A Comparative Study on the Expression Levels of the HMEI Gene Family in Hirudinaria manillensis and Whitmania pigra

Leeches are classic animal-derived traditional Chinese medicines for activating blood circulation and eliminating blood stasis. The hematophagous Hirudinaria manillensis and the non-hematophagous Whitmania pigra are the two most widely used medicinal leeches, but their HMEI (H. manillensis elastase inhibitor) gene family, a gene family containing candidate elastase-inhibitor paralogs, has not been systematically compared across species and geographical populations. Here, we performed RNA-Seq on head tissues from five populations of each species and compared the expression of all HMEI paralogs. The 38 paralogs formed 9 descriptive sequence groups. Total HMEI expression showed no significant interspecies difference, yet the two species showed distinct transcriptional patterns in the sampled populations: expression in H. manillensis was concentrated in a few paralogs with ultra-high abundance, whereas expression in W. pigra was distributed across multiple moderately expressed paralogs. Total HMEI expression did not differ significantly among H. manillensis populations, whereas significant inter-population differences were detected in W. pigra. Four expression-based candidates for functional validation were identified: HMEI_Hman06 and HMEI_Hman15 from H. manillensis, and HMEI_Wpig01 and HMEI_Wpig17 from W. pigra. These results characterize interspecies and population-associated variation in HMEI expression, providing a transcriptomic resource for comparative biochemical and functional characterization of HMEI paralogs.