Basidiomycete fungi are an underexplored source of specialized metabolites with potential biotechnological applications. The environmental triggers that activate fungal biosynthetic pathways are not well understood. This study investigated how nutritional competition with the bacterium Escherichia coli affects the wood-decaying fungus Fomitopsis betulina, using both metabolomics and transcriptomics to characterize chemical and gene-expression responses to co-culture.
When grown on solid medium in the presence of E. coli, F. betulina produced an inhibition zone in which bacterial growth was suppressed. A mass spectrometry–based metabolomics analysis of material from the inhibition zone identified several compounds that were enriched compared with surrounding medium. Reported enriched molecules included a Sumiki’s acid derivative, calcium diformate, and sulfuric acid. The authors interpret these findings as consistent with acidification of the medium within the interaction zone, which may contribute to bacterial growth inhibition.
Although F. betulina has previously been reported to produce an antibacterial alkaloid, piptamine, the current study did not detect piptamine or structurally related derivatives under the experimental conditions applied. The authors note that these compounds were not observed in their metabolomic profiles from the co-culture assays.
In liquid co-culture experiments, the presence of E. coli led to rapid consumption of available glucose. This resulted in carbon-starvation conditions for the fungus and was associated with a 44% reduction in fungal biomass relative to control conditions. The rapid depletion of the primary carbon source established a nutritional stress environment that served as the dominant physiological perturbation in the liquid co-cultures.
Global transcriptomic analyses of F. betulina exposed to bacterial competition revealed extensive metabolic reprogramming. Genes involved in carbon acquisition and nutrient transport were upregulated, consistent with cellular attempts to scavenge limited resources. Transcripts associated with redox homeostasis and general stress adaptation also increased, indicating engagement of protective and homeostatic mechanisms in response to the altered environment.
Among the transcriptional changes, a homolog of the Velvet regulatory complex was upregulated. The Velvet complex is recognized as a central regulator of fungal development and specialized metabolism in other fungal systems. Concurrently, the co-culture condition induced expression of genes located in multiple biosynthetic gene clusters, including those annotated for terpene, polyketide, and fungal RiPP (ribosomally synthesized and post-translationally modified peptide) pathways. These expression changes indicate activation of secondary metabolism programs in F. betulina during bacterial competition.
The combined chemical and transcriptomic data indicate that bacterial competition can act as a potent environmental trigger for nutritional stress and secondary metabolism in F. betulina. The detection of acidification-associated compounds in the solid-medium inhibition zone suggests one mechanism for interkingdom antagonism in that setting. In liquid culture, nutrient depletion rather than localized chemical secretion appears to be the primary stressor driving transcriptional shifts.
These observations support the concept that co-culture with bacteria is a viable strategy to induce otherwise silent or low-expression biosynthetic pathways in basidiomycetes. The authors propose that fungal–bacterial co-culture could serve as an alternative or complementary approach to direct extraction-based screening for identifying high-value specialized metabolites from basidiomycete fungi.
Co-culture of Fomitopsis betulina with Escherichia coli produced measurable antagonism on solid medium and caused rapid glucose depletion and carbon-starvation in liquid culture. These conditions led to broad transcriptional reprogramming of fungal metabolism, upregulation of a Velvet homolog, and induction of multiple biosynthetic gene clusters (terpene, polyketide, and fungal RiPP). Previously reported antibacterial piptamine was not detected under the experimental conditions. The study concludes that bacterial competition is an effective trigger for nutritional stress and activation of secondary metabolism in this basidiomycete and highlights co-culture as a promising method to reveal specialized metabolite pathways.