Phylogenetic Relationships, Speciation, and Origin of Armillaria in the Northern Hemisphere: A Lesson Based on rRNA and Elongation Factor 1-Alpha

Armillaria species have a global distribution and play various roles in the natural ecosystems, e.g., pathogens, decomposers, and mycorrhizal associates. However, their taxonomic boundaries, speciation processes, and origin are poorly understood. Here, we used a phylogenetic approach with 358 samplings from Europe, East Asia, and North America to delimit the species boundaries and to discern the evolutionary forces underpinning divergence and evolution. Three species delimitation methods indicated multiple unrecognized phylogenetic species, and biological species recognition did not reflect the natural evolutionary relationships within Armillaria; for instance, biological species of A. mellea and D. tabescens are divergent and cryptic species/lineages exist associated with their geographic distributions in Europe, North America, and East Asia. While the species-rich and divergent Gallica superclade might represent three phylogenetic species (PS I, PS II, and A. nabsnona) that undergo speciation. The PS II contained four lineages with cryptic diversity associated with the geographic distribution. The genus Armillaria likely originated from East Asia around 21.8 Mya in early Miocene when Boreotropical flora (56–33.9 Mya) and the Bering land bridge might have facilitated transcontinental dispersal of Armillaria species. The Gallica superclade arose at 9.1 Mya and the concurrent vicariance events of Bering Strait opening and the uplift of the northern Tibetan plateau might be important factors in driving the lineage divergence.

Global distribution and richness of Armillaria species inferred from public databases and amplicon sequencing datasets

Armillaria is a globally distributed fungal genus most notably recognized as economically important plant pathogens that are found predominantly in forest and agronomic systems. Armillaria has more recently received attention for ecologically diverse roles as woody plant decomposers and as mycorrhizal symbionts with specific plants. Previous phylogenetic analyses suggest that around 50 species are recognized globally. Despite this previous work, no studies have analyzed the global species richness and distribution of the genus using data derived from fungal community sequencing datasets. In order to assess the global diversity and species richness of Armillaria, we mined publicly available sequencing datasets derived from numerous primer pairs for the ribosomal operon. Our estimates reveal that species richness ranges from 50 to 60 species, depending on which marker is used. Eastern Asia represents the biogeographic region with the highest species richness. We also assess the overlap of species across geographic regions and propose some hypotheses regarding the drivers of variability in species diversity and richness between different biogeographic regions.

Transcriptomics reveals the mycoparasitic strategy of the mushroom Entoloma abortivum on species of the mushroom Armillaria

During mycoparasitism, a fungusーthe hostーis parasitized by another fungusーthe mycoparasite. The genetic underpinnings of these relationships have been best characterized in Ascomycete fungi. However, within Basidiomycete fungi, there are rare instances of mushroom-forming species parasitizing the reproductive structures, or sporocarps, of other mushroom-forming species. One of the most enigmatic of these occurs between Entoloma abortivum and species of Armillaria, where hyphae of E. abortivum are hypothesized to disrupt the development of Armillaria sporocarps, resulting in the formation of carpophoroids. However, it remains unknown whether carpophoroids are the direct result of a mycoparasitic relationship. To address the nature of this unique interaction, we analyzed gene expression of field-collected Armillaria and E. abortivum sporocarps and carpophoroids. Transcripts in the carpophoroids are primarily from E. abortivum, supporting the hypothesis that this species is parasitizing Armillaria. Most notably, we identified differentially expressed E. abortivum β-trefoil-type lectins in the carpophoroid, which we hypothesize bind to Armillaria cell wall galactomannoproteins, thereby mediating recognition between the mycoparasite and the host. The most significantly upregulated E. abortivum transcripts in the carpophoroid code for oxalate decarboxylasesーenzymes that degrade oxalic acid. Oxalic acid is a virulence factor in many plant pathogens, including Armillaria species, however, E. abortivum has evolved a sophisticated strategy to overcome this defense mechanism. The number of gene models and genes that code for carbohydrate-active enzymes in the E. abortivum transcriptome were reduced compared to other closely related species, perhaps as a result of the specialized nature of this interaction.

Towards the Biological Control of Devastating Forest Pathogens from the Genus Armillaria

Research Highlights: A large scale effort to screen, characterize, and select Trichoderma strains with the potential to antagonize Armillaria species revealed promising candidates for field applications. Background and Objectives: Armillaria species are among the economically most relevant soilborne tree pathogens causing devastating root diseases worldwide. Biocontrol agents are environment-friendly alternatives to chemicals in restraining the spread of Armillaria in forest soils. Trichoderma species may efficiently employ diverse antagonistic mechanisms against fungal plant pathogens. The aim of this paper is to isolate indigenous Trichoderma strains from healthy and Armillaria-damaged forests, characterize them, screen their biocontrol properties, and test selected strains under field conditions. Materials and Methods: Armillaria and Trichoderma isolates were collected from soil samples of a damaged Hungarian oak and healthy Austrian spruce forests and identified to the species level. In vitro antagonism experiments were performed to determine the potential of the Trichoderma isolates to control Armillaria species. Selected biocontrol candidates were screened for extracellular enzyme production and plant growth-promoting traits. A field experiment was carried out by applying two selected Trichoderma strains on two-year-old European Turkey oak seedlings planted in a forest area heavily overtaken by the rhizomorphs of numerous Armillaria colonies. Results: Although A. cepistipes and A. ostoyae were found in the Austrian spruce forests, A. mellea and A. gallica clones dominated the Hungarian oak stand. A total of 64 Trichoderma isolates belonging to 14 species were recovered. Several Trichoderma strains exhibited in vitro antagonistic abilities towards Armillaria species and produced siderophores and indole-3-acetic acid. Oak seedlings treated with T. virens and T. atrobrunneum displayed better survival under harsh soil conditions than the untreated controls. Conclusions: Selected native Trichoderma strains, associated with Armillaria rhizomorphs, which may also have plant growth promoting properties, are potential antagonists of Armillaria spp., and such abilities can be exploited in the biological control of Armillaria root rot.