J. Hopf, C. Kudary, S. Lee, D. Donahue, V. Ploplis, F.J. Castellino
Berthiaume Institute for Precision Health / University of Notre Dame,
United States
Keywords: ESKAPEE, anti-MDR, anti-AMR, wound healing, anti-inflammatory
Summary:
Statement of Purpose Escalating antimicrobial resistance and the slow pace of new antibiotic discovery demand therapeutic strategies that bypass or supplement conventional antibiotics. We developed phage-architecture–inspired antibacterial nanoparticles (PhANPs)—silica cores decorated with discontinuous shells of silver-alloyed gold nanospheres, with or without cysteine-bearing antimicrobial peptides—to achieve broad-spectrum pathogen clearance and restore antibiotic sensitivity. This work focuses on their antibiofilm and wound-healing performance in infected rat wounds. Methods PhANPs (SiO₂@Au@Ag; 65 nm or 130 nm cores) were synthesized to replicate the spacing of bacteriophage surface spikes, promoting efficient bacterial membrane contact. Peptide-free ANPs and peptide-functionalized pep@ANPs were evaluated in both soluble and immobilized formats. A panel of pathogens representing ESKAPE and implant-associated species—S. aureus (MRSA), P. aeruginosa, E. faecalis, C. striatum, K. pneumoniae, A. baumannii, and S. epidermidis—was tested through CFU reduction, growth-curve assays, and confocal LIVE/DEAD imaging. Cytocompatibility was determined using HaCaT keratinocytes and MG-63 osteoblast-like cells. For in vivo testing, full-thickness wounds in Sprague-Dawley rats were infected with MRSA or carbapenem-resistant A. baumannii and treated topically once daily for 12 days with PhANP@peptide, free peptide, or placebo. Wound closure, bacterial burden, and inflammation were monitored macroscopically and histologically. Results PhANPs displayed potent antibiofilm activity both on material surfaces and within complex wound environments. Immobilized coatings completely prevented bacterial colonization and biofilm maturation on implant-grade steel. In pre-formed biofilms, a single treatment with PhANP@peptides disaggregated biofilm structure and eliminated viable cells, producing sterile surfaces that resisted recolonization. In the rat wound model, PhANP@peptides induced rapid wound stabilization within 24 hours and accelerated closure throughout the 12-day course. Treated wounds remained free of pus or inflammation, while untreated and peptide-only controls exhibited persistent exudate and delayed epithelialization. Quantitative analysis revealed a >99.999% reduction in recoverable bacteria relative to controls, consistent with the disruption of biofilm-embedded populations rather than transient planktonic clearance. Histological examination confirmed restored epidermal integrity, well-organized granulation tissue, and absence of necrosis. The nanoparticles’ silver alloy component contributed to durable local antibacterial protection without exceeding safe silver ion release thresholds. PhANP@peptide also enhanced the performance of sub-MIC antibiotics: co-administration with ertapenem or ceftriaxone produced additive to synergistic suppression of bacterial growth. Repeated bacterial exposure over 1,000 generations failed to generate resistance, underscoring the mechanical and multivalent mode of action that disrupts cell membranes and biofilm scaffolds simultaneously. No cytotoxicity or inflammatory response was observed in keratinocyte or osteoblast assays, indicating compatibility with skin and implant applications. Conclusions Phage-architecture nanoparticles promote both rapid infection control and regenerative wound repair by combining bactericidal, antibiofilm, and pro-healing functions in a single nanostructure. In infected rat wounds, they eradicated multidrug-resistant bacteria, dismantled biofilms, and restored normal healing kinetics with minimal inflammation. Their success in preventing and resolving biofilms positions these materials as next-generation coatings for implants and dressings for acute and chronic wounds. The integration of strong antibiofilm efficacy, wound-healing acceleration, and antibiotic re-sensitization within a biocompatible, antibiotic-free platform supports advancement toward translational testing for trauma, burn, and surgical infection management.