Differential anti-microbial properties between the burying beetle Nicrophorus quadripunctatus and the short sexton beetle Necrodes littoralis.

To identify the differential properties of anti-microbial peptides (AMPs) and proteins in the burying beetle Nicrophorus quadripunctatus and the short sexton beetle Necrodes littoralis, we performed comparative transcriptomic analyses, quantified gene expression levels, examined structural variations, and evaluated anti-microbial and hemolytic activities of representative AMPs, including cecropin B and defensin. Two AMPs (cecropin B and defensin) and four anti-microbial proteins (cathelicidin, lysozyme, major royal jelly protein, and peptidoglycan recognition proteins) were predominantly detected. Defensin was the most highly expressed AMP in both species, whereas cecropin B showed relatively lower expression. Cecropin B and defensin in N. quadripunctatus exhibited 52.2- and 100.9-fold higher relative transcription levels, respectively, compared with N. littoralis, representing the two largest expression differences among the examined genes. Notably, substantial sequence variation was observed in the mature peptide regions of both AMPs between the two species. The α-helices of both AMPs from N. quadripunctatus exhibited longer segments than those from N. littoralis as a structural feature. Defensin of N. littoralis exhibited significantly higher anti-microbial activity against Gram-positive bacteria compared with that of N. quadripunctatus. In addition, only cecropin B exhibited anti-microbial activity against Gram-negative bacteria, and neither of the two AMPs showed detectable hemolytic activity. Overall, this study provides the first comparative transcriptomic, structural, and functional characterization of AMPs and anti-microbial proteins in N. quadripunctatus and N. littoralis, offering important foundational insights for the future development of bacteria-specific AMPs as alternatives to conventional antibiotics.