A recombinant fungal defensin-like peptide-P2 combats multidrug-resistant Staphylococcus aureus and biofilms

Area of Science:

• Microbiology
• Drug Discovery
• Immunology

Background:

• Methicillin-resistant Staphylococcus aureus (MRSA) poses a significant threat due to antibiotic resistance, intracellular persistence, biofilm formation, and persister cells.
• Fungal defensin-like peptides (DLPs) are a promising class of antimicrobial agents with potent antibacterial properties.

Purpose of the Study:

• To identify and characterize novel fungal DLPs for their potential as antibacterial agents against multidrug-resistant (MDR) Staphylococcus aureus.
• To evaluate the efficacy of the most promising DLP candidate (P2) against MDR S. aureus, including its activity against intracellular bacteria, biofilms, and persister cells, as well as its immunomodulatory effects and in vivo therapeutic potential.

Main Methods:

• Nine novel fungal DLPs were identified and expressed in Pichia pastoris.
• Antibacterial, anti-biofilm, and anti-persister activities of the DLPs were tested against MDR S. aureus.
• In vitro assays assessed P2's effects on intracellular bacteria in macrophages and its mechanism of action.
• In vivo studies evaluated P2's therapeutic efficacy, immunomodulatory effects, and impact on bacterial translocation and organ injury in infected mice.

Main Results:

• Fungal DLP P2 demonstrated high activity (MIC < 2 μg/mL), low toxicity, no observed resistance, and high stability.
• P2 effectively reduced intracellular MDR S. aureus in macrophages (80-97%) by disrupting bacterial DNA and membranes.
• P2 inhibited/eradicated biofilms, killed 99% of vancomycin-resistant persister bacteria, and modulated inflammatory responses.
• In vivo, P2 (5 mg/kg) achieved 100% survival in infected mice, outperforming vancomycin (30 mg/kg), and alleviated organ damage.

Conclusions:

• The novel fungal DLP P2 exhibits broad-spectrum antibacterial activity, potent anti-biofilm and anti-persister effects, and favorable immunomodulatory properties.
• P2 demonstrates significant therapeutic potential against MDR Staphylococcus aureus infections, including intracellular and biofilm-associated forms.
• P2 represents a promising candidate for the development of novel antimicrobial agents to combat challenging staphylococcal infections.