Small RNAs regulation and genomic harmony: insights into allopolyploid evolution in marsh orchids (Dactylorhiza)

O_LIHybridization and polyploidy are major drivers of plant diversification, often accompanied by shifts in gene expression and genome composition. Small RNAs (smRNAs) are thought to influence such genomic changes, particularly through their interactions with transposable elements (TEs). C_LIO_LIWe comparatively quantified smRNAs in established sibling allopolyploids Dactylorhiza majalis and D. traunsteineri and their diploid progenitors to assess how independent allopolyploidization events shaped smRNA landscapes. C_LIO_LIDespite independent origins, the allotetraploids exhibited substantial overlap in smRNA composition, including transgressive accumulation of smRNA near genes related to transcriptional regulation, cell division and stress response. Consistently, TE-associated 24 nt smRNAs more closely resembled the paternal and larger genome, while shorter smRNAs typically reflected the maternal and smaller genome. Nevertheless, distinct patterns were also evident: the older D. majalis showed greater accumulation of smRNAs near genes involved in transcriptional and translational regulation, while the younger D. traunsteineri displayed stronger non-additive patterns, suggesting ongoing resolution of post-polyploid meiotic and mitotic instability. C_LIO_LIOur results reveal both convergence and divergence in smRNA landscapes among independently formed allopolyploids. Our study highlights the central role of smRNAs in resolving genomic conflict, with possible implications for functional divergence and ecological innovation during polyploid evolution. C_LI Plain-language summaryWe studied two sibling marsh orchid species that arose through hybridization and genome doubling. By comparing their small RNA molecules and how these associate with genes, we found both shared and species-specific patterns. These differences reflect how the orchids genomes have changed since formation, and may explain how repeated polyploidy events contribute to genetic and ecological diversity in plants.