The floral microbiome is of significant relevance to plant reproduction and crop productivity. While plant genotype is key to floral microbiome assembly, whether and how genotypic variation in floral traits and plant-level mutualistic and antagonistic interactions at the rhizosphere and phyllosphere influence the microbiome in the anthosphere remain little known. Using a factorial field experiment that manipulated biotic interactions belowground (mycorrhizae treatments) and aboveground (herbivory treatments) in three strawberry genotypes, we assessed how genotypic variation in flower abundance and size and plant-level biotic interactions influence the bidirectional relationships between floral volatile organic compounds (VOCs) and the floral microbiome using structural equation modeling. We found that plant genotype played a stronger role, overall, in shaping the floral microbiome than biotic interactions with mycorrhizae and herbivores. Genotypic variation in flower abundance and size influenced the emission of floral VOCs, especially terpenes (e.g. α- and β-pinene, ocimene isomers) and benzenoids (e.g. p-anisaldehyde, benzaldehyde), which in turn affected floral bacterial and fungal communities. While the effects of biotic interactions on floral traits including VOCs were weak, mycorrhizae treatments (mycorrhizae and herbivory + mycorrhizae) affected the fungal community composition in flowers. These findings improve our understanding of the mechanisms by which plant genotype influences floral microbiome assembly and provide the first evidence that biotic interactions in the rhizosphere and phyllosphere can influence the floral microbiome, and offer important insights into agricultural microbiomes.

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