Hygromycin B Is a Highly Versatile Inducer of Secondary Metabolite Production Applicable Across Diverse Fungal Taxa

Hygromycin B, a well-known antibiotic agent, was recently reported as an inducer of secondary metabolites for a fungal strain of Fusarium sp. In this study, we verified the versatility and usefulness of hygromycin B as an inducer of fungal secondary metabolite production. We demonstrate that hygromycin B induces the production of various fungal secondary metabolites, including polyketides, peptides, and terpenes, in 71% of 28 fungal strains belonging to the Ascomycota phylum, specifically within the Eurotiomycetes, Sordariomycetes, and Dothideomycetes classes. We also demonstrate the discovery of novel compounds, named hannocateol (1) and shirazines A (2) and B (3), from the hygromycin B-supplemented cultured materials. The production of the novel compounds increased with hygromycin B-supplementation up to 100 μg/g (maximum >100-fold) but not by HDAC inhibitors, protein synthesis inhibitors against prokaryotes, and antifungal agents. Notably, protein synthesis inhibitors against eukaryotes similar to hygromycin B, such as 5-fluorouracil and paromomycin, could induce the production of 1 and 2, suggesting that its mechanism would be associated with protein synthesis inhibition. We proved that hygromycin B is a highly versatile inducer of secondary metabolite production applicable across diverse fungal taxa, making it a simple and powerful tool for uncovering cryptic biosynthetic pathways and discovering novel bioactive compounds.