Bacterial communities on upper and lower leaf surfaces show distinct seasonal response patterns

Leaf surfaces represent one of the largest microbial habitats on Earth, which can be divided into upper and lower leaf surfaces. Upper and lower leaf surfaces offer contrasting microenvironments that vary through the seasons. This variation has rarely been quantified; however, in spite of it being highly relevant for leaf-microbiome contributions to terrestrial ecosystems, especially forests. To address this gap, we tracked bacterial communities on both surfaces of the same pooled leaves of Quercus robur in situ from spring to autumn, using genetically identical ramets to control host and genotypic variation. We combined 16S rRNA sequencing with analyses of leaf structural and physiological traits to link bacterial taxonomic, phylogenetic, and assembly characteristics. The two surfaces hosted distinct bacterial communities whose divergence intensified through the growing season. Upper-surface bacterial diversity remained relatively stable and was mainly associated with dynamic leaf traits, whereas lower-surface communities showed faster compositional and assembly-related shifts and were associated with both dynamic and stable leaf traits. These contrasting seasonal patterns reveal that within-leaf heterogeneity represents a fundamental axis of variation for bacterial seasonal dynamics, providing new insight into how microhabitat structure and host traits jointly shape phyllosphere functioning.