Using rice-based solid-state fermentation, the researchers cultivated I. obliquus mycelia and detected three important triterpenoids: inotodiol, ergosterol, and lanosterol. By combining chemical analysis, time-series transcriptomics, and quantitative polymerase chain reaction (qPCR) validation, the study linked increasing triterpenoid accumulation with rising expression of key biosynthetic genes. These findings provide a genetic foundation for future work to improve sustainable production of Chaga-derived compounds and to clarify fungal triterpenoid biosynthesis.
Inonotus obliquus is a medicinal macrofungus that naturally grows on birch trees in high-latitude regions, but its sclerotia develop slowly and overharvesting may threaten natural resources. Triterpenoids from this fungus, especially lanostane-type compounds such as inotodiol, lanosterol, and ergosterol-related molecules, have attracted interest for their biological activities and potential pharmaceutical value. Previous studies have explored elicitors, culture conditions, and omics approaches to increase triterpenoid production, yet the specific biosynthetic route leading to compounds such as inotodiol remains insufficiently understood. These limitations highlight the need to investigate how I. obliquus produces and modifies triterpenoids under controllable cultivation conditions.
A study (DOI:10.48130/panfungi-0025-0002) published in Panfungi on 02 February 2026 by Hirokazu Kawagishi & Chengwei Liu’s team, Northeast Forestry University, reports that rice-grown mycelia can accumulate key triterpenoids while revealing candidate genes associated with their biosynthesis and modification.
To investigate triterpenoid production, the researchers first cultivated I. obliquus strain CT5 on a rice-based solid medium and extracted metabolites from mycelia at different growth stages. Inotodiol was purified using silica gel chromatography and semi-preparative high-performance liquid chromatography (HPLC), and its structure was confirmed by nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography–mass spectrometry (LC-MS). The team then used gas chromatography–mass spectrometry (GC-MS) and HPLC to detect and quantify lanosterol, ergosterol, and inotodiol at 5, 10, and 15 days of cultivation. The chemical results showed a clear time-dependent increase in all three triterpenoids. At 15 days, the levels reached 7.33 mg/kg for inotodiol, 14.61 mg/kg for ergosterol, and more than 108.65 mg/kg for lanosterol, indicating that the mycelial stage can serve as a practical source of these compounds. Because natural sclerotium formation is slow, this rice-based cultivation approach offers a shorter and more controllable route for obtaining medically valuable triterpenoids. The researchers next performed RNA sequencing on mycelia collected at 5 and 10 days to capture gene-expression changes during the period when triterpenoid levels were rising. Sequencing generated 40.54 Gb of high-quality data, with Q30 values above 98.24% and high alignment rates to the reference genome. Comparative transcriptome analysis identified 316 differentially expressed genes, including 180 upregulated and 136 downregulated genes. Functional annotation showed enrichment in carbohydrate metabolism, amino acid metabolism, secondary metabolite biosynthesis, catalytic activity, and transport-related functions. Further analysis focused on the mevalonate (MVA) pathway, the main fungal route for terpene biosynthesis, and the downstream ergosterol pathway. Genes encoding squalene synthase, squalene epoxidase, lanosterol synthase, lanosterol 14α-demethylase, and sterol C-22 desaturase showed increasing expression over time, matching the observed rise in triterpenoid content. qPCR validation confirmed the transcriptome trends. The team also identified cytochrome P450s and short-chain dehydrogenases (SDRs) as possible modifying enzymes that may help diversify lanostane-type triterpenoids in I. obliquus.
Overall, the study provides both a cultivation strategy and a molecular roadmap for exploring triterpenoid biosynthesis in I. obliquus. By linking metabolite accumulation with candidate biosynthetic and modification genes, the work advances understanding of how Chaga mushroom produces valuable triterpenoid compounds. Future studies using isotope labeling, heterologous expression, and gene knockout or knockdown experiments could verify the functions of these candidate genes and support more efficient production of fungal natural products.
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References
DOI
10.48130/panfungi-0025-0002
Original Source URL
https://doi.org/10.48130/panfungi-0025-0002
Funding Information
This work was supported by the Fundamental Research Funds for the Central Universities (No. 2572023AW40), the National Natural Science Foundation of China (Nos 32370069 and U22A20369), and the Natural Science Foundation of Heilongjiang Province of China (No. LH2023C035).
About Panfungi
Fungi comprise one of Earth's most diverse and ecologically vital kingdoms, spanning macroscopic forms such as edible and medicinal mushrooms, as well as pathogenic species, slime molds, and innumerable microscopic organisms. Panfungi (e-ISSN 3142-7847) is an open access, online journal committed to publishing high-quality, rigorously peer-reviewed original research articles, reviews, methodological developments, and perspective articles that advance the comprehensive study of fungal biology and its applications.