One Enzyme Lets a Common Root Fungus Infect Plants Across the Globe

A fungus that quietly lives inside the roots of healthy plants across Europe has turned out to be one of the most revealing pathogens ever studied at the root–soil interface. In a sweeping genomic and functional analysis published in Nature Microbiology, researchers report that Plectosphaerella cucumerina, a soil-borne fungus long considered an emerging but poorly understood crop pathogen, is in fact a core member of the root mycobiota of the model plant Arabidopsis thaliana. More strikingly, the team has pinpointed a single carbohydrate-degrading enzyme that acts as a master key, allowing the fungus to breach the roots of multiple unrelated plant species. The finding offers a rare mechanistic explanation for how some fungi achieve global distribution while infecting an astonishing breadth of hosts.

The story began with a reanalysis of fungal DNA sequencing data from 291 Arabidopsis root samples collected at 18 sites across Europe. From more than 26 million sequenced reads targeting the fungal internal transcribed spacer region, the researchers identified 338 fungal sequence variants present in roots. Only five of these were reproducibly detected in more than 80 percent of the samples, defining a tiny multispecies core mycobiota. Among these five, one variant stood out: it was detected in 98 percent of root samples, and when the team cross-referenced it against 488 cultured fungal isolates from four of the sampling sites, 18 percent of those isolates matched it perfectly. The organism behind this ubiquitous signature was Plectosphaerella cucumerina.

To understand how widespread this fungus really is, the researchers queried the GlobalFungi database, a global repository of fungal occurrence data from environmental sequencing studies. Plectosphaerella sequences appeared in soil, mosses, topsoil, rhizosphere and root samples distributed across the planet, and the plants associated with these detections spanned a phylogenetically diverse array of monocot and dicot hosts. Among the five most prevalent root-associated fungal variants, Plectosphaerella showed the strongest enrichment at the soil–root interface, confirming that it is not merely a soil dweller but an active root colonizer with a host range that cuts across the plant tree of life.

With the ecological stage set, the team assembled a collection of 72 Plectosphaerella strains isolated from plants as diverse as mosses, grasses and flowering dicots, gathered from multiple continents. Using PacBio long-read sequencing, they generated high-quality genome assemblies ranging from 35.5 to 40.5 million base pairs. A phylogeny built from 5,466 single-copy genes revealed two subspecies and a small outgroup, with a burst of transposable elements in one subspecies. Yet the most important result was what the genomes did not show: when the researchers tested whether the host plant from which each strain had been isolated explained variation in the fungi’s repertoires of carbohydrate-active enzymes, proteases and candidate effectors, it did not. Fungal phylogeny accounted for roughly half of the variance in these gene families, but host identity had no detectable effect. The fungus, in other words, carries no signature of specialization on particular plants.

Functional assays reinforced this picture of a generalist. Most isolates, regardless of their host of origin, were detrimental to Arabidopsis seedlings grown in sterile culture, and two Arabidopsis-derived strains retained their harmful effects across 12 geographically diverse Arabidopsis accessions. All 72 isolates consumed carbon sources with nearly identical profiles, showing a particular aptitude for plant cell wall-derived monosaccharides. The modest genetic variation among strains therefore seemed unlikely to explain the fungus’s remarkable environmental and host flexibility on its own, pointing the investigators toward gene regulation rather than gene content as the decisive factor.

To test this, the researchers performed controlled recolonization experiments in which germ-free Arabidopsis, tomato and barley seedlings were inoculated with a single fungal strain originally isolated from Arabidopsis roots. The results were strikingly host-dependent. The fungus significantly reduced a combined index of plant survival and growth by 19.5 percent in Arabidopsis and 10.7 percent in tomato, both dicots, but had no measurable effect on barley, a monocot. Quantitative PCR showed that the fungus colonized the roots of all three species, including barley, but confocal microscopy revealed a crucial difference: in Arabidopsis, fungal hyphae penetrated deep into the peridermal and cortical cell layers, establishing extensive endophytic growth throughout the root system. In tomato, invasion was confined largely to the epidermal layer. In barley, hyphae clustered around root hairs but never entered the root interior, suggesting that the monocot’s immune system or structural barriers halt the fungus at the surface.

The transcriptomic explanation for this host specificity emerged when the team sequenced fungal gene expression inside the roots of all three plants. Of 3,399 fungal genes significantly regulated during root colonization compared with growth on inert membranes, most responded in a host-specific manner, revealing remarkable transcriptional plasticity. Genes involved in catabolism were disproportionately represented among the upregulated sets, and genes encoding carbohydrate-active enzymes, the molecular tools fungi use to dismantle plant cell walls, were strongly overrepresented among induced genes in every host condition. Crucially, the specific enzymes activated mirrored the cell wall chemistry of each host. In the pectin-rich walls of the two dicots, the fungus induced pectin-degrading enzymes; in barley, whose walls are dominated by xylans and mixed-linkage glucans, it switched on xylan-degrading enzymes instead. The fungus, it appears, reads the chemical composition of its host’s walls and tailors its enzymatic arsenal accordingly.

One gene rose above all others in this analysis: a single-copy gene encoding a secreted enzyme from glycoside hydrolase family 64, a candidate β-1,3-glucanase whose expression was induced more than 100-fold inside roots regardless of host species. Structural prediction placed it close to a biochemically characterized β-1,3-glucanase from a soil bacterium, an enzyme with strict specificity for β-1,3-linked glucose chains. These linkages occur in plant callose and mixed-linkage glucans, but they are also the principal immunogenic components of fungal cell walls, meaning the enzyme could serve the fungus in two ways: dismantling host defenses or masking its own wall from immune detection. When the researchers deleted the gene using CRISPR-Cas9 genome editing, confirmed by whole-genome resequencing to be a clean single insertion, the consequences were dramatic. Pathogenicity on Arabidopsis and tomato dropped by 62 to 77 percent, and root colonization fell by 18 to 24 percent, while growth in artificial media and colonization of barley were unaffected. The enzyme, in short, is a specific determinant of infection in the hosts the fungus can actually invade.

The evolutionary reach of this mechanism became apparent when the team surveyed 2,534 fungal genomes. GH64 genes are significantly enriched in plant saprotrophs, pathogens and endophytes, and multiple species in the destructive genus Colletotrichum carry a single copy. Deleting the orthologous gene in Colletotrichum incanum, a root pathogen from a different fungal family that diverged from Plectosphaerella long ago, reduced fungal load in Arabidopsis roots by 68.5 percent and increased plant shoot weight by 10 percent. Notably, the gene was dispensable for infection of Nicotiana benthamiana, consistent with its low expression in that host’s roots. Two independently evolved pathogens thus rely on the same enzyme family to infect the same host, demonstrating that host-induced carbohydrate-active enzymes can couple fungal virulence with multihost compatibility. Because core root fungi are normally held in check by bacterial commensals and plant immune metabolites in nature, the authors suggest that disease emerges when these restraints fail. The GH64 enzyme and its relatives now stand out as promising targets for protecting crops against broad-host-range root pathogens, and the study more broadly reframes fungal host range not as a fixed genomic property but as a dynamic regulatory capacity, unlocked one enzyme at a time by the chemical cues of whatever root the fungus encounters.

Subject of Research: Carbohydrate-active enzymes enabling multihost root infection by the fungus Plectosphaerella cucumerina

Article Title: Carbohydrate-active enzymes from a core root mycobiota member enable infection of multiple plant hosts

Article References: Raja-Kumar, R.-S., Mesny, F., Basak, A. K., Newfeld, J., Chesneau, G., Entila, F., Lee, T., Rigerte, L., Carvajal Acevedo, S., Hüttel, B., Crous, P. W., Maciá-Vicente, J. G., Stewart, H., Ryan, M., Fakhoury, A. M., Sacristán, S., Aitouguinane, M., Batisson, I., Dumontet, S., … Hacquard, S. (2026). Carbohydrate-active enzymes from a core root mycobiota member enable infection of multiple plant hosts. Nature Microbiology. https://doi.org/10.1038/s41564-026-02492-3

Image Credits: AI Generated

DOI: 10.1038/s41564-026-02492-3

Keywords: Plectosphaerella cucumerina, root microbiome, CAZymes, GH64, β-1,3-glucanase, plant pathology, Arabidopsis thaliana, Colletotrichum incanum, fungal genomics, cell wall degradation, host range, CRISPR gene editing