Diplodia tip blight pathogen's virulence empowered through host switch.

Increased drought combined with emerging pathogens poses an increased threat to forest health. This is attributable to the unpredictable behaviour of forest pathosystems, which can favour fungal pathogens over the host under persistent drought stress conditions. Diplodia sapinea (≡ Sphaeropsis sapinea) is one of the most severe pathogens in Scots pine (Pinus sylvestris) causing Diplodia tip blight (conifer blight) under certain environmental conditions. Recently, the fungus has also been isolated from non-conifer hosts, indicating that it has a broader host range than previously known. In this study we compared the impact of different levels of water availability on necrosis length caused by D. sapinea strains isolated as endophytes (eight strains isolated from asymptomatic Scots pine) and pathogens (five strains isolated from symptomatic Scots pine) and five strains isolated from symptomatic non-pine hosts. For all strains the decreased water availability increased the necrosis length in Scots pine shoots. The isolates from non-pine hosts caused the most severe reactions under all water availabilities. The results of the study indicate the likelihood that effects of climatic changes such as drought will drive D. sapinea damage in Scots pine-dominated forests and increase mortality rates in affected trees. Further, the higher necrosis in the Scots pines caused by strains that had performed a host switch are concerning with regard to future scenarios thus increasing infection pressure on Scots pine from unknown sources.

Fungi Associated With Woody Tissues of European Beech and Their Impact on Tree Health.

Filamentous fungi associated with woody tissues of European Beech (Fagus sylvatica) and isolated from diseased trees and healthy trees were examined in relation to their impact on tree health. To this end, classical culture-based isolation methods, in planta inoculations and fungal identification using ITS-barcode and morphological characters were used. Stem endophytes of healthy beech saplings collected in German forests were isolated to determine endophyte communities in woody stem tissues. Pathogenicity tests were performed on living potted beech saplings using twelve selected fungal pathogens and wood inhabiting fungi (Hypocreales, Botryosphaeriales, and Xylariales) originating mainly from European beech with symptoms of the complex disease Vitality loss, or from bark necroses, or known to be common endophytes of beech. The impact of these ascomycetous fungi with respect to tree health was discussed. The potential influences of endophytic fungi of beech and of test conditions are discussed in relation to the success of inoculation. All tested fungal strains except for Neonectria ditissima were able to establish themselves post inoculation in the beech stems and caused necroses when there was sufficient water, but at different severities. Under the experimental conditions, Botryosphaeria corticola was shown to be the most virulent tested latent pathogen against F. sylvatica. In the context of climate change and global warming, the tested Botryosphaeriaceae are able to play a primary role in the disease progress of Vitality loss of Beech. The key role of Neonectria coccinea in causing bark necroses and the loss of vitality in beech was confirmed because the tested strain induced large lesions on the beech saplings.

Highly Clonal Structure and Abundance of One Haplotype Characterise the Diplodia sapinea Populations in Europe and Western Asia.

Diplodia sapinea is a cosmopolitan endophyte and opportunistic pathogen having occurred on several conifer species in Europe for at least 200 years. In Europe, disease outbreaks have increased on several Pinus spp. in the last few decades. In this study, the genetic structure of the European and western Asian D. sapinea population were investigated using 13 microsatellite markers. In total, 425 isolates from 15 countries were analysed. A high clonal fraction and low genetic distance between most subpopulations was found. One single haplotype dominates the European population, being represented by 45.3% of all isolates and found in nearly all investigated countries. Three genetically distinct subpopulations were found: Central/North European, Italian and Georgian. The recently detected subpopulations of D. sapinea in northern Europe (Estonia) share several haplotypes with the German subpopulation. The northern European subpopulations (Latvia, Estonia and Finland) show relatively high genetic diversity compared to those in central Europe suggesting either that the fungus has existed in the North in an asymptomatic/endophytic mode for a long time or that it has spread recently by multiple introductions. Considerable genetic diversity was found even among isolates of a single tree as 16 isolates from a single tree resulted in lower clonal fraction index than most subpopulations in Europe, which might reflect cryptic sexual proliferation. According to currently published allelic patterns, D. sapinea most likely originates from North America or from some unsampled population in Asia or central America. In order to enable the detection of endophytic or latent infections of planting stock by D. sapinea, new species-specific PCR primers (DiSapi-F and Diplo-R) were designed. During the search for Diplodia isolates across the world for species specific primer development, we identified D. africana in California, USA, and in the Canary Islands, which are the first records of this species in North America and in Spain.

The Diplodia Tip Blight Pathogen Sphaeropsis sapinea Is the Most Common Fungus in Scots Pines' Mycobiome, Irrespective of Health Status-A Case Study from Germany.

The opportunistic pathogen Sphaeropsis sapinea (≡Diplodia sapinea) is one of the most severe pathogens in Scots pine, causing the disease Diplodia tip blight on coniferous tree species. Disease symptoms become visible when trees are weakened by stress. Sphaeropsis sapinea has an endophytic mode in its lifecycle, making it difficult to detect before disease outbreaks. This study aims to record how S. sapinea accumulates in trees of different health status and, simultaneously, monitor seasonal and age-related fluctuations in the mycobiome. We compared the mycobiome of healthy and diseased Scots pines. Twigs were sampled in June and September 2018, and filamentous fungi were isolated. The mycobiome was analyzed by high-throughput sequencing (HTS) of the ITS2 region. A PERMANOVA analysis confirmed that the mycobiome community composition significantly differed between growth years (p < 0.001) and sampling time (p < 0.001) but not between healthy and diseased trees. Sphaeropsis sapinea was the most common endophyte isolated and the second most common in the HTS data. The fungus was highly abundant in symptomless (healthy) trees, presenting in its endophytic mode. Our results highlight the ability of S. sapinea to accumulate unnoticed as an endophyte in healthy trees before the disease breaks out, representing a sudden threat to Scots pines in the future, especially with increasing drought conditions experienced by pines.

Consequences of Sphaeropsis tip blight disease for the phytohormone profile and antioxidative metabolism of its pine host.

Phytopathogenic fungi infections induce plant defence responses that mediate changes in metabolic and signalling processes with severe consequences for plant growth and development. Sphaeropsis tip blight, induced by the endophytic fungus Sphaeropsis sapinea that spreads from stem tissues to the needles, is the most widespread disease of conifer forests causing dramatic economic losses. However, metabolic consequences of this disease on bark and wood tissues of its host are largely unexplored. Here, we show that diseased host pines experience tissue dehydration in both bark and wood. Increased cytokinin and declined indole-3-acetic acid levels were observed in both tissues and increased jasmonic acid and abscisic acid levels exclusively in the wood. Increased lignin contents at the expense of holo-cellulose with declined structural biomass of the wood reflect cell wall fortification by S. sapinea infection. These changes are consistent with H2 O2 accumulation in the wood, required for lignin polymerization. Accumulation of H2 O2 was associated with more oxidized redox states of glutathione and ascorbate pools. These findings indicate that S. sapinea affects both phytohormone signalling and the antioxidative defence system in stem tissues of its pine host during the infection process.

Genetic diversification of the chestnut blight fungus Cryphonectria parasitica and its associated hypovirus in Germany.

Chestnut blight in south-western Germany was first reported in 1992 and is since expanding in distribution. Here we investigated the invasion history of Cryphonectria parasitica and its associated hypovirus. For this, we characterized 284 isolates collected between 1992 and 2012 for hypovirulence, vegetative compatibility (vc), mating type, and microsatellite haplotype. A total of 27 haplotypes and 15 vc types were observed, although the C. parasitica population analyzed is currently dominated to 50 % by one haplotype and to 64 % by the vc type EU-2. Structure analysis indicated two divergent genetic pools. Over 66 % of the haplotypes belonged to a pool probably originating from northern Italy. Further diversification is expected due to ongoing sexual recombination, but also to new migration and additional introductions. Cryphonectria hypovirus 1 (CHV-1) was found in four of five C. parasitica populations from Baden-Württemberg. Genetic analysis of the 35 CHV-1 isolates obtained revealed that they all belong to the German subtype, although they have clearly diverged from the first German hypovirus isolated in 1992. Our study suggests that C. parasitica has been introduced into Germany several times from two different gene pools, whereas the hypovirus most probably has a single origin.