PROJECT SUMMARY/ABSTRACT Periprosthetic infections (PPIs) occur in 1-4% of primary total joint replacement and up to 30% of revision arthroplasty and are difficult to prevent or treat due to the tendency of bacteria, especially Staphylococcus aureus (S. aureus), to colonize and form biofilms on implant surfaces and to invade the surrounding bone matrices. Conventional antibiotic prophylaxis and treatments do not adequately address this challenge. We recently showed that anti-fouling zwitterionic polymers grafted from Ti6Al4V intramedullary (IM) implants, when combined with a single systemic injection of vancomycin, effectively inhibited S. aureus colonization on implant surfaces and PPI in murine femoral canals, significantly outperforming either treatment alone. Here, we aim to facilitate the clinical translation of this promising synergistic anti-PPI strategy by engineering a robust anti- fouling block copolymer dip-coating as an off-the-shelf product for metallic implants, and an effective and safe synergistic antibiotic prophylaxis regimen for sustained protection during primary implant insertion and implant revision, respectively, against methicillin-sensitive and methicillin-resistant (MRSA) S. aureus PPIs. In Aim 1, high-molecular weight polymethacrylate block copolymers with anti-fouling zwitterionic sidechains and metal surface-binding sidechains are prepared by sequential Reverse Addition Fragmentation Chain Transfer (RAFT) polymerization and copper-catalyzed azide-alkyne cycloaddition (CuAAC). Taking advantage of the excellent control over the degree of polymerization by RAFT and the high-fidelity CuAAC modification of sidechains, the respective block lengths and chemical nature and spatial density of metal alloy surface-binding residues are modularly altered to identify an optimal copolymer composition and dip-coating protocol to achieve consistent and stable anti-fouling coating on Ti6Al4V that can sustain flow wash, sterilization and over-the-shelf storage. Coated surfaces are characterized by water contact angels, x-ray photoelectron spectroscopy, and resistance to non-specific protein adsorption and bacterial colonizations in vitro. In Aims 2 and 3, the top anti-fouling dip- coating chosen in Aim 1 is applied to Ti6Al4V IM pins and examined, along with synergistic antibiotic prophylaxis, for the efficacy and safety in achieving long-term inhibition of PPI and recurrent PPI caused by methicillin-sensitive S. aureus or MRSA following the insertion of primary and revision implants in rats, respectively. The degree of PPI or recurrent PPI as a function of pin coating and the timing/frequency of synergistic antibiotic prophylaxis are longitudinally monitored by complete blood counts and µCT quantification of cortical bone thickening, and by endpoint quantification of bacteria on the retrieved pin, torsion test and histology/electron microscopy characterizations of explanted femurs over the course of 6 months. Long-term safety of the co...