We have analyzed the transcriptome of Scolopendra subspinipes mutilans using RNA sequencing and identified severalantimicrobial peptide candidates. Among the peptides, named scolopendrasins, were selected based on the physicochemicalproperties of antimicrobial peptides via an in silico analysis. As a result, we evaluated the antimicrobial activities ofscolopendrasins against Gram positive and negative bacteria including Candida albicans by radial diffusion assay and colonycount assay. We also investigated the cytotoxicity of scolopendrasins through hemolysis assay. We found that the actionof scolopendrasins involved binding to the surface of microorganisms via a specific interaction with lipopolysaccharides,lipoteichoic acid, and peptidoglycans, which are components of the bacterial membrane. These results will provide a basisfor developing therapeutic agents such as peptide antibiotics.