Revisiting the Asian Buffalo Leech (Hirudinaria manillensis) Genome: Focus on Antithrombotic Genes and Their Corresponding Proteins

Abstract

Leeches are well-known annelids due to their obligate blood-feeding habits. Some leech species secrete various biologically active substances which have important medical and pharmaceutical value in antithrombotic treatments. In this study, we provided a high-quality genome of the Asian buffalo leech (Hirudinaria manillensis), based on which we performed a systematic identification of potential antithrombotic genes and their corresponding proteins. Combining automatic and manual prediction, we identified 21 antithrombotic gene families including fourteen coagulation inhibitors, three platelet aggregation inhibitors, three fibrinolysis enhancers, and one tissue penetration enhancer. A total of 72 antithrombotic genes, including two pseudogenes, were identified, including most of their corresponding proteins forming three or more disulfide bonds. Three protein families (LDTI, antistasin, and granulin) had internal tandem repeats containing 6, 10, and 12 conserved cysteines, respectively. We also measured the anticoagulant activities of the five identified hirudins (hirudin_Hman1 ~ hirudin_Hman5). The results showed that three (hirudin_Hman1, hirudin_Hman2, and hirudin_Hman5), but not the remaining two, exhibited anticoagulant activities. Our study provides the most comprehensive collection of antithrombotic biomacromolecules from a leech to date. These results will greatly facilitate the research and application of leech derivatives for medical and pharmaceutical purposes in the treatment of thrombotic diseases.

Annelid comparative genomics and the evolution of massive lineage-specific genome rearrangement in bilaterians

“…Those with an accompanying publication are Acholoe squamosa (Adkins et al 2023), Alitta virens (Fletcher et al 2023), Lepidonotus clava (Darbyshire et al 2022), Piscicola geometra (Doe et al 2023), and Sthenelais limicola (Darbyshire et al 2023). Genomes from other sources with accompanying publications are: Branchipolynoe longqiensis (He et al 2023), Hirudinaria manillensis (Liu et al 2023), Metaphire vulgaris (Jin et al 2020), Owenia fusiformis , (Martín-Zamora et al 2023), Paraescarpia echinospica (Sun et al 2021) , Streblospio benedicti (Zakas et al 2022), Sipunculus nudus (Zheng et al 2023), and Urechis unicinctus (Cheng et al 2024).…”

A Chromosome-Level Genome Assembly of the Non-Hematophagous Leech Whitmania pigra (Whitman 1884): Identification and Expression Analysis of Antithrombotic Genes

“…The DNA and RNA sequencing were performed with reference to our recently published article [27]. Briefly, a HiFi SMRTbell library was created using the SMRTbell Express Template Prep Kit 2.0 (PacBio, Menlo Park, CA, USA) and were sequenced by the PacBio Sequel II platform.…”

“…Recently, we used state-of-the-art third-generation sequencing (PacBio HiFi) and next-generation sequencing (Illumina Hi-C, Survey and RNA-seq) to obtain a nearly complete chromosome-scale genome of a blood-feeding H. manillensis. Based on the highquality genome, we systematically identified 72 antithrombotic genes involving 21 gene families [27], many of which were not predicted in previous studies.…”

“…Here, we used the third generation (PacBio HiFi) and the next-generation sequencing methods (Illumina Hi-C, Survey and RNA-Seq) to obtain a nearly complete chromosomescale genome of the W. pigra. With a so called BRAKER-plus gene prediction strategy which combined process-oriented and manual prediction approaches [27], we systematically identified the antithrombotic related genes of W. pigra. Combined with the results from H. manillensis [27], we compared the gene constitutions between the nonhematophagous and hematophagous leeches.…”

“…With a so called BRAKER-plus gene prediction strategy which combined process-oriented and manual prediction approaches [27], we systematically identified the antithrombotic related genes of W. pigra. Combined with the results from H. manillensis [27], we compared the gene constitutions between the nonhematophagous and hematophagous leeches. Meanwhile, we provided RNA-seq data of the W. pigra and H. manillensis and calculated the expression levels of the antithrombotic genes of the two species.…”