Search Results (7)

Leech therapy is a biotherapeutic approach that has been traditionally used for centuries and is currently being re-evaluated in modern medicine. The efficacy of this treatment is attributed to various bioactive compounds found in leech saliva, which exhibit anticoagulant, anti-inflammatory, antioxidant, and anticancer properties. It has been demonstrated that leech saliva possesses the potential to modulate inflammatory processes and apoptotic mechanisms. In this study, the therapeutic potential of the saliva of Hirudo verbana was evaluated, and its biological and pharmacological effects were comprehensively investigated. The anticancer effects, antioxidant capacity, and anti-inflammatory activity of the crude leech saliva were assessed using human umbilical vein endothelial cells and epithelial ovarian cancer cells. The chemical composition of the saliva was analyzed using gas chromatography–mass spectrometry, while the protein content was determined by the Bradford assay. Antioxidant activity was measured using the 2,2-diphenyl-1-picrylhydrazyl radical scavenging assay, inflammatory effects were evaluated by Enzyme-Linked ImmunoSorbent Assay, and cell viability was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The findings revealed that crude leech saliva had a minimal effect on healthy cells but showed a selective effect on the viability of ovarian cancer cells. At low concentrations (3.13%), 99.16% of healthy cells remained viable, whereas this rate decreased to 89.25% in cancer cells; at high concentrations (50%), cell viability in cancer cells declined to 63.02%. Gas chromatography–mass spectrometry analysis identified compounds such as gibberellic acid and 6-[(4-methoxyphenyl)methoxy]-4,4,5,7,8-pentamethyl-3H-chromen-2-one, which demonstrated high affinity for the antiapoptotic proteins Bcl-2 and Survivin in molecular docking analyses. In conclusion, the crude leech saliva was confirmed to possess anti-inflammatory, antioxidant, and anticancer properties. However, further biochemical and clinical research is needed to elucidate the underlying mechanisms of these biological effects in greater detail. Full article

The gastrointestinal microbiota of medicinal leeches is particularly interesting due to their blood-feeding habits, increasing medical use, and risk of pathogen transmission. Three groups of Hirudo verbana were used to study the leech microbiota: farmed leeches fasting for a long time, farmed leeches recently fed with bovine blood, and wild specimens fed with amphibian blood. The microbiota of the leeches’ mouth, pharynx, crop, and intestine was analyzed. Metasequencing analyses were performed using amplification of the 16S rRNA V3-V4 region on a NovaSeq Illumina platform. The relative abundance of bacterial microbiota included environmental bacteria from the families Rhizobiaceae, Comamonadaceae, Sphingobacteriaceae, Phreatobacteraceae, Myxococcaceae, Chitinophagaceae, Rhodospirillaceae, and Bdellovibrionaceae, as well as symbiotic/probiotic bacteria such as Mucinivorans, Aeromonas, Vagococcus, Lactobacillales, and Morganella. Significant differences were found in the different regions of the digestive system among the three groups of leeches, and environmental bacteria were present in all groups to varying degrees. A negative correlation was found between the dominant environmental and the symbiotic/probiotic bacteria. In contrast, a positive correlation was found between environmental and symbiotic/probiotic bacteria, indicating their association with host factors. Microbiota diversity, abundance, and bacterial correlations may be influenced by factors such as the leech’s fasting state, blood meal source, and environmental conditions. The identified opportunistic pathogens, such as Rickettsia, Anaplasma, and Treponema, identified for the first time in H. verbana, should be taken into consideration when using this leech in hirudotherapy. Our results show that extensive screening for opportunistic and pathogenic agents should be performed on leeches intended for medical use. Long-fasting leeches and leeches cultured in specialized farms are recommended for hirudotherapy. Full article

This study presents the first data on the distribution of the Mediterranean medicinal leech Hirudo verbana in Romania, as well as the first molecular characterization of the Romanian populations, using mitochondrial DNA markers. In total, 133 specimens from 19 populations in Romania were surveyed for mitochondrial COI and 12S markers. Available GenBank sequences from other localities throughout Europe were included to place the Romanian populations in a broader phylogeographical context. The Romanian H. verbana samples belong to 23 COI haplotypes, 4 of which are previously known from Serbia and Ukraine, while 19 haplotypes are herein reported for the first time. For 12S, the number of haplotypes identified was 12, from which only 5 are unique to Romania. The Romanian H. verbana cluster within the previously reported eastern phylogroup, known to occur from Serbia to North Caucasus, Turkey, and Uzbekistan. The mismatch distribution analysis and the results of neutrality tests suggest that the Romanian H. verbana currently undergoes a population dispersal. This study also revealed that the lower Danube basin is a hotspot of genetic diversity of H. verbana, where different lineages from the Balkan and Anatolian glacial refugia mixed in the postglacial period. Species Distribution Models (SDMs) indicated that the main ecological variables influencing the current distribution of H. verbana are wetland coverage and elevation. Following the IUCN Red List categories and criteria, the present conservation status of the Mediterranean medicinal leech H. verbana in Romania was assessed as Vulnerable (VU). Our study contributes towards a more refined understanding of the genetic diversity of H. verbana and of the factors that led to its current geographical distribution in Romania and Europe. Full article

The invertebrate leech Hirudo verbana represents a powerful experimental animal model for improving the knowledge about the functional interaction between the extracellular matrix (ECM) and cells within the tissue microenvironment (TME), and the key role played by ECM stiffness during development and growth. Indeed, the medicinal leech is characterized by a simple anatomical organization reproducing many aspects of the basic biological processes of vertebrates and in which a rapid spatiotemporal development is well established and easily assessed. Our results show that ECM structural organization, as well as the amount of fibrillar and non-fibrillar collagen are deeply different from hatching leeches to adult ones. In addition, the changes in ECM remodelling occurring during the different leech developmental stages, leads to a gradient of stiffness regulating both the path of migratory cells and their fates. The ability of cells to perceive and respond to changes in ECM composition and mechanics strictly depend on nuclear or cytoplasmic expression of Yes-Associated Protein 1 (YAP1), a key mediator converting mechanical signals into transcriptional outputs, expression, and activation. Full article

Space and time coherent mapping (STCM) is a technology developed in our laboratory for improved matrix-assisted laser desorption ionization (MALDI) time of flight (TOF) imaging mass spectrometry (IMS). STCM excels in high spatial resolutions, which probe-based scanning methods cannot attain in conventional MALDI IMS. By replacing a scanning probe with a large field laser beam, focusing ion optics, and position-sensitive detectors, STCM tracks the entire flight trajectories of individual ions throughout the ionization process and visualizes the ionization site on the sample surface with a subcellular scale of precision and a substantially short acquisition time. Results obtained in thinly sectioned leech segmental ganglia and epididymis demonstrate that STCM IMS is highly suited for (1) imaging bioactive lipid messengers such as endocannabinoids and the mediators of neuronal activities in situ with spatial resolution sufficient to detail subcellular localization, (2) integrating resultant images in mass spectrometry to optically defined cell anatomy, and (3) assembling a stack of ion maps derived from mass spectra for cluster analysis. We propose that STCM IMS is the choice over a probe-based scanning mass spectrometer for high-resolution single-cell molecular imaging. Full article

Until the beginning of the 21st century, the famous medicinal leech was thought to be represented by only one species, Hirudo medicinalis. However, recent publications have demonstrated that under that name, at least five different species of medicinal leeches were hidden. During the last decade, the biogeography of Western-Palaearctic leeches has begun to unravel, untangling their diversity in practically all of Europe, except for its westernmost peninsula, Iberia. Hirudo medicinalis has been repeatedly reported from Iberia, but those records were considered questionable. We discovered H. verbana in northern Spain, constituting its first record in Iberia. Using an integrative approach (combining morpho-anatomical data and molecular analyses using three genes, COI,12S rRNA, and ITS2), two endemic and geographically separated Iberian lineages have been found. One of them is easily distinguished by its distinctive colour-pattern and is described as H. verbana bilineata ssp. nov. We characterized the new subspecies morphologically, ecologically, and genetically. We also established its phylogenetic relationships with other European Hirudo spp. and confirm the presence of H. troctina in Iberia, occurring as far as 43° lat. N. Iberian H. verbana records constitute its westernmost known distribution to date. The provided distribution pattern of H. verbana contributes to a better understanding of the complexity of Iberia as a glacial refugium/cradle for endemisms, harbouring populations with a high degree of genetic structure that began to settle throughout the Pleistocene. Iberian Hirudo populations are declining in recent decades and there is an urgent need to assess their conservation status and to initiate conservation measures to reverse their decline. Full article

Recent studies performed on the invertebrate model Hirudo verbana (medicinal leech) suggest that the T2 ribonucleic enzyme HvRNASET2 modulates the leech’s innate immune response, promoting microbial agglutination and supporting phagocytic cells recruitment in challenged tissues. Indeed, following injection of both lipoteichoic acid (LTA) and Staphylococcus aureus in the leech body wall, HvRNASET2 is expressed by leech type I granulocytes and induces bacterial aggregation to aid macrophage phagocytosis. Here, we investigate the HvRNASET2 antimicrobial role, in particular assessing the effects on the Gram-negative bacteria Escherichia coli. For this purpose, starting from the three-dimensional molecule reconstruction and in silico analyses, the antibacterial activity was evaluated both in vitro and in vivo. The changes induced in treated bacteria, such as agglutination and alteration in wall integrity, were observed by means of light, transmission and scanning electron microscopy. Moreover, immunogold, AMPs (antimicrobial peptides) and lipopolysaccharide (LPS) binding assays were carried out to evaluate HvRNASET2 interaction with the microbial envelopes and the ensuing ability to affect microbial viability. Finally, in vivo experiments confirmed that HvRNASET2 promotes a more rapid phagocytosis of bacterial aggregates by macrophages, representing a novel molecule for counteracting pathogen infections and developing alternative solutions to improve human health. Full article