Karau, Melissa J.

Introduction Infection following extensor mechanism reconstruction with Marlex mesh is a devastating and limb threatening complication. The purpose of this study was to evaluate whether vancomycin-loaded mesh reduced staphylococcal infection in a murine extensor mechanism reconstruction model with Marlex mesh. Methods Mouse-sized, eight-layer Marlex mesh constructs were fabricated to simulate clinical reconstruction and soaked in a vancomycin solution (50 mg/mL) for in vitro and in vivo testing. Three methicillin-resistant clinical isolates from human periprosthetic knee infections were evaluated: Staphylococcus aureus, Staphylococcus epidermidis , and Staphylococcus lugdunensis. In vitro testing assessed vancomycin loading, mechanical properties, antibacterial activity, and early biofilm inhibition. For in vivo experiments, plain or vancomycin-loaded Marlex meshes were implanted into the murine extensor mechanism and inoculated intraoperatively (5 per group per isolate). Meshes and surrounding tissues were harvested on postoperative day 9 for quantitative culture. Results For all three isolates, vancomycin-loaded Marlex mesh exhibited sustained antibacterial activity for up to 3 days in vitro based on zone of inhibition testing (days 1–3, p < 0.0001 for each isolate). In vitro testing of biofilm inhibition demonstrated significant reduction in biofilm-associated colony-forming units at 6 hours after inoculation and remained low thereafter (p<0.05 for each isolate). In vivo , vancomycin-loaded Marlex mesh reduced bacterial burden on meshes and surrounding tissues by > 3 log₁₀ compared with plain mesh for all three isolates (p<0.01 for both mesh and tissue for all isolates). Conclusion Vancomycin-loaded Marlex mesh provided sustained local antibacterial activity and significantly reduced staphylococcal infection without compromising mechanical properties in a murine extensor mechanism reconstruction model. This approach may represent a promising adjunct to reduce infection risk in extensor mechanism reconstructions with Marlex mesh after total knee arthroplasty.

Background: Infection following extensor mechanism reconstruction (EMR) with Marlex mesh (C.R. Bard) is devastating and limb-threatening. The purpose of this study was to evaluate whether vancomycin-loaded mesh reduces staphylococcal infection in a murine EMR model. Methods: Mouse-sized, 8-layer mesh constructs were fabricated to simulate clinical reconstruction and soaked in a vancomycin solution (50 mg/mL) for in vitro and in vivo testing. Three methicillin-resistant clinical isolates from human periprosthetic Staphylococcus aureus, Staphylococcus epidermidis, and Staphylococcus lugdunensis knee infections were evaluated. In vitro testing assessed vancomycin loading capacity, mechanical properties, antibacterial activity, and inhibition of biofilm formation. For in vivo experiments, plain or vancomycin-loaded mesh was implanted into C57BL/6 mouse extensor mechanisms, which were intraoperatively contaminated with 1 of the 3 study isolates (5 per group). Mesh and surrounding tissue were harvested for quantitative culture on postoperative day 9. Results: For all 3 isolates, vancomycin-loaded mesh produced zones of inhibition and exhibited antibacterial activity through at least 3 days in vitro (p < 0.0001). Mechanical testing revealed that loading mesh with vancomycin did not negatively impact tensile strength, hysteresis, or suture pull-through properties. In vitro testing of inhibition of biofilm formation demonstrated reduced colony-forming units 6 hours after inoculation, which remained low throughout the 9-day experimental period (p < 0.05 for all isolates). In vivo, vancomycin-loaded mesh reduced bacterial burden on mesh and in surrounding tissue by >3 log 10 compared with plain mesh for all 3 isolates (p < 0.01 for both mesh and tissue). Conclusions: Vancomycin-loaded mesh provided in vitro antibacterial activity without compromising mesh mechanical properties and reduced staphylococcal infection in a murine EMR model. Further work is needed to evaluate translational potential and safety before clinical application. Clinical Relevance: Mesh-associated infection after EMR is devastating and difficult to treat. These preclinical findings suggest that local vancomycin delivery may serve as an adjunct to systemic prophylaxis, warranting further study before clinical use.

Aims Extensor mechanism disruption is a major complication following total knee arthroplasty, with treatment often consisting of extensor mechanism reconstruction utilizing monofilament polypropylene mesh (Marlex mesh, C. R. Bard, Inc). Infection following this procedure is devastating, often resulting in revision surgery and mesh removal, with permanent limb dysfunction. This study evaluated the in vitro ability of vancomycin-loaded polypropylene mesh to inhibit growth of isolates of three Staphylococcus species from patients with periprosthetic joint infection. Methods The mesh was prepared in two configurations – single-layer and eight-layer – and soaked for ten minutes in either sterile water (control) or vancomycin solution (50 mg/ml), after which the mesh was placed on tryptic soy agar plates inoculated with staphylococci and incubated at 37°C for 18 to 24 hours. Zones of inhibition were measured daily over seven days, with mesh transferred to freshly inoculated plates daily. Testing was performed in triplicate, with experiments repeated three times for each bacterial isolate. A total of 30 clinical isolates were tested (five methicillin-susceptible Staphylococcus epidermidis , five methicillin-resistant S. epidermidis , ten methicillin-resistant Staphylococcus aureus , and ten Staphylococcus lugdunensis ). Results Across all species, the eight-layer vancomycin-loaded mesh produced statistically significantly larger median zones of inhibition than the single-layer mesh. The antibacterial activity of the eight-layer mesh persisted through to day 7, while the single-layer mesh showed reduced activity by days 4 to 5. Conclusion Compared with single-layer vancomycin-loaded mesh, multilayer vancomycin-loaded mesh, which mimics clinical application, provides enhanced and sustained antibacterial activity in vitro. Cite this article: Bone Joint Res 2026;15(8):998–1004.

Abstract A low-cost, easy-to-adopt noninvasive imaging modality for differentiating bacterial infection from inflammation is desirable to accurately diagnose bacterial infections. Herein, a bacteria-specific PET probe, [68Ga]Ga-BP, was developed and evaluated in rat models of foreign body-associated osteomyelitis induced by methicillin-resistant Staphylococcus aureus, Pseudomonas aeruginosa, and Klebsiella pneumoniae and was directly compared with sterile inflammation and uninstrumented uninfected controls. [68Ga]Ga-BP-based imaging of MRSA-infected tibiae was also compared to [68Ga]Ga-citrate and [18F]FDG. After iron depletion, dynamic PET/CT imaging was performed at 15–120 min postinjection of radiotracers, followed by assessment of ex vivo biodistribution. [68Ga]Ga-BP successfully detected tibial infection and demonstrated &gt;6-fold higher uptake in infected tibiae with all three bacterial species versus contralateral tibiae via PET imaging. [68Ga]Ga-BP outperformed [68Ga]Ga-citrate and [18F]FDG in identifying MRSA infection. [68Ga]Ga-BP represents a promising, noninvasive PET imaging agent for direct bacterial detection and bacterial infection treatment monitoring.

Abstract Background Phage therapy has emerged as a generally safe approach, as demonstrated in preclinical and clinical studies. However, data on efficacy, including optimal dosing frequency, remain limited. MRSE is a leading cause of foreign body-associated osteomyelitis and a potential target for phage therapy. Here, the activity of phage K alone and in combination with vancomycin (VAN), administered once (QD) or three times (TID) daily, against biofilms formed by two orthopedic infection-associated MRSE isolates on orthopedic screws was evaluated.Fig 1.in vitro multi-dose assay of methicillin-resistant S. epidermidis biofilm-coated cortical bone screws.IDRL, Infectious Diseases Research Laboratory; DMEM, Dulbecco's Modified Eagle Medium; PBS, phosphate-buffered saline; CFU, colony-forming units. Created in https://BioRender.comFig 2.Quantities of biofilm on screws after 24 hours of treatment.*Log10 CFU/cm² of biofilm on screws before treatments. Biofilm burden of the MRSE isolates prior to treatment ranged from 5.7 to 6.3 log10 CFU/cm². A Kruskal-Wallis test was performed to evaluate the statistical significance of biofilm reduction between treatment groups. The minimum inhibitory concentration (MIC), minimum biofilm inhibitory concentration (MBIC), and minimum biofilm bactericidal concentration (MBBC) of vancomycin for both IDRL-9410 and IDRL-9403 were 2, 2, and &gt;128 µg/mL, respectively. Methods MRSE IDRL-9410 and IDRL-9403 biofilms were formed on titanium cortical bone screws (D 1.5 mm, L 4 mm, surface area 0.33 cm²) in 0.2 mL of tryptic soy broth containing 10⁶ CFU/mL (Fig 1); testing was performed in triplicate on two separate days. After 24 hours, screws were rinsed with PBS and randomly assigned to: Dulbecco’s Modified Eagle Medium (DMEM); vancomycin (VAN, 20 μg/mL) in DMEM; phage K (10⁹ PFU/mL) in DMEM; or VAN and phage K in DMEM. Every 8 hours, screws from the TID groups were transferred to new plates containing fresh treatment, with QD groups left in the original material. 24 hours post-treatment, screws were quantitatively cultured. Bactericidal activity was defined as ≥ 3 log₁₀ CFU/cm² reduction compared to the DMEM group, and synergy as ≥ 2 log₁₀ CFU/cm² reduction in the combination group compared to VAN or phage alone. Results After 24 hours, bactericidal activity was observed for QD and TID VAN-phage K groups against IDRL-9410 (p &lt; 0.05 and p &lt; 0.01, respectively) and IDRL-9403 (both p &lt; 0.01). Synergy was observed in the VAN-phage K QD group, with a 2.2 log₁₀ CFU/cm² reduction for IDRL-9410 compared to VAN alone (Fig 2). 3.0 and 5.3 log₁₀ CFU/cm² reductions were observed in the VAN TID groups for IDRL-9410 and IDRL-9403, respectively, compared to the DMEM group (both p &lt; 0.01). There was no statistically significant difference in biofilm reduction between the QD and TID groups for phage K alone or VAN-phage K for either strain. Conclusion Combination treatment with phage K and VAN, administered QD or TID, showed greater biofilm reduction compared to phage K alone. Increasing phage dosing frequency did not enhance biofilm reduction under the conditions studied. Disclosures Robin Patel, MD, a patent on Bordetella pertussis/parapertussis PCR issued, a patent on a device/method for sonication, a patent on PET imaging of bacterial infection: a patent on Bordetella pertussis/parapertussis PCR issued, a patent on a device/method for sonication, a patent on PET imaging of bacterial infection|MicuRx Pharmaceuticals and bioMérieux: Grant/Research Support|PhAST, Day Zero Diagnostics, DEEPULL DIAGNOSTICS, S.L., Nostics, HealthTrackRx, bioMérieux and CARB-X: Advisor/Consultant|Up-to-Date and the Infectious Diseases Board Review Course: Honoraria

Abstract Background Staphylococci are the most frequent microorganisms in periprosthetic joint infection (PJI) and treatment remains challenging. Phage is being considered as a potential treatment. Here, the in vitro activity of phages SaMD07FSphi1or SaNSI1469phi1, alone and in combination with vancomycin against staphylococcal biofilms was evaluated. Figure 1 In vitro activity of phage SaMD07FSphi1 against S. aureus and S. epidermidis biofilms Figure 2 In vitro activity of phage SaNSI1469phi1 against S. aureus and S. epidermidis biofilms Methods Biofilms were grown at 37°C on 4 cm stainless steel K-wires in tryptic soy broth (TSB) with 106 cfu/ml of S. epidermidis or S. aureus (8 PJI isolates with phage activity against planktonically-grown bacteria). After 24 h, 3 K-wires were quantitatively cultured to define pre-treatment bacterial loads, with the remaining assigned to one of four treatment groups: TSB, phage (4 x 106 plaque forming units/ml), vancomycin (16 µg/ml) or phage/vancomycin. K-wires were incubated for 6 or 24 h at 37°C, rinsed, sonicated, and quantitatively cultured. Testing was performed in triplicate and on 3 separate days and mean log10 cfu/K-wire reduction factors (LRFs) versus TSB calculated. Results Phage SaMD07FSphi1 (Figure 1) plus vancomycin showed LRFs of 2.8 (p, 0.0003) and 2.3 (p, 0.0177) log10 cfu/K-wire at 24 h for S. aureus IDRL-6121 and S. aureus IDRL-8454, respectively, whereas phage alone showed was 3.1 (p, 0.0003) and 3.0 (p, 0.0003), respectively. All treatments were ≤1.0 log10 cfu/K-wire 24 h LRFs for S. aureus IDRL-6175, S. epidermidis IDRL-5966 and S. epidermidis IDRL-6108.This phage plus vancomycin showed LRFs of 0.6 log10 cfu/K-wire at 6 h for S. epidermidis IDRL-5966 and S. aureus IDRL-6175, with bacterial regrowth at 24 h. Phage SaNSI1469phi1 (Figure 2) plus vancomycin showed 24 h LRFs of 2.0 (p, 0.02) and 3.2 (p, 0.003) log10 cfu/K-wire for S. epidermidis IDRL-6955 and S. aureus IDRL-7299, respectively, whereas phage alone was 0.2 (p, 0.17) and 0.5 (p, 0.03) log10 cfu/K-wire, respectively. All treatments showed ≤1.0 log10 cfu/K-wire 24-hour LRFs log10 cfu/K-wire for S. epidermidis IDRL-10273. This phage alone showed 6 h LRFs of 1.0 and 1.3 for S. epidermidis IDRL-6955 and S. aureus IDRL-7299, with bacterial regrowth at 24 h. Conclusion Phages SaMD07FSphi1 and SaNSI1469phi1 alone and with vancomycin showed activity against staphylococcal biofilms formed by 4 isolates. Increased activity was observed at 6 h in some cases, with bacterial regrowth at 24 h. Disclosures Robin Patel, MD, a patent on Bordetella pertussis/parapertussis PCR issued, a patent on a device/method for sonication, a patent on PET imaging of bacterial infection: a patent on Bordetella pertussis/parapertussis PCR issued, a patent on a device/method for sonication, a patent on PET imaging of bacterial infection|MicuRx Pharmaceuticals and bioMérieux: Grant/Research Support|PhAST, Day Zero Diagnostics, DEEPULL DIAGNOSTICS, S.L., Nostics, HealthTrackRx, bioMérieux and CARB-X: Advisor/Consultant|Up-to-Date and the Infectious Diseases Board Review Course: Honoraria

ABSTRACT Biofilms hinder wound infection healing, making wound infections a healthcare challenge for which innovative treatments are needed. A 1.77 cm 2 electrochemical bandage (e-bandage), which generates the biocide hypochlorous acid (HOCl), has shown promising anti-biofilm activity in vitro , ex vivo , and in vivo . The e-bandage is a three-electrode system operationalized by a hydrogel. In addition to providing a conductive electrolyte for the electrodes, the hydrogel provides a protective barrier for the wound. How different hydrogels impact e-bandage functionality is not completely defined. Here, the in vitro activity of seven clinically available hydrogels (xanthan gum, 3M, Duoderm, Prontosan, Purilon, Skintegrity, and Solosite) was evaluated with 1.77 cm 2 HOCl-generating e-bandages using a potentiostat with a multiplexer in a membrane biofilm model against methicillin-resistant Staphylococcus aureus IDRL-6169 and Acinetobacter baumannii ATCC-17978. When applied with polarized HOCl-producing e-bandages, all evaluated hydrogels except Solosite yielded ≥5.5 log 10 CFU/cm 2 reductions in bacterial quantities after 6 h of treatment, with most being below the detection limit at that time point. Prontosan exhibited antibacterial activity independent of e-bandage. Results of this study inform the selection of clinically available hydrogels that may be suitable for use with a 1.77 cm 2 HOCl-producing e-bandage. IMPORTANCE In this study, it was shown that some clinically available hydrogels, combined with a 1.77 cm 2 HOCl-generating electrochemical bandage, deliver time-dependent antimicrobial activity against methicillin-resistant Staphylococcus aureus and Acinetobacter baumannii biofilms, offering a potential approach to treat wound infections, including those caused by antimicrobial-resistant bacteria.

ABSTRACT Periprosthetic joint infection (PJI) is a serious complication of total joint arthroplasty (TJA), with Staphylococcus aureus being one of the most common causative organisms. Irrigation and debridement with component retention (IDCR) is often performed to manage PJI. Antimicrobial treatment that targets S. aureus biofilms on prosthesis surfaces and within bone may be especially helpful when retaining a prosthesis. Phages, viruses that specifically infect bacteria, are a potential therapeutic consideration. Using a biofilm time kill assay in bovine synovial fluid, bactericidal activity of vancomycin or phage K (32 µg/mL and 10 11 pfu/mL, respectively) as well as the combination of phage K (10 10 pfu/mL) and vancomycin (32 µg/mL) was observed against a methicillin‐resistant S. aureus (MRSA) strain. Using a murine PJI model, mice infected with MRSA underwent IDCR followed by phage administration into the joint space with or without systemically administered vancomycin. Mice receiving both phage and vancomycin had significant reductions in MRSA on the prosthesis surface compared to untreated or vancomycin‐treated mice. Phage K showed bactericidal activity in vitro and was active when locally delivered in combination with systemically administered vancomycin in a murine PJI model managed with IDCR.

Background/Objectives: Staphylococci are the most common causes of periprosthetic joint infection (PJI); new antimicrobials are needed to manage these difficult infections. Nisin A is a lantibiotic peptide derived from Lactococcus lactis that has antimicrobial activity against Gram-positive bacteria, including staphylococci, and is an FDA-approved preservative used in the food and dairy industry. Here, the in vitro nisin A susceptibility of PJI-associated staphylococci was assessed. Methods: The minimum inhibitory concentrations (MICs), minimum biofilm inhibitory concentrations (MBICs), and minimum biofilm bactericidal concentrations (MBBCs) of nisin A were measured by broth microdilution against 106 staphylococcus isolates isolated from PJI. MICs were assessed using 5 × 105 CFU/mL plus nisin A. For MBICs, biofilms were grown on pegged lids for 6 h, followed by 20 h of treatment. For MBBCs, pegged lids were transferred to plates containing media only for 20 h. The results were determined as the lowest concentrations with no visual growth. Two-dimensional MICs with nisin A and vancomycin were assessed for 20 isolates. Fractional inhibitory concentrations (FICs) were calculated to determine synergistic, additive, antagonistic, or indifferent interactions. Results: The MIC that inhibited 90% of S. aureus and S. epidermidis was 4 µg/mL, apart from for the MRSA subset (8 µg/mL). The MBIC that inhibited 90% of isolates was 4 µg/mL. The MBBCs ranged from 4 to 256 µg/mL. When tested together, nisin A and vancomycin yielded an FIC between 1.25 and 1.5, indicative of indifference, except for one isolate each of MRSA and MSSA, for which an additive effect (FIC of 1) was observed. Conclusions: Nisin A showed inhibitory activity against staphylococci that cause PJI.

ABSTRACT Contezolid has activity against methicillin-resistant Staphylococcus aureus (MRSA), a common cause of orthopedic infection. In a rat model of foreign body osteomyelitis, 1.3 log 10 cfu/g bone and 0.5 log 10 cfu/k-wire reduction in MRSA was found in tibiae ( P = 0.0186) and K-wires, respectively, of contezolid-treated compared to untreated rats. No MRSA was recovered from animals receiving rifampin alone or in combination with contezolid or vancomycin. There was no emergence of resistance to contezolid, vancomycin, or rifampin.