Genome mining and metabolomics reveal context-dependent biosynthetic potential in a core poplar dark septate endophyte.

Dark septate endophytes (DSEs) are ubiquitous root-colonizing fungi characterized by melanized, septate hyphae and a frequent association with plants in stressful or nutrient-poor environments. Although DSEs are increasingly recognized for context-dependent effects on host stress resilience, the specialized metabolic capacity of many DSE lineages remains poorly resolved. Hyaloscypha finlandica is a recurrent root-associated fungus of Populus and other hosts, making it a useful system for understanding symbiotic mechanisms using multi-omics analyses. Here, we combined comparative biosynthetic gene cluster (BGC) prediction across 41 ascomycete genomes with untargeted metabolomics of H. finlandica PMI 746 grown in a defined minimal medium and a complex plant-derived medium. Benchmarking antiSMASH, FunBGCeX, and BGC-Prophet revealed broad conservation of PKS-, NRPS-, and terpene-associated biosynthetic capacity among DSEs. LC-MS/MS profiling detected strong medium-associated shifts in the extracellular chemical footprint of H. finlandica and identified putative terpenoid-, polyketide-, peptide-, siderophore-, and DHN-melanin-related features through spectral matching and in silico chemical-class inference. Authentic-standard comparison supported a probable assignment of 7-oxodehydroabietic acid, whereas most other annotations remain putative or class-level assignments. Integrating genomic and metabolomic evidence identified representative candidate intersections between encoded biosynthetic capacity and observed chemistry, but definitive BGC-product links remain targets for future genetic and structural validation. Together, these results provide a public paired-omics resource for a core Populus-associated DSE and a conservative framework for interpreting fungal multi-omics in non-model endophytes.IMPORTANCEDark septate endophytes (DSEs) are widespread fungal symbionts that influence plant performance, yet their encoded and expressed metabolic capacities remain largely uncharacterized. This study provides a paired comparative genome-mining and untargeted metabolomics resource for Hyaloscypha finlandica, a recurrent Populus-associated DSE. By explicitly distinguishing biosynthetic gene cluster (BGC) prediction, spectral annotation, authentic-standard support, and unresolved BGC-Prophet assignment, this work provides a transparent reference for future studies of DSE specialized metabolism. The resulting data sets support comparative analyses across fungal endophytes and help prioritize candidate metabolites and gene clusters for targeted validation.