Beyond detection: Unveiling microbial dynamics in oak seedlings using isolation and high-throughput amplicon sequencing

Background Forest nurseries are critical for producing resilient nursery stock for reforestation and planting of ornamental trees, yet the microbial communities associated with nursery-grown plants remain poorly characterized. Quercus robur L. seedlings from seven Czech forest nurseries were analyzed to assess microbial diversity, co-occurrence patterns, and environmental drivers. Results Microbial communities were characterized using high-throughput amplicon sequencing (HTAS), fungal isolation, and comprehensive soil chemistry. HTAS revealed broader taxonomic and functional profiles compared to isolation, which selectively enriched for fast-growing pathogens. Both methods were found to be complementary, emphasizing the value of methodological integration. Fungal communities exhibited pronounced site-specific beta diversity and responded strongly to inorganic soil parameters, particularly calcium, phosphorus, and pH/CaCl ₂ . Bacterial communities were more spatially cohesive and primarily associated with humification-related factors. Trends in fungal alpha diversity were observed in relation to organic matter fractions (e.g., C _ox , D _H ). Network analysis revealed predominantly positive intra- and inter-kingdom interactions, indicating potential cooperation or niche complementarity. Pathogens were embedded within broader trophic networks, and their ecological behavior appeared modulated by co-occurring taxa, supporting the pathobiome concept. Conclusions Mineral soil properties were identified as key drivers of microbial community structure and trophic composition in oak seedling tissues. Despite standardized nursery conditions, edaphic variability exerted strong filtering effects on fungal and bacterial communities. These findings provide ecological insight into seedling–microbe interactions and offer a basis for microbiome informed nursery management strategies.