The research team, drawn from institutions in Israel, the United States, and China, used quantitative gas chromatography coupled with tandem mass spectrometry (GC–MS/MS) to identify and measure the major soluble sugars present at four defined stages of germination: dormant conidia, the establishment of polarity, fully germinated conidia with elongated germ tubes, and the first appearance of hyphal branching. This is the first time such a quantitative, stage-resolved portrait of sugar composition has been produced for these filamentous fungi. Previous studies of spore carbohydrates in other models relied on less sensitive methods or examined only a single developmental snapshot, leaving the dynamics of the transition largely hidden.

In dormant conidia of T. asperelloides strain T203, one molecule dominated everything else: mannitol, a sugar alcohol that made up roughly 75 percent of the total measured soluble sugars and polyols. Mannitol is a long-recognized fungal reserve compound, valued both as a compact energy store and as a compatible solute that protects cells against drought, heat, and oxidative damage. But the moment the spore committed to polar growth, its mannitol reserves collapsed, falling to just 13 percent of the measured sugar pool—a sixfold drop. Then, surprisingly, the sugar came back. As the germlings progressed through germ tube extension and first branching, mannitol climbed again, reaching about 26 percent of the pool by the final stage.

Trehalose, a disaccharide famous for stabilizing proteins and membranes under stress, told the mirror-image story. In dormant conidia it accounted for a modest 6 percent of the measured sugars, roughly thirteenfold less than mannitol. As germination proceeded, trehalose surged to around 65 percent of the pool in germinated conidia and remained high at about 50 percent during first branching. Meanwhile, other tested sugars—glucose, fructose, sorbitol, and myo-inositol—stayed comparatively stable throughout. The picture that emerges is of a carefully choreographed handoff: the spore spends its mannitol savings to power the earliest steps of outgrowth, then progressively rebuilds a trehalose-rich reserve as the new hypha establishes itself, before quietly topping up the mannitol again.

That pattern turns out to be anything but universal. When the researchers extended their profiling to four reference species spanning the Sordariomycetes—the insect pathogens Metarhizium anisopliae and Cordyceps militaris, the wheat-infecting plant pathogen Fusarium graminearum, and the saprophytic model Neurospora crassa—they found sharply different storage strategies. Dormant conidia of T. asperelloides, M. anisopliae, and C. militaris were all mannitol-dominated, echoing the Trichoderma pattern. In contrast, the resting spores of N. crassa and F. graminearum held glucose as their most abundant measured carbohydrate. Once germination began, most of the species shifted toward glucose accumulation, presumably funneling reserves into glycolysis. T. asperelloides alone made the distinctive transition from mannitol to trehalose dominance before partially re-accumulating mannitol, marking it as an outlier even among close relatives.

To understand what genetic machinery underlies these shifting pools, the team integrated the sugar measurements with time-resolved RNA sequencing data covering the same developmental window. The transcriptomic landscape mirrored the metabolic one. In T. asperelloides, the most dramatic transcriptional upheaval occurred during the first transition, from dormant conidia to polar growth, when 238 metabolic-process genes were upregulated and 1,353 were downregulated—a massive clearing of the dormant transcriptome consistent with the degradation or dilution of stored messenger RNAs that are no longer needed once growth resumes. Later transitions showed a more balanced mix, with several hundred genes uniquely induced during germ tube formation and branching, reflecting renewed carbohydrate biosynthesis and central carbon metabolism coming online.

Within this broad remodeling, one gene stood out. mpd1, which encodes the enzyme mannitol-1-phosphate 5-dehydrogenase, showed its highest transcript abundance in dormant conidia and declined sharply during germination—precisely matching the collapse of the mannitol pool. Its predicted partner, mtd1, encoding mannitol dehydrogenase, is thought to catalyze the reverse reaction, oxidizing mannitol back to fructose. Together these two enzymes have long been viewed as forming the classic mannitol cycle, proposed more than four decades ago: fructose-6-phosphate is reduced to mannitol-1-phosphate, dephosphorylated to mannitol for storage, and then oxidized back to fructose when carbon is needed. If that textbook cycle were the whole story, sabotaging its biosynthetic arm should cripple mannitol accumulation.

To test that prediction, the researchers attempted to delete mpd1, but the fungus proved stubborn: gene replacement attempts yielded only heterokaryotic strains that retained the native allele, a result that itself hints at how important the pathway may be. The team therefore turned to an antisense RNA strategy, generating two independent strains in which mpd1 transcript levels were strongly and reproducibly reduced. The outcome was striking. Despite the severe transcriptional knockdown, mannitol content in dormant conidia was indistinguishable from wild type, the full soluble-sugar profile across all four germination stages was essentially unchanged, germination dynamics proceeded normally, and the knockdown strains grew on mannitol as a sole carbon source just as well as their wild-type counterparts.

The only visible response to the perturbation was a compensatory shift in mtd1. Rather than a simple across-the-board increase, the knockdown strains showed a stage-dependent redistribution of mtd1 expression, with elevated levels appearing already at polarity establishment—earlier than in wild type—suggesting the fungus reallocates flux through alternative enzymatic routes to maintain its mannitol reserves. The finding echoes earlier work in the plant pathogen Botrytis cinerea, where even double mutants lacking both canonical mannitol enzymes still synthesized and degraded the polyol and grew on it as a sole carbon source, implying unrecognized mannitol phosphorylation pathways and additional dehydrogenase-like activities. Fungal mannitol metabolism, it appears, is not a simple closed loop but a flexible, multi-branched network with built-in redundancy.

The comparative dimension of the study added an evolutionary layer to this biochemical picture. Using Bayesian network inference on the expression dynamics of mannitol-, trehalose-, glycolysis-, and TCA-cycle-associated genes, the researchers reconstructed regulatory modules for each of the five species and compared their structures using Jensen–Shannon divergence. Several core interactions were conserved across all species, confirming an ancient metabolic architecture, but other connections were lineage-specific—including edges found only in Cordyceps and a set shared specifically by Trichoderma and Metarhizium. Notably, the network-based similarity tree only partially matched the species’ organismal phylogeny, with T. asperelloides and M. anisopliae clustering together in a way that seems to reflect their shared mannitol-rich storage strategy rather than close ancestry alone. The authors caution that these network edges represent inferred associations from a limited number of sampling points, not validated causal interactions, and that future work will require complete knockout combinations and direct metabolic flux measurements to dissect the compensatory mechanisms in detail. Even so, the study delivers a clear conceptual message: storage-carbohydrate metabolism during fungal germination is robust, developmentally coordinated, and wired into central carbon metabolism through a resilient framework that tolerates the loss of individual components. For a genus whose members protect crops worldwide by parasitizing plant pathogens, understanding how spores budget their chemical reserves could ultimately inform how we deploy them—and it establishes T. asperelloides as a powerful new systems-level model for dissecting how fungi transition from dormancy to growth.

Subject of Research: Storage-carbohydrate metabolism and its transcriptional regulation during conidial germination in Sordariomycete fungi

Article Title: Conserved storage-carbohydrate metabolic modules are rewired during germination of Trichoderma asperelloides and other Sordariomycetes

Article References: Tenennbaum, B., Yakubovich, E., Wang, Y.-W., Meng, G., Moonjely, S., Trail, F., Ben Ari, J., Wang, Z., Dong, C., Townsend, J. P., & Yarden, O. (2026). Conserved storage-carbohydrate metabolic modules are rewired during germination of Trichoderma asperelloides and other Sordariomycetes. Frontiers in Fungal Biology, 7, Article 1930613. https://doi.org/10.3389/ffunb.2026.1930613

Image Credits: AI Generated

DOI: 10.3389/ffunb.2026.1930613

Keywords: Trichoderma asperelloides, conidial germination, mannitol, trehalose, Sordariomycetes, GC-MS/MS, metabolic networks, Bayesian network inference, RNA-seq, fungal metabolism, mpd1, redundancy