A synergistic culture dependent and independent approach reveals a conserved wheat seed mycobiome

The occurrence of pathogenic fungal taxa associated with wheat (Triticum aestivum L.) seeds is well studied, but less is known about non-pathogenic taxa of the wheat seed mycobiome. The goal of our research is to characterize wheat seed fungal endophyte diversity with a synergistic culture dependent and independent experimental approach. Four publicly available winter wheat cultivars developed in the southeastern United States with varying phenotypic and disease resistance traits were examined over a period of two years: Catawba, Hilliard, Shirley, and USG 3640. Our culture dependent methods involving two nutrient media generated 645 fungal isolates representing twelve genera sampled from multiple cultivars. Metabarcoding analysis identified a broader range of fungal taxa and a greater number of unique sequences than culture dependent methods. When examining fungal diversity across cultivars and years, richness decreased in 2021 for both culture dependent and independent approaches. However, wheat seed fungal community structure was stable across cultivars and years. Our results highlight the importance of combining culture independent and dependent methods to capture and establish a diverse endophytic fungal catalog associated with the wheat seed and highlight areas where future culture dependent efforts can focus their efforts.