A truncated Sph12-38 with potent antimicrobial activity showing resistance against bacterial challenge in Oryzias melastigma

Antimicrobial peptides (AMPs) represent an efficient part of innate immunity and are found in a variety of life. Among them Histone 2A (H2A), as a promising class of AMPs, attracts great attention, but the in vivo mechanism of H2A derived AMP is still less known. Based on the acquisition of Sphistin, a synthetic 38-amino acid H2A derived peptide from Scylla paramamosain, as reported in our previous study, was truncated into three short fragments (Sph12-38, Sph20-38 and Sph30-38) and further investigated for its possible functional domains. The antimicrobial activities of these analogs against different Gram-positive bacteria, Gram-negative bacteria and fungi were illustrated. Among the analogs, Sph12-38 showed a stronger activity with a much lower minimum inhibitory concentration (3 μM) against Staphylococcus aureus, Corynebacterium glutamicum, Micrococcus lysodeikticus Fleming, Bacillus subtilis, Pseudomonas fluorescens, Aeromonas hydrophila and A. sobria in comparison with the reported Sphistin. A leakage of intracellular content was described in E. coli treated with Sph12-38. Unlike Sphistin which mainly disrupts the membrane integrity, Sph12-38 could also combine the A. sobria genomic DNA with a minimum concentration of 6 μM and was located intracellularly in cells observed under confocal laser scanning microscope imaging. In comparison with the control group of Oryzias melastigma injected with A. sobria alone, the group treated with a mixture of Sph12-38 and A. sobria showed a higher survival rate 7 days post-injection. Furthermore, in a pretreatment assay at 6 h, a higher survival rate was observed in the group injected with the mixture of Sph12-38 and A. sobria. Taken together, the synthetic peptide of Sph12-38 had a potent antimicrobial activity against bacteria. However, Sph12-38 had no cytotoxicity towards the hemolymph of S. paramamosain. Our study suggested that, as with Sph12-38, the H2A derived peptides were more likely prone to exert their activities in vivo through the truncated fragments while defending against different species of pathogens.