Discovery and Mechanism of Action of a Novel Antimicrobial Peptide from an Earthworm.

The robust innate immune system of the earthworm provides a potential source of natural antimicrobial peptides (AMPs). However, the cost and high rediscovery rate of direct separation and purification limits their discovery. Genome sequencing of numerous earthworm species facilitates the discovery of new antimicrobial peptides. Through predicting potential antimicrobial peptides in the open reading frames of the Eisenia andrei genome and sequence optimization, a novel antimicrobial peptide, named EWAMP-R (RIWWSGGWRRWRW), was identified. EWAMP-R demonstrated good activity against various bacteria, including drug-resistant strains. The antibacterial mechanisms of EWAMP-R were explored through molecular simulation and wet-laboratory experiments. These experiments demonstrated that the bacterial membrane may be one of the targets of EWAMP-R but that there may be different interactions with Gram-negative and Gram-positive bacterial membranes. EWAMP-R can disrupt bacterial membrane integrity; however, at low concentrations, it appears that EWAMP-R may get through the membrane of Escherichia coli instead of damaging it directly, implying the existence of a secondary response. Gene expression studies identified that in E. coli, only the apoptosis-like cell death (ALD) pathway was activated, while in Staphylococcus aureus, the MazEF pathway was also upregulated, limiting the influence of the ALD pathway. The different antimicrobial actions against Gram-positive and -negative bacteria can provide important information on the structure-activity relationship of AMPs and facilitate AMP design with higher specificity. This study identified a new source of antibacterial agents that has the potential to address the increasingly serious issue of antibiotic resistance.