Comparative Study of Hirustasin Superfamily Gene Expression in Two Medicinal Leeches, Hirudinaria manillensis and Whitmania pigra

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Background/Objectives: Leeches constitute a pharmacologically significant animal group in traditional medicine due to their antithrombotic peptides, which include numerous members of the hirustasin gene superfamily. However, a comparative expression profile of this pharmaceutically important family across different leech species is lacking. Methods: This study conducted a comparative transcriptomic analysis of hirustasin gene superfamily expression in the hematophagous leech Hirudinaria manillensis and the non-hematophagous leech Whitmania pigra. Results: The total expression of the hirustasin gene superfamily, quantified as transcripts per million (TPM), showed no significant difference (p = 0.237) between H. manillensis (11,802.60 ± 1596.59) and W. pigra (8623.12 ± 965.96). However, both species exhibited pronounced intergenic expression heterogeneity. Five dominantly expressed genes (TPM > 1000) in H. manillensis and three in W. pigra were identified, collectively comprising 81% and 62% of the total hirustasin gene superfamily expression per species, respectively. Critically, the dominantly expressed genes exhibited no phylogenetic correspondence between species. Integrating expression profiles with phylogenetic reconstruction identified five high-potential candidate genes: poecistasin_Hman2, hirustasin_like_Hman01, hirustasin_like_Hman11, guamerin_Wpig, and bdellastasin_Wpig. Population-level analysis revealed marked population-specific expression patterns in H. manillensis, contrasting with minimal inter-population divergence in W. pigra. Nevertheless, geographically distinct populations of both species showed significant variation in the expression of their respective dominantly expressed genes. Conclusions: These findings provide a set of high-priority candidate genes and insights into their expression characteristics, serving as a starting point for subsequent functional validation and, when integrated with other screening methods, for future antithrombotic drug discovery.

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