Possible Biological Control of Ash Dieback Using the Mycoparasite Hymenoscyphus Fraxineus Mitovirus 2.

Invasive fungal diseases represent a major threat to forest ecosystems worldwide. As the application of fungicides is often unfeasible and not a sustainable solution, only a few other control options are available, including biological control. In this context, the use of parasitic mycoviruses as biocontrol agents of fungal pathogens has recently gained particular attention. Since the 1990s, the Asian fungus Hymenoscyphus fraxineus has been causing lethal ash dieback across Europe. In the present study, we investigated the biocontrol potential of the mitovirus Hymenoscyphus fraxineus mitovirus 2 (HfMV2) previously identified in Japanese populations of the pathogen. HfMV2 could be successfully introduced via co-culturing into 16 of 105 HfMV2-free isolates. Infection with HfMV2 had contrasting effects on fungal growth in vitro, from cryptic to detrimental or beneficial. Virus-infected H. fraxineus isolates whose growth was reduced by HfMV2 showed overall a lower virulence on ash (Fraxinus excelsior) saplings as compared with their isogenic HfMV2-free lines. The results suggest that mycoviruses exist in the native populations of H. fraxineus in Asia that have the potential for biological control of ash dieback in Europe. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Distribution, causal agents, and infection dynamic of emerging ink disease of sweet chestnut in Southern Switzerland.

Emerging diseases caused by both native and exotic pathogens represent a main threat to forest ecosystems worldwide. The two invasive soilborne pathogens Phytophthora cinnamomi and Phytophthora × cambivora are the causal agents of ink disease, which has been threatening Castanea sativa in Europe for several centuries and seems to be re-emerging in recent years. Here, we investigated the distribution, causal agents, and infection dynamics of ink disease in southern Switzerland. A total of 25 outbreaks were identified, 19 with only P. cinnamomi, 5 with only P. × cambivora, and 1 with both species. Dendrochronological analyses showed that the disease emerged in the last 20-30 years. Infected trees either died rapidly within 5-15 years post-infection or showed a prolonged state of general decline until death. Based on a generalized linear model, the local risk of occurrence of ink disease was increased by an S-SE aspect of the chestnut stand, the presence of a pure chestnut stand, management activities, the proximity of roads and buildings, and increasing annual mean temperature and precipitation. The genetic structure of the local P. cinnamomi population suggests independent introductions and local spread of the pathogen.

Global invasion history of the emerging plant pathogen Phytophthora multivora.

BACKGROUND: global trade in living plants and plant material has significantly increased the geographic distribution of many plant pathogens. As a consequence, several pathogens have been first found and described in their introduced range where they may cause severe damage on naïve host species. Knowing the center of origin and the pathways of spread of a pathogen is of importance for several reasons, including identifying natural enemies and reducing further spread. Several Phytophthora species are well-known invasive pathogens of natural ecosystems, including Phytophthora multivora. Following the description of P. multivora from dying native vegetation in Australia in 2009, the species was subsequently found to be common in South Africa where it does not cause any remarkable disease. There are now reports of P. multivora from many other countries worldwide, but not as a commonly encountered species in natural environments. RESULTS: a global collection of 335 isolates from North America, Europe, Africa, Australia, the Canary Islands, and New Zealand was used to unravel the worldwide invasion history of P. multivora, using 10 microsatellite markers for all isolates and sequence data from five loci from 94 representative isolates. Our population genetic analysis revealed an extremely low heterozygosity, significant non-random association of loci and substantial genotypic diversity suggesting the spread of P. multivora readily by both asexual and sexual propagules. The P. multivora populations in South Africa, Australia, and New Zealand show the most complex genetic structure, are well established and evolutionary older than those in Europe, North America and the Canary Islands. CONCLUSIONS: according to the conducted analyses, the world invasion of P. multivora most likely commenced from South Africa, which can be considered the center of origin of the species. The pathogen was then introduced to Australia, which acted as bridgehead population for Europe and North America. Our study highlights a complex global invasion pattern of P. multivora, including both direct introductions from the native population and secondary spread/introductions from bridgehead populations.

Temporal changes in pathogen diversity in a perennial plant-pathogen-hyperparasite system.

Hyperparasites can affect the evolution of pathosystems by influencing the stability of both pathogen and host populations. However, how pathogens of perennial hosts evolve in the presence of a hyperparasite has rarely been studied. Here, we investigated temporal changes in genetic diversity of the invasive chestnut blight pathogen Cryphonectria parasitica in the presence of its parasitic mycovirus Cryphonectria hypovirus 1 (CHV1). The virus reduces fungal virulence and represents an effective natural biocontrol agent against chestnut blight in Europe. We analysed genome-wide diversity and CHV1 prevalence in C. parasitica populations in southern Switzerland that were sampled twice at an interval of about 30 years. Overall, we found that both pathogen population structure and CHV1 prevalence were retained over time. The results suggest that recent bottlenecks have influenced the structure of C. parasitica populations in southern Switzerland. Strong balancing selection signals were found at a single vegetative incompatibility (vic) locus, consistent with negative frequency-dependent selection imposed by the vegetative incompatibility system. High levels of mating among related individuals (i.e., inbreeding) and genetic drift are probably at the origin of imbalanced allele ratios at vic loci and subsequently low vc type diversity. Virus infection rates were stable at ~30% over the study period and we found no significant impact of the virus on fungal population diversity. Consequently, the efficacy of CHV1-mediated biocontrol was probably retained.

Temporal and Spatial Genetic Population Structure of Cryphonectria parasitica and Its Associated Hypovirus Across an Invasive Range of Chestnut Blight in Europe.

Chestnut blight has spread throughout Europe since the introduction of its causal agent, Cryphonectria parasitica, >70 years ago. In our study, we analyzed the diversity of vegetative compatibility (vc) and microsatellite genotypes of C. parasitica, as well as sequence diversity of Cryphonectria hypovirus 1 (CHV1) in six populations from Switzerland, Croatia, and North Macedonia. Resampling of local populations that were already investigated more than a decade ago allowed us to analyze the spatial and temporal population structure across an invasive range of the pathogen in Europe. Regardless of which genetic marker was used, the >60-year-old Swiss and Croatian populations had high population diversity, whereas more recent North Macedonian populations were mostly clonal. These diversity differences between the investigated populations remained stable over time. A high diversity of CHV1 was observed in all three countries, with North Macedonian strains forming a separate cluster from strains obtained in other countries. No correlation between vc diversity and CHV1 prevalence was observed, suggesting a well-established and maintained natural hypovirulence in all countries, further corroborated by an observed increase in genetic diversity of Croatian C. parasitica populations over time, without collapse of CHV1 prevalence.

Patterns of Genetic Diversification in the Invasive Hybrid Plant Pathogen Phytophthora × alni and Its Parental Species P. uniformis.

In pathogenic fungi and oomycetes, interspecific hybridization may lead to the formation of new species having a greater impact on natural ecosystems than the parental species. From the early 1990s, a severe alder (Alnus spp.) decline due to an unknown Phytophthora species was observed in several European countries. Genetic analyses revealed that the disease was caused by the triploid hybrid P. × alni, which originated in Europe from the hybridization of P. uniformis and P. × multiformis. Here, we investigated the population structure of P. × alni (158 isolates) and P. uniformis (85 isolates) in several European countries using microsatellite markers. Our analyses confirmed the genetic structure previously observed in other European populations, with P. uniformis populations consisting of at most two multilocus genotypes (MLGs) and P. × alni populations dominated by MLG Pxa-1. The genetic structure of P. × alni populations in the Czech Republic, Hungary and Sweden seemed to reflect the physical isolation of river systems. Most rare P. × alni MLGs showed a loss of heterozygosity (LOH) at one or a few microsatellite loci compared with other MLGs. This LOH may allow a stabilization within the P. × alni genome or a rapid adaptation to stress situations. Alternatively, alleles may be lost because of random genetic drift in small, isolated populations, with no effect on fitness of P. × alni. Additional studies would be necessary to confirm these patterns of population diversification and to better understand the factors driving it.

Deciphering the worldwide invasion of the Asian long-horned beetle: A recurrent invasion process from the native area together with a bridgehead effect.

Retracing introduction routes is crucial for understanding the evolutionary processes involved in an invasion, as well as for highlighting the invasion history of a species at the global scale. The Asian long-horned beetle (ALB) Anoplophora glabripennis is a xylophagous pest native to Asia and invasive in North America and Europe. It is responsible for severe losses of urban trees, in both its native and invaded ranges. Based on historical and genetic data, several hypotheses have been formulated concerning its invasion history, including the possibility of multiple introductions from the native zone and secondary dispersal within the invaded areas, but none have been formally tested. In this study, we characterized the genetic structure of ALB in both its native and invaded ranges using microsatellites. In order to test different invasion scenarios, we used an approximate Bayesian "random forest" algorithm together with traditional population genetics approaches. The strong population differentiation observed in the native area was not geographically structured, suggesting complex migration events that were probably human-mediated. Both native and invasive populations had low genetic diversity, but this characteristic did not prevent the success of the ALB invasions. Our results highlight the complexity of invasion pathways for insect pests. Specifically, our findings indicate that invasive species might be repeatedly introduced from their native range, and they emphasize the importance of multiple, human-mediated introductions in successful invasions. Finally, our results demonstrate that invasive species can spread across continents following a bridgehead path, in which an invasive population may have acted as a source for another invasion.

Are traded forest tree seeds a potential source of nonnative pests?

The international seed trade is considered relatively safe from a phytosanitary point of view and is therefore less regulated than trade in other plants for planting. However, the pests carried by traded seeds are not well known. We assessed insects and fungi in 58 traded seed lots of 11 gymnosperm and angiosperm tree species from North America, Europe, and Asia. Insects were detected by X-raying and molecular methods. The fungal community was characterized using high-throughput sequencing (HTS) and by growing fungi on non-selective agar. About 30% of the seed lots contained insect larvae. Gymnosperms contained mostly hymenopteran (Megastigmus spp.) and dipteran (Cecidomyiidae) larvae, while angiosperms contained lepidopteran (Cydia latiferreana) and coleopteran (Curculio spp.) larvae. HTS indicated the presence of fungi in all seed lots and fungi grew on non-selective agar from 96% of the seed lots. Fungal abundance and diversity were much higher than insect diversity and abundance, especially in angiosperm seeds. Almost 50% of all fungal exact sequence variants (ESVs) found in angiosperms were potential pathogens, in comparison with around 30% of potentially pathogenic ESVs found in gymnosperms. The results of this study indicate that seeds may pose a greater risk of pest introduction than previously believed or accounted for. A rapid risk assessment suggests that only a small number of species identified in this study is of phytosanitary concern. However, more research is needed to enable better risk assessment, especially to increase knowledge about the potential for transmission of fungi to seedlings and the host range and impact of identified species.

Assessing the Phytosanitary Risk Posed by an Intraspecific Invasion of Cryphonectria parasitica in Europe.

Intraspecific cryptic invasions may occur when new strains of an invasive species are introduced into an area where this species had already been introduced previously. In plant pathogens, such invasions are not well studied, even if, potentially, they can have severe consequences. Here, we investigated the effects of a potential intraspecific invasion in Europe of Cryphonectria parasitica, the causal agent of chestnut blight. Specifically, we tested the hypotheses that (i) non-European strains are more virulent on Castanea sativa than those already present in Europe because they have never encountered this new host, and (ii) the variation in virulence among strains is higher within native than within introduced populations. In a greenhouse, 2-year-old C. sativa seedlings were inoculated with Cryphonectria parasitica strains from South Korea, the United States, and Switzerland, and lesion development and seedling mortality were recorded weekly. Additionally, growth and sporulation of the strains were measured in vitro on agar medium at 15 and 24°C. Although lesion growth was similar for all strains, seedlings inoculated with strains from South Korea and Switzerland died faster than seedlings inoculated with strains from the United States. Moreover, in vitro strains from South Korea grew faster and produced more spores at both temperatures than the strains from the other two countries. In conclusion, our results did not support the two hypotheses. All strains, regardless of their origin, were found to be highly virulent on the inoculated chestnut seedlings. Nevertheless, current phytosanitary measures to avoid the introduction of new genotypes of C. parasitica into Europe should be further implemented.

Role of Fresh Dead Wood in the Epidemiology and the Biological Control of the Chestnut Blight Fungus.

The invasive fungus Cryphonectria parasitica, the causal agent of chestnut blight, is able to survive and sporulate on the bark of fresh dead Castanea sativa wood for at least 2 years. Here, we experimentally investigated the role of fresh dead wood in the epidemiology of chestnut blight, specifically in the spread of the hyperparasitic virus Cryphonectria hypovirus 1, which acts as biocontrol agent of C. parasitica. A total of 152 artificially initiated, virulent bark cankers in four chestnut stands were treated with virus-infected asexual spores originating either from sporulating dead wood or from a spore suspension. Molecular markers for both the virus and the fungal carrier were used to examine the spread of the applied biocontrol virus. Fourteen months after treatment, 42 to 76% of the conidial spray-treated cankers and 50 to 60% of the cankers exposed to a sporulating dead stem had been virus infected by the applied hypovirulent conidia in all four study sites. Virus infection reduced canker expansion and promoted canker healing (callusing). Thus, fresh chestnut dead wood may play an important role in supporting the successful spread of natural hypovirulence in chestnut forests. Further, combined with the application of virus-infected conidial suspensions, it may help promote the establishment of artificially released hypoviruses in chestnut stands to control chestnut blight.

Insights into the phylogeny of Northern Hemisphere Armillaria: Neighbor-net and Bayesian analyses of translation elongation factor 1-α gene sequences.

Armillaria possesses several intriguing characteristics that have inspired wide interest in understanding phylogenetic relationships within and among species of this genus. Nuclear ribosomal DNA sequence-based analyses of Armillaria provide only limited information for phylogenetic studies among widely divergent taxa. More recent studies have shown that translation elongation factor 1-α (tef1) sequences are highly informative for phylogenetic analysis of Armillaria species within diverse global regions. This study used Neighbor-net and coalescence-based Bayesian analyses to examine phylogenetic relationships of newly determined and existing tef1 sequences derived from diverse Armillaria species from across the Northern Hemisphere, with Southern Hemisphere Armillaria species included for reference. Based on the Bayesian analysis of tef1 sequences, Armillaria species from the Northern Hemisphere are generally contained within the following four superclades, which are named according to the specific epithet of the most frequently cited species within the superclade: (i) Socialis/Tabescens (exannulate) superclade including Eurasian A. ectypa, North American A. socialis (A. tabescens), and Eurasian A. socialis (A. tabescens) clades; (ii) Mellea superclade including undescribed annulate North American Armillaria sp. (Mexico) and four separate clades of A. mellea (Europe and Iran, eastern Asia, and two groups from North America); (iii) Gallica superclade including Armillaria Nag E (Japan), multiple clades of A. gallica (Asia and Europe), A. calvescens (eastern North America), A. cepistipes (North America), A. altimontana (western USA), A. nabsnona (North America and Japan), and at least two A. gallica clades (North America); and (iv) Solidipes/Ostoyae superclade including two A. solidipes/ostoyae clades (North America), A. gemina (eastern USA), A. solidipes/ostoyae (Eurasia), A. cepistipes (Europe and Japan), A. sinapina (North America and Japan), and A. borealis (Eurasia) clade 2. Of note is that A. borealis (Eurasia) clade 1 appears basal to the Solidipes/Ostoyae and Gallica superclades. The Neighbor-net analysis showed similar phylogenetic relationships. This study further demonstrates the utility of tef1 for global phylogenetic studies of Armillaria species and provides critical insights into multiple taxonomic issues that warrant further study.

Virulence and Stump Colonization Ability of Armillaria borealis on Norway Spruce Seedlings in Comparison to Sympatric Armillaria Species.

Although Armillaria borealis is one of the closest relatives of the aggressive root rot pathogen A. ostoyae, little is known about its ecology. In central and northern Europe, A. borealis often co-occurs with A. ostoyae or A. cepistipes, a weak pathogen, in conifer or mixed-forest stands. In this study, the virulence of 10 A. borealis, nine A. ostoyae, and five A. cepistipes isolates toward 2- and 4-year-old potted Norway spruce (Picea abies) seedlings was assessed. In addition, the ability of all isolates to colonize fresh stumps cut from 6-year-old Norway spruce seedlings was tested. All inoculations were done by insertion of Armillaria-colonized hazelnut stem segments into the soil substrate. On the 2-year-old seedlings, A. borealis and A. ostoyae showed, overall, a similar virulence 29 months after inoculation whereas, on the 4-year-old seedlings, A. ostoyae was more virulent. The third species in the experiment, A. cepistipes, caused almost no seedling mortality. Six months after cutting, the highest percentage of stumps was colonized by A. ostoyae (94.3%), followed by A. borealis (85.2%), and A. cepistipes (78.4%). Our inoculation experiments show that A. borealis has a clear pathogenic potential toward Norway spruce seedlings. However, compared with A. ostoyae, damage caused by A. borealis may decrease more rapidly with increasing tree age. Similar to other Armillaria species, A. borealis seems to be an efficient colonizer of fresh stumps.

Dynamics of Cryphonectria hypovirus infection in chestnut blight cankers.

Virulent strains of the chestnut blight fungus Cryphonectria parasitica cause lethal bark cankers on chestnut trees. Infection of C. parasitica with Cryphonectria hypovirus 1 in Europe biologically controls this disease, leading to nonlethal and inactive cankers. Unexpectedly, virus-free C. parasitica strains have been isolated from inactive cankers. In this study, we compared the virulence of virus-infected and virus-free C. parasitica strains isolated from either inactive or active cankers on chestnut seedlings and sprouts. In the seedling experiment, we assessed canker growth and seedling mortality. In the sprout experiment, we also assessed canker growth and made fungal reisolations to determine virus infection and immigration of foreign vegetative compatibility (vc) types over a period of 13 years in a coppice forest. Overall, the virulence of virus-free C. parasitica strains isolated from inactive versus active cankers did not differ. Significant differences were only attributed to virus infection. Virus infection and fungal strain composition in cankers changed over time. Foreign vc types immigrated into cankers and virus-free cankers became virus-infected within a few years. Most of the cankers were callused over time and became inactive. However, we observed that the virus did not always persist in these cankers. This study demonstrates that virus spread occurs effectively in European chestnut forests and that this biocontrol system is highly dynamic.

Development of new polymorphic microsatellite markers for three closely related plant-pathogenic Phytophthora species using 454-pyrosequencing and their potential applications.

Phytophthora spp. (oomycetes) are causal agents of devastating diseases on a high number of crops, ornamentals, and native plants worldwide. Neutral molecular markers are increasingly being used to investigate the genetic population structure and possible pathways of spread of different plant pathogens, including Phytophthora spp. In this study, polymorphic microsatellite markers were developed for three species of the former Phytophthora citricola species complex-namely, P. multivora, P. plurivora, and P. pini (P. citricola I)-using the 454-pyrosequencing technique. In total, 35 polymorphic microsatellite loci were found and further characterized: 11 for P. plurivora, 16 for P. multivora, and 8 for P. pini. Microsatellites with dinucleotide motifs repeated 6 to 10 times were the most common for all three species. On average, 65 alleles per species and 5.3 alleles per locus were detected. Most loci were characterized by a low observed heterozygosity, which might be due to the homothallic mating system of the three Phytophthora spp. targeted. Cross amplification of the newly developed markers was tested on 17 Phytophthora spp. belonging to five different internal transcribed spacer clades. Transferability success was generally low and decreased with increasing genetic distance from the species to the three target species. A set of four loci was selected to easily discriminate P. plurivora, P. multivora, and P. pini on the basis of presence or absence of a polymerase chain reaction amplicon on an agarose gel.

A new multilocus approach for a reliable DNA-based identification of Armillaria species.

In this paper we highlight and critically discuss limitations to molecular methods for identification of fungi via the example of the basidiomycete genus Armillaria. We analyzed a total of 144 sequences of three DNA regions commonly used for identifying fungi (ribosomal IGS-1 and ITS regions, translation elongation factor-1 alpha gene) from 48 specimens of six Armillaria species occurring in Europe (A. cepistipes, A. ostoyae, A. gallica, A. borealis, A. mellea, A. tabescens). Species were identified by comparing newly obtained sequences with those from the NCBI database, phylogenetic analyses and PCR-RFLP analyses of the three regions considered. When analyzed separately, no single gene region could unambiguously identify all six Armillaria species because of low interspecific and high intrasequence variability. We therefore developed a multilocus approach, which involves the stepwise use of the three regions. Following this scheme, all six species could be clearly discriminated. Our study suggests that, to improve the reliability of DNA-based techniques for species identification, multiple genes or intergenic regions should be analyzed.

Climate, host and geography shape insect and fungal communities of trees.

Non-native pests, climate change, and their interactions are likely to alter relationships between trees and tree-associated organisms with consequences for forest health. To understand and predict such changes, factors structuring tree-associated communities need to be determined. Here, we analysed the data consisting of records of insects and fungi collected from dormant twigs from 155 tree species at 51 botanical gardens or arboreta in 32 countries. Generalized dissimilarity models revealed similar relative importance of studied climatic, host-related and geographic factors on differences in tree-associated communities. Mean annual temperature, phylogenetic distance between hosts and geographic distance between locations were the major drivers of dissimilarities. The increasing importance of high temperatures on differences in studied communities indicate that climate change could affect tree-associated organisms directly and indirectly through host range shifts. Insect and fungal communities were more similar between closely related vs. distant hosts suggesting that host range shifts may facilitate the emergence of new pests. Moreover, dissimilarities among tree-associated communities increased with geographic distance indicating that human-mediated transport may serve as a pathway of the introductions of new pests. The results of this study highlight the need to limit the establishment of tree pests and increase the resilience of forest ecosystems to changes in climate.

Microsatellite Genotyping in the Chestnut Blight Fungus Cryphonectria parasitica.

This chapter describes the use of polymorphic microsatellite (simple sequence repeats, SSR) markers for genotyping isolates of Cryphonectria parasitica, the causal agent of chestnut blight. The SSR presented are particularly useful to characterize the genetic population structure of this invasive fungal pathogen, including invasion history (e.g., possible source population, introduced genotypes) and reproduction mode (sexual vs. asexual). Microsatellite markers can also be used to track fungal strains in laboratory and field experiments.

Novel RNA viruses from the native range of Hymenoscyphus fraxineus, the causal fungal agent of ash dieback.

The native Japanese population of the fungus Hymenoscyphus fraxineus, the causal agent of ash dieback in Europe, was screened for viruses using a high-throughput sequencing method. Five RNA viruses were detected in 116 fungal isolates sequenced via Illumina RNA-seq platform, with an overall virus prevalence of 11.2%. The viruses were completely sequenced by RNA ligase mediated rapid amplification of cDNA ends (RLM-RACE) followed by Sanger sequencing. The sequences appear to represent new species from three established families (Mito-, Endorna- and Partitiviridae), one recognized genus (Botybirnavirus) and a negative-sense single-stranded RNA virus in the order Bunyavirales from the proposed family "Mybuviridae". The highest prevalence was found for the mitovirus (7.8%), that had two genomic forms (linear and circular), while the other viruses were detected each in one isolate. Co-infection of a mitovirus and an endornavirus was also observed in one of the infected isolates. Here we describe the molecular characterization of the identified viruses. This study expands the diversity of viruses in H. fraxineus and provides the basis for investigating the virus-mediated control of ash dieback in Europe.

Worldwide diversity of endophytic fungi and insects associated with dormant tree twigs.

International trade in plants and climate change are two of the main factors causing damaging tree pests (i.e. fungi and insects) to spread into new areas. To mitigate these risks, a large-scale assessment of tree-associated fungi and insects is needed. We present records of endophytic fungi and insects in twigs of 17 angiosperm and gymnosperm genera, from 51 locations in 32 countries worldwide. Endophytic fungi were characterized by high-throughput sequencing of 352 samples from 145 tree species in 28 countries. Insects were reared from 227 samples of 109 tree species in 18 countries and sorted into taxonomic orders and feeding guilds. Herbivorous insects were grouped into morphospecies and were identified using molecular and morphological approaches. This dataset reveals the diversity of tree-associated taxa, as it contains 12,721 fungal Amplicon Sequence Variants and 208 herbivorous insect morphospecies, sampled across broad geographic and climatic gradients and for many tree species. This dataset will facilitate applied and fundamental studies on the distribution of fungal endophytes and insects in trees.

Emergence and diversification of a highly invasive chestnut pathogen lineage across southeastern Europe.

Invasive microbial species constitute a major threat to biodiversity, agricultural production and human health. Invasions are often dominated by one or a small number of genotypes, yet the underlying factors driving invasions are poorly understood. The chestnut blight fungus Cryphonectria parasitica first decimated the North American chestnut, and a more recent outbreak threatens European chestnut stands. To unravel the chestnut blight invasion of southeastern Europe, we sequenced 230 genomes of predominantly European strains. Genotypes outside of the invasion zone showed high levels of diversity with evidence for frequent and ongoing recombination. The invasive lineage emerged from the highly diverse European genotype pool rather than a secondary introduction from Asia or North America. The expansion across southeastern Europe was mostly clonal and is dominated by a single mating type, suggesting a fitness advantage of asexual reproduction. Our findings show how an intermediary, highly diverse bridgehead population gave rise to an invasive, largely clonally expanding pathogen.