Isolation, characterization, and antibacterial activity of a novel bacteriocin-like substance (PAM3-2) produced by marine-derived Acinetobacter pittii

Abstract Background The emergence and spread of multidrug-resistant bacterial pathogens threaten public health, livestock production and food safety. Marine-derived bacteria are a potentially valuable but underexplored source of antimicrobial substances. This study aimed to isolate and characterize an antibacterial substance from the marine-derived Acinetobacter pittii ( A. pittii ) strain AM3-2, previously reported to display broad antibacterial activity, and to evaluate its antibacterial properties and preliminary mode of action. Results A proteinaceous bacteriocin-like antibacterial substance, designated PAM3-2, was obtained from the cell-free fermentation supernatant of marine-derived A. pittii by acid precipitation, chromatography and activity-guided fractionation. The active fraction showed an apparent molecular-mass about 25 kDa by tricine‑sodium dodecyl sulfate‑polyacrylamide gel electrophoresis. PAM3-2 displayed concentration-dependent inhibitory activity against both Gram-positive and Gram-negative indicator bacteria; the greatest susceptibility was observed for Salmonella enterica subsp. enterica serovar Paratyphi B ( S. Paratyphi B) with minimum inhibitory concentration of 20.45 µg/mL. PAM3-2 exhibited high physicochemical stability, retaining more than 70% of its antibacterial activity after treatment at 121 °C for 20 min, maintaining substantial activity over a broad pH range (pH 2–8), and showing considerable resistance to proteolytic enzymes. Scanning electron microscopy and leakage assays were consistent with loss of membrane integrity, accompanied by increased extracellular potassium, lactate dehydrogenase, nucleic acids and proteins. Conclusions PAM3-2 is a proteinaceous bacteriocin-like antibacterial substance produced by marine-derived A. pittii with high physicochemical stability. PAM3-2 exhibits broad-spectrum antibacterial activity, particularly against S . Paratyphi B, and its antibacterial effect is associated with disruption of membrane integrity.