Diplodia sapinea as a contributing factor in the crown dieback of Scots pine (Pinus sylvestris) after a severe drought

The frequency and impact of drought on global ecosystems have increased within the last century, while drought has affected tree health in many regions. Diplodia sapinea is a widespread, opportunistic pathogen infecting most conifers, causing Diplodia tip blight, thriving on hosts impacted by stress such as drought, heat, or mechanical wounding. In summer of 2018, a large-scale drought was recorded all over Europe. In late summer, pine trees all over Gotland showed crown dieback, where necrotic twigs and needles were found, especially in the upper part of the crowns. Symptoms were consistent with a potential outbreak of D. sapinea. Effects of the combination of drought and Diplodia tip blight on mortality or recovery of Scots pine in Nordic conditions are unknown. This study confirmed the presence and potential contribution of D. sapinea in the observed damages of Scots pine. Shoot blight and drought led to crown defoliation which was observed one year post-drought, while trees showed a clear recovery of newly grown shoots within the second year. Severely affected pines (>70% of the upper third of the crown with shoot blight) showed increased mortality. Recovery of the surviving trees was independent of previous dieback levels. Diplodia sapinea was most abundant in twigs with shoot blight of the symptomatic trees compared to healthy-looking twigs from the same trees and asymptomatic trees in affected and healthy pine stands. Sampling on affected and healthy sites showed possible endophytic infections with low abundance within healthy-looking twigs. Spore deposition of D. sapinea was monitored on healthy and affected sites for two consecutive years after crown damages occurred to confirm the presence of the opportunistic pathogen in the affected region. Spore deposition was observed during all seasons and correlated with high precipitation during sampling. Our observations provide insights into the emergence of Diplodia tip blight in the Northern countries and underline the potential impact of D. sapinea on tree health in the course of a changing climate

Deciphering common and specific transcriptional immune responses in pea towards the oomycete pathogens Aphanomyces euteiches and Phytophthora pisi

Primers used in this study for RT-qPCR analysis. (DOCX 20 kb

Transcriptomic analysis identifies candidate genes for Aphanomyces root rot disease resistance in pea

BackgroundAphanomyces euteiches is a soil-borne oomycete that causes root rot in pea and other legume species. Symptoms of Aphanomyces root rot (ARR) include root discoloration and wilting, leading to significant yield losses in pea production. Resistance to ARR is known to be polygenic but the roles of single genes in the pea immune response are still poorly understood. This study uses transcriptomics to elucidate the immune response of two pea genotypes varying in their levels of resistance to A. euteiches.ResultsIn this study, we inoculated roots of the pea (P. sativum L.) genotypes 'Linnea' (susceptible) and 'PI180693' (resistant) with two different A. euteiches strains varying in levels of virulence. The roots were harvested at 6 h post-inoculation (hpi), 20 hpi and 48 hpi, followed by differential gene expression analysis. Our results showed a time- and genotype-dependent immune response towards A. euteiches infection, involving several WRKY and MYB-like transcription factors, along with genes associated with jasmonic acid (JA) and abscisic acid (ABA) signaling. By cross-referencing with genes segregating with partial resistance to ARR, we identified 39 candidate disease resistance genes at the later stage of infection. Among the genes solely upregulated in the resistant genotype 'PI180693', Psat7g091800.1 was polymorphic between the pea genotypes and encoded a Leucine-rich repeat receptor-like kinase reminiscent of the Arabidopsis thaliana FLAGELLIN-SENSITIVE 2 receptor.ConclusionsThis study provides new insights into the gene expression dynamics controlling the immune response of resistant and susceptible pea genotypes to A. euteiches infection. We present a set of 39 candidate disease resistance genes for ARR in pea, including the putative immune receptor Psat7g091800.1, for future functional validation

Fungal Communities Vectored by Ips sexdentatus in Declining Pinus sylvestris in Ukraine: Focus on Occurrence and Pathogenicity of Ophiostomatoid Species

Drought-induced stress and attacks by bark beetle Ips sexdentatus currently result in a massive dieback of Pinus sylvestris in eastern Ukraine. Limited and fragmented knowledge is available on fungi vectored by the beetle and their roles in tree dieback. The aim was to investigate the fungal community vectored by I. sexdentatus and to test the pathogenicity of potentially aggressive species to P. sylvestris. Analysis of the fungal community was accomplished by combining different methods using insect, plant, and fungal material. The material consisted of 576 beetles and 96 infested wood samples collected from six sample plots within a 300 km radius in eastern Ukraine and subjected to fungal isolations and (beetles only) direct sequencing of ITS rDNA. Pathogenicity tests were undertaken by artificially inoculating three-to-four-year-old pine saplings with fungi. For the vector test, pine logs were exposed to pre-inoculated beetles. In all, 56 fungal taxa were detected, 8 exclusively by isolation, and 13 exclusively by direct sequencing. Those included nine ophiostomatoids, five of which are newly reported as I. sexdentatus associates. Two ophiostomatoid fungi, which exhibited the highest pathogenicity, causing 100% dieback and mortality, represented genera Graphium and Leptographium. Exposure of logs to beetles resulted in ophiostomatoid infections. In conclusion, the study revealed numerous I. sexdentatus-vectored fungi, several of which include aggressive tree pathogens

Evaluation of pea genotype PI180693 partial resistance towards aphanomyces root rot in commercial pea breeding

The cultivation of vining pea (Pisum sativum) faces a major constraint with root rot diseases, caused by a complex of soil-borne pathogens including the oomycetes Aphanomyces euteiches and Phytophtora pisi. Disease resistant commercial varieties are lacking but the landrace PI180693 is used as a source of partial resistance in ongoing pea breeding programs. In this study, the level of resistance and their interaction with A. euteiches virulence levels of six new back-crossed pea breeding lines, deriving from the cross between the susceptible commercial cultivar Linnea and PI180693, were evaluated for their resistance towards aphanomyces root rot in growth chamber and green house tests. Resistance towards mixed infections by A. euteiches and P. pisi and commercial production traits were evaluated in field trials. In growth chamber trials, pathogen virulence levels had a significant effect on plant resistance, as resistance was more consistent against A. euteiches strains exhibiting high or intermediate virulence compared with lowly virulent strains. In fact, line Z1701-1 showed to be significantly more resistant than both parents when inoculated with a lowly virulent strain. In two separate field trials in 2020, all six breeding lines performed equally well as the resistant parent PI180693 at sites only containing A. euteiches, as there were no differences in disease index. In mixed infections, PI180693 exhibited significantly lower disease index scores than Linnea. However, breeding lines displayed higher disease index scores compared with PI180693, indicating higher susceptibility towards P. pisi. Data on seedling emergence from the same field trials suggested that PI180693 was particularly sensitive towards seed decay/damping off disease caused by P. pisi. Furthermore, the breeding lines performed equally well as Linnea in traits important for green pea production, again emphasizing the commercial potential. In summary, we show that the resistance from PI180693 interacts with virulence levels of the pathogen A. euteiches and is less effective towards root rot caused by P. pisi. Our results show the potential use of combining PI180693 partial resistance against aphanomyces root rot with commercially favorable breeding traits in commercial breeding programs

Identifying Fraxinus excelsior tolerant to ash dieback: Visual field monitoring versus a molecular marker

Ash dieback (ADB) caused by the pathogen Hymenoscyphus fraxineus is the cause of massive mortality of Fraxinus spp. in Europe. The aim of this work was to check for the presence of the molecular marker for ADB tolerance in mapped healthy-looking F. excelsior trees, and to compare its occurrence in trees exhibiting severe ADB symptoms. Monitoring of 135 healthy-looking F. excelsior on the island of Gotland, Sweden, showed that after 3-4 years 99.3% of these trees had 0%-10% crown damage, thus remaining in a similar health condition as when first mapped. After 5-6 years, 94.7% of these trees had 0%-10% crown damage. Molecular analysis of leaf tissues from 40 of those showed the presence of the molecular marker in 34 (85.0%) trees, while it was absent in 6 (15.0%) trees. Analysis of leaf tissues from 40 severely ADB-diseased trees showed the presence of the molecular marker in 17 (42.5%) trees, but its absence in 23 (57.5%) trees (p < .0001). The results demonstrated that monitoring of healthy-looking F. excelsior is a simple and straightforward approach for the selection of presumably ADB-tolerant ash for future breeding. The cDNA-based molecular marker revealed moderate capacity on its own to discriminate between presumably ADB-tolerant and susceptible F. excelsior genotypes

Host genotype interacts with aerial spore communities and influences the needle mycobiome of Norway spruce

The factors shaping the composition of the tree mycobiome are still under investigation. We tested the effects of host genotype, site, host phenotypic traits, and air fungal spore communities on the assembly of the fungi inhabiting Norway spruce needles. We used Norway spruce clones and spore traps within the collection sites and characterized both needle and air mycobiome communities by high-throughput sequencing of the ITS2 region. The composition of the needle mycobiome differed between Norway spruce clones, and clones with high genetic similarity had a more similar mycobiome. The needle mycobiome also varied across sites and was associated with the composition of the local air mycobiome and climate. Phenotypic traits such as diameter at breast height or crown health influenced the needle mycobiome to a lesser extent than host genotype and air mycobiome. Altogether, our results suggest that the needle mycobiome is mainly driven by the host genotype in combination with the composition of the local air spore communities. Our work highlights the role of host intraspecific variation in shaping the mycobiome of trees and provides new insights on the ecological processes structuring fungal communities inhabiting woody plants

Host genotype interacts with aerial spore communities and influences the needle mycobiome of Norway spruce

The factors shaping the composition of the tree mycobiome are still under investigation. We tested the effects of host genotype, site, host phenotypic traits, and air fungal spore communities on the assembly of the fungi inhabiting Norway spruce needles. We used Norway spruce clones and spore traps within the collection sites and characterized both needle and air mycobiome communities by high-throughput sequencing of the ITS2 region. The composition of the needle mycobiome differed between Norway spruce clones, and clones with high genetic similarity had a more similar mycobiome. The needle mycobiome also varied across sites and was associated with the composition of the local air mycobiome and climate. Phenotypic traits such as diameter at breast height or crown health influenced the needle mycobiome to a lesser extent than host genotype and air mycobiome. Altogether, our results suggest that the needle mycobiome is mainly driven by the host genotype in combination with the composition of the local air spore communities. Our work highlights the role of host intraspecific variation in shaping the mycobiome of trees and provides new insights on the ecological processes structuring fungal communities inhabiting woody plants.This research was supported by the Swedish research council for Environment, Agricultural Sciences and Spatial Planning, FORMAS, project 2016-00798. M.E. and H.D.C were also supported by Formas project 2017-00402. J.O. was partially supported by the 'Ramon y Cajal' fellowship RYC-2015-17459. The authors would like to thank the owners of the seed orchards, Svenska skogsplantor AB and Sodra skogsagarna AB, for allowing us to sample the trees and assisting with the air mycobiome sampling. The authors also thank Antonio Rizzi, Rena Gadjieva, Maria Jonsson, and Katarina Ihrmark for their assistance with the laboratory and field work. The authors would like to acknowledge the support of the National Genomics Infrastructure (NGI)/Uppsala, Genome Center and UPPMAX for assisting us in massive parallel sequencing and computational infrastructure. Work performed at NGI/Uppsala Genome Center was funded by RFI/VR and Science for Life Laboratory, Sweden

Genetic diversity of the pea root pathogen Aphanomyces euteiches in Europe

The oomycete pathogen Aphanomyces euteiches causes root rot in various legume species. In this study we focused on A. euteiches causing root rot in pea (Pisum sativum), thereby being responsible for severe yield losses in pea production. We aimed to understand the genetic diversity of A. euteiches in Europe, covering a north-to-south gradient spanning from Sweden, Norway and Finland to the UK, France and Italy. A collection of 85 European A. euteiches strains was obtained, all isolated from infected pea roots from commercial vining pea cultivation fields. The strains were genotyped using 22 simple-sequence repeat markers. Multilocus genotypes were compiled and the genetic diversity between individual strains and population structure between countries was analysed. The population comprising strains from Italy was genetically different and did not share ancestry with any other population. Also, strains originating from Finland and the eastern parts of Sweden were found to be significantly different from the other populations, while strains from the rest of Europe were more closely related. A subset of 10 A. euteiches strains from four countries was further phenotyped on two susceptible pea genotypes, as well as on one genotype with partial resistance towards A. euteiches. All strains were pathogenic on all pea genotypes, but with varying levels of disease severity. No correlation between the genetic relatedness of strains and virulence levels was found. In summary, our study identified three genetically distinct groups of A. euteiches in Europe along a north-to-south gradient, indicating local pathogen differentiation