From decay to discovery: A new antimicrobial peptide from Chrysomya megacephala (Diptera: Calliphoridae) larvae.

Abstract

The larvae of Chrysomya megacephala (Diptera: Calliphoridae) thrive in environments rich in decaying organic matter and dead animals, which are often colonized by bacteria. This adaptation suggests the larvae possess antimicrobial substances that provide them with a defense against bacterial infection. The hemolymph from around 200 C. megacephala larvae previously exposed to methicillin-resistant Staphylococcus aureus (MRSA) (clinical isolate) was collected, purified, and isolated utilizing Reverse Phase High Performance Liquid Chromatography (RP-HPLC). Fractions exhibiting antimicrobial properties underwent further analysis using Quadrupole Time-of-Flight Liquid Chromatography Mass Spectrometry (QTOF-LCMS). The resulting mass spectra were translated into amino acid sequences through de novo algorithms. Antimicrobial prediction tools from repositories such as APD3, Database of Antimicrobial Activity and Structure of Peptides (DBSAAP), Database of Antimicrobial Peptide (DBAMP), and Collection of Antimicrobial Peptide Version 4 (CAMPR4) were employed to validate the peptide sequences. We found the peptide (HGCGRLSKWFRQPGLLLSVKR) exhibited partial similarity with C. megacephala's heat shock protein 70 (hsp70). The peptide demonstrated activity against Staphylococcus aureus, including MRSA, Micrococcus luteus, Staphylococcus epidermidis, and Bacillus subtilis with a minimum inhibitory concentration (MIC) of 0.06 mg/ml and an inhibition zone ranging from 8 to 11 mm at a concentration of 1 mg/ml. The cytotoxicity test of the peptide on immortalized keratinocyte cells (HaCat) and human corneal epithelial cells (HCEC) revealed an average cell viability > 80% at concentration of 0.06 mg/ml. In conclusion, the novel peptide (HGCGRLSKWFRQPGLLLSVKR) exhibits a broad spectrum of activity against Gram-positive bacteria while demonstrating minimal toxicity towards human cells.