Abstract

Fragrance is a crucial quality trait in economically significant crops. However, enhancing a species with either no scent or weak fragrance presents substantial challenges. Distant hybridization serves an effective strategy to improve the fragrance traits in such species. This study examined the phenotypic variations and fragrance changes in Epidendrum floribundum 'GZSL' (♀), which has a strong fragrance but short single-flower blooming duration, Encyclia tampensis 'LRY' (♂), which exhibits weaker fragrance but longer blooming duration, as well as their distant hybrids × Epicyclia 'LY'. Results revealed that 'LY' exhibited heterosis in aroma intensity, plant height, and inflorescence length, with most heterosis indices exceeding 1.6. Further analysis of floral VOCs (volatile organic compounds) across the three materials indicated that differences in aroma intensity during peak bloom were primarily due to the quantity and variety of terpenoid VOCs, rather than their relative content. Using PLS-DA and VIP value analysis, we identified VOCs such as linalool, β-myrcene, ( E )-β-ocimene, ( E )-2-nonenal, hotrienol, and bicyclosesquiphellandrene as potential key contributors to the strong aroma intensity of 'LY'. RNA-seq and KEGG analyses showed that the upregulated expression of structural genes in the terpenoid biosynthesis pathway contributed to the richer and higher release of terpenoid VOCs in 'LY' and 'GZSL' compared to 'LRY'. Additionally, transcription factors such as ERFs, MYBs, and bHLHs may regulate terpenoid biosynthesis. These findings offer valuable insights for improving fragrance intensity in Orchidaceae through intergeneric distant hybridization.