected nuts out of the total number of nuts sampled (%), determined by combining the results of morphological identification of isolates obtained from nuts, and their typing through a newly developed taxon-specific molecular assay. Disease incidence ranged from 20 to 93%, depending on site. Geostatistical analyses revealed that, despite the clustering of sites (P 0·05). This finding suggests that the disease is influenced by site-dependent factors whose scale (c. 7·5–15·6 km) is consistent with the climate variability throughout the sampling region. Multivariate analyses on maximum, mean and minimum temperatures and on rainfall showed that warmer temperatures were associated with higher levels of disease incidence. The temperatures of months before nut harvesting were selected as predictors for partial least squares regression (PLSR) models (GnoMods) of G. castanea incidence. External validation on data collected either on sites or in years not used for model fitting showed the good predictive abilities of the GnoMods (Spearman's ρobs/pred > 0·72, P

The North American fungal pathogen Heterobasidion irregulare is currently distributed in pine and oak stands along 103 km of coastline west of Rome, Italy. This paper reviews and expands the knowledge on impacts, pathways of introduction and invasion, factors driving the invasion and on the dispersal abilities of this pathogen in Italy. Further, an integrated disease management programto minimize the spread of the fungus in Europe is suggested, based both on the published literature and on new findings reported here. Observational and genetic evidence support a single introduction through infected wood during World War II, and a subsequent invasion through spore dispersal. Experimental evidence suggests transmission potential of the pathogen rather than hypersusceptibility of native hosts is the major determinant of invasion. The current range of H. irregulare is too vast to suggest eradication, however, we recommend minimizing the risk of spread of H. irregulare outside the zone of infestation while reducing the magnitude of infestations within its current range. We provide evidence suggesting the mostcost-effectivemanagement approach hinges on preventing the saprobic establishment of the fungus in stumps in a 'buffer' area surrounding the current zone of infestation. © 2014 Institute of Chartered Foresters. All rights reserved.

The North American fungal pathogen Heterobasidion irregulare is invading and threatening pine stands in Italy and is freely hybridizing with the native species Heterobasidion annosum. Susceptibility of native hosts has been excluded as a factor driving H. irregulare invasion. Here we tested whether H. irregulare and H. annosum differ in their ability to saprobically colonize pine wood, and whether saprobic growth is correlated to fruiting bodies production. When inoculated in pine logs, H. irregulare genotypes colonized a volume of wood significantly larger than H. annosum genotypes. Heterobasidion irregulare significantly exceeded H. annosum in all parameters used as metrics of fruiting body production, including number and size of fruiting bodies, and pores perimeter index (PPI), an index summarizing the amount of surface available for spore production. Number of fruiting bodies and PPI were significantly correlated with volume of wood colonized by Heterobasidion genotypes. Results may explain why H. irregulare has been reported to sporulate more abundantly than H. annosum and provide explanations for its high transmission potential in Italy. This knowledge implies that approaches to control the spread of H. irregulare should be aimed at limiting saprobic establishment of the fungus rather than focusing on identifying more tolerant tree species.

Habitat preference of an invasive fungal tree pathogen is here compared with that of a sympatric and native closely related congener to test the hypothesis that the invasive ability of the exotic organism may be linked to its capacity to colonize substrates unavailable to the indigenous relative. We compared the distribution of infectious airspora of the North American Heterobasidion irregulare introduced into Italy with that of the native H. annosum, both regarded to be able to establish only in the presence of conifers. Geostatistical and statistical analyses were employed to test for association between both species and five Mediterranean vegetation types. Results show that H. annosum is positively associated with pines and negatively associated with deciduous oaks. The probability of finding its spores decreases to almost 0 at distances over 500 m from pines, and this species is virtually absent in pure oak forests. Spores of H. irregulare are present irrespective of vegetation type, and this species can be found not only where pines are present, but also in pure oak forests. This knowledge implies that spread of H. irregulare is not limited by the fragmented distribution of pine woodlands in central Italy and is essential to both predict and hinder its progress in Europe.