Molecular based identification and phylogenetic relationship of the leech Hirudinaria manillensis from India by using mitochondrial cytochrome c oxidase subunit I gene.

BACKGROUND: A molecular approach for the identification of unknown species by the using mitochondrial cox1 gene is an effective and reliable as compared with morphological-based identification. Hirudinaria manillensis referred to as Asian Buffalo Leech, is found in South Asia and traditionally used as medicine owing to its medicinal properties. METHODS AND RESULTS: The study aimed to isolate and identify the leech species using cox1 gene sequencing and their phylogenetic relationships. The nucleotide sequences of cytochrome c oxidase subunit I (cox1) mitochondrial genes were analyzed for species identification and the phylogenetic relationship of crucial therapeutic leech Hirudinaria manillensis. The isolated DNA from the leech sample was amplified with cox1 gene-specific primers. BLAST results with the H. manillensis sequence showed 89.24% homology with H. manillensis and phylogenetic tree analysis revealed the genetic relationship with other GenBank submitted sequences. CONCLUSION: The present study concluded that the cox1 gene could be an effective way to identify the leech H. manillensis and provided sufficient phylogenetic information to distinguish H. manillensis indicating a significant mtDNA-based approach to species identification.

In vitro anticancer activity of Hirudinaria manillensis methanolic extract and its validation using in silico molecular docking approach.

Cancer has emerged as a potentially lethal illness, which recently upsurged in the mortality rate. Animal-derived compounds could be promising targets with higher efficacy and low toxicity in anticancer therapy. The present study aimed to explore the presence of anticancer potential compounds in Hirudinaria manillensis methanolic extract and their anticancer potential against various cancer cell types and target identification by Auto dock method. Initially, the identification of bioactive compounds was achieved by GC-MS analysis followed by the anticancer activity by MTT assay against A549, HeLa, MDA-MB-231, MG-63, and MOLT-4. Further, the effect of a lead compound on the cancer cell target was analyzed by the Auto dock method. GC-MS analysis results revealed the presence of 25 different bioactive compounds including anticancer potential compounds, such as Lupeol, Carvacrol, and Demecolcine. Interestingly, MTT assay results demonstrated the anticancer potential of Hirudinaria manillensis extract (LE) against various cancer cell lines, such as A549 (54.60 µg/ml), HeLa (19.93 µg/ml), MDA-MB-231 (20.23 µg/ml), MG-63 (20.04 µg/ml), and MOLT-4 (171.8 µg/ml), respectively. Among these cell types, the maximum inhibition was observed against HeLa with the IC50 concentration of 19.93 µg/ml. Furthermore, Demecolcine compound was docked with the EGFR tyrosine kinase showed the binding affinity of the docked complex was predicted to be - 6.2 kcal/mol. Thus, we conclude that H. manillensis has a significant anticancer effect on human cancer cell lines and could be used as a natural target which paves the way for further studies on biomedical applications in cancer therapeutics.