Diversity and selective pressures of anticoagulants in three medicinal leeches (Hirudinida: Hirudinidae, Macrobdellidae)

Abstract

Although medicinal leeches have long been used as treatment for various ailments because of their potent anticoagulation factors, neither the full diversity of salivary components that inhibit coagulation, nor the evolutionary selection acting on them has been thoroughly investigated. Here, we constructed expressed sequence tag libraries from salivary glands of two species of medicinal hirudinoid leeches, Hirudo verbana and Aliolimnatis fenestrata, and identified anticoagulant-orthologs through BLASTx searches. The data set then was augmented by the addition of a previously constructed EST library from the macrobdelloid leech Macrobdella decora. The identified orthologs then were compared and contrasted with well-characterized anticoagulants from a variety of leeches with different feeding habits, including non-sanguivorous species. Moreover, four different statistical methods for predicting signatures of positive and negative evolutionary pressures were used for 10 rounds each to assess the level and type of selection acting on the molecules as a whole and on specific sites. In total, sequences showing putative BLASTx-orthology with five and three anticoagulant-families were recovered in the A. fenestrata and H. verbana EST libraries respectively. Selection pressure analyses predicted high levels of purifying selection across the anticoagulant diversity, although a few isolated sites showed signatures of positive selection. This study represents a first attempt at mapping the anticoagulant repertoires in a comparative fashion across several leech families.

Be ready at any time: postprandial synthesis of salivary proteins in salivary gland cells of the haematophagous leech Hirudo verbana

“…These bioactive substances are produced and stored in unicellular salivary gland cells in the anterior body part of the leech and secreted during feeding (Ascenzi et al, 1995;Baskova et al, 1992;Baskova and Zavalova, 2001;Hildebrandt and Lemke, 2011;Kvist et al, 2013;Müller et al, 2015). We have recently shown (Hildebrandt and Lemke, 2011), and confirmed in this study (Fig.…”

“…The European leeches Hirudo medicinalis and Hirudo verbana are model animals for studying the biology of haemotaphagous leeches in general and for investigating the pharmacological properties of their salivary gland proteins with respect to potential targets in host animals (Kvist et al, 2013;Siddall et al, 2007). These leeches feed on body fluids of amphibians, fish, birds, reptiles and mammals (Ceylan and Erbatur, 2012;Dickinson and Lent, 1984;Kutschera and Roth, 2005;Merilä and Sterner, 2002;Sawyer, 1986).…”

“…During feeding, leeches release bioactive salivary proteins and peptides (Baskova et al, 2004;Baskova and Zavalova, 2001;Hildebrandt and Lemke, 2011), produced in unicellular salivary glands in the anterior segments of the leech body, into the wound (Hildebrandt and Lemke, 2011). Only a handful of these substances out of up to 100 that may be present in leech saliva (Baskova et al, 2004) are known so far, among them enzymes, anti-inflammatory or antimicrobial agents and inhibitors like anti-coagulants (Ascenzi et al, 1995;Baskova et al, 1992Baskova et al, , 2008Baskova and Zavalova, 2001;Deckmyn et al, 1995;Greinacher and Warkentin, 2008;Gronwald et al, 2008;Hildebrandt and Lemke, 2011;Kvist et al, 2013;Müller et al, 2015;Rigbi et al, 1996;Vilahur et al, 2004). During one blood meal that lasts at least 20-30 min (Lent et al, 1988), up to 1 mg of salivary protein is secreted into the wound (Lemke et al, 2013), resulting in partial or complete emptying of the approximately 40,000 salivary gland cells (Lemke et al, 2013).…”

Transcriptomics and differential gene expression in Whitmania pigra (Annelida: Clitellata: Hirudinida: Hirudinidae): Contrasting feeding and fasting modes

“…F I G U R E 2 Results from the gene ontology analysis (examined using topGO), illustrating the log scale proportional devotion of the salivary transcripts to various processes regarding biological process, cellular component, and molecular function anticoagulant studies (e.g., Min, Sarkar, & Siddall, 2010;Kvist, Min, & Siddall, 2013;Kvist et al, 2014;Siddall et al, 2016;Tessler, Marancik et al, 2018) including several putative orthologs beyond the archetypal sequences (the putative guamerin sequence from W. pigra was aligned with several known, putatively paralogous factor Xa inhibitors derived from leeches, in order to tease out the most likely ortholog for this sequence). Phylogenetic analyses were performed in order to robustly infer orthology between the newly derived sequences and the archetypal anticoagulants.…”

Effects of light spectrum and intensity on growth, survival and physiology of leech (Whitmania pigra ) larvae under the rearing conditions

“…Whitmania pigra Whitman has been used as a traditional Chinese medicine in cardiovascular diseases and other chronic diseases for more than 2000 years, which demonstrates wide pharmacological effects such as anti-platelet agglomeration and decreasing blood lipids (Chinese Pharmacopoeia Editorial Committee 2015; Chen, Li, Li, Wang, Li, Ni & Koike 2011;Kvist, Min & Siddall 2013;Wang, Zhao, Wang, Song, Liang & Ji 2014). In recent years, leech (Whitmania pigra, Whitmania acranulata, Hirudo nipponica et al) populations have declined dramatically due to over-exploitation for medicinal purposes and environmental pollution (Laima 2003).…”