Abstract
Background: Cationic antimicrobial peptides (AMPs) have attracted considerable attention to source antimicrobial as well as antibiofilm technology solutions. In vitro and in vivo antimicrobial efficacies of three short-chain AMPs, Indolidicin, Cecropin A (1–7) – Melittin (CAMA), lactoferricin (17–30), and their combination were explored for the first time against multi-drug resistant enteroaggregative Escherichia coli (MDR-EAEC). Methods and materials: Minimum Inhibitory Concentrations (MICs) and Minimum Bactericidal Concentrations (MBCs) of AMPs were determined by broth micro-dilution technique. The AMPs were tested for its stability at temperatures (70 °C and 90 °C), proteases (Proteinase-K and trypsin) and physiological concentration of cationic salts (NaCl and MgCl2); safety in sheep RBCs and cell lines (RAW 264.7 and HEp-2); adverse effect on beneficial flora and membrane permeability. In vitro dose- and time-dependent time kill assay was also carried out. Later, in vivo studies of AMPs (alone and in combination) employing Galleria mellonella (survival assay, bacterial burden, haemocyte density, melanisation assay, cytotoxicity assay and histopathological examination) was carried out. Results: All the three AMPs were stable (except for Indolicidin with trypsin), tested safe at lower MIC and no adverse effect against beneficial flora was observed. AMPs alone and in combination, exhibited bacterial membrane permeability. In vitro time-kill assay revealed concentration-cum-time dependent clearance of MDR-EAEC in the AMP-treated groups, while, in the in vivo G. mellonella experiment, the infected group treated with AMPs (alone and in combination) revealed an increased survival rate, immunomodulatory effect, reduced MDR-EAEC counts and were tested safe to the larval cells which concurred with histopathological observations. Conclusion: The AMPs seem to be promising candidates for further investigations using appropriate animal models for MDR-EAEC strains.
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