In a previous work, we performed de novo RNA sequencing of Allomyrina dichotoma using next generation sequencing and identified several antimicrobial peptide candidates based on transcriptome analysis. Among them, a cationic antimicrobial peptide, allomyrinasin, was selected bioinformatically based on its physicochemical properties. Here, we assessed the antimicrobial and anti-inflammatory activities of allomyrinasin against microorganisms and mouse macrophage Raw264.7 cells. Allomyrinasin showed antimicrobial activities against various microbes and decreased the nitric oxide production of the lipopolysaccharide-induced Raw264.7 cells. Furthermore, quantitative RT-PCR and ELISA revealed that allomyrinasin reduced cytokine expression levels in the Raw264.7 cells. We also identified inducible nitric oxide synthase, cyclooxygenase-2 expression, and PGE 2 production through western blot analysis and ELISA. We confirmed that allomyrinasin bound to bacterial cell membranes via a specific interaction with lipopolysaccharides. Taken together, these data indicate that allomyrinasin has antimicrobial and anti-inflammatory activities as exemplified in lipopolysaccharide-induced Raw264.7 cells. We have provided a potentially useful antimicrobial peptide candidate that has both antimicrobial and anti-inflammatory activities.
We compared the mRNA expression profile of the Harmonia axyridis larvae that were either untreated or treated with LPS. The extracted mRNAs were subjected to ACP RT-PCR analysis using a combination of arbitrary primers and oligo (dT) primer. Among the 47 DEGs differentially expressed, we identified a cDNA showing homology with defensin-like antibacterial peptide. The cDNA showed a putative 32-residue signal sequence and a 50-residue mature peptide named harmoniasin. We also investigated the antibacterial activity of the harmoniasin analog, which exhibited potent antibacterial activities against Gramnegative and-positive bacteria strains and it also evidenced no hemolytic activity.
The centipede Scolopendra subpinipes mutilans is a medicinally important arthropod species. However, its transcriptome is not currently available and transcriptome analysis would be useful in providing insight into a molecular level approach. Hence, we performed de novo RNA sequencing of S. subpinipes mutilans using next-generation sequencing. We generated a novel peptide (scolopendrasin II) based on a SVM algorithm, and biochemically evaluated the in vitro antimicrobial activity of scolopendrasin II against various microbes. Scolopendrasin II showed antibacterial activities against gram-positive and -negative bacterial strains, including the yeast Candida albicans and antibiotic-resistant gram-negative bacteria, as determined by a radial diffusion assay and colony count assay without hemolytic activity. In addition, we confirmed that scolopendrasin II bound to the surface of bacteria through a specific interaction with lipoteichoic acid and a lipopolysaccharide, which was one of the bacterial cell-wall components. In conclusion, our results suggest that scolopendrasin II may be useful for developing peptide antibiotics.
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