Ebru Oral

Introduction: Prosthetic joint infection (PJI) is a severe complication of orthopedic surgery, driven by biofilm-forming bacteria such as Staphylococcus aureus . Conventional antibiotics are limited in penetrating biofilms or remaining localized at the infection site. We engineered a targeted nanocarrier system by conjugating DNA aptamers specific to S. aureus biofilm components onto poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) encapsulating vancomycin. PLGA NPs were synthesized using flow-focusing microfluidics, followed by surface functionalization via EDC/NHS chemistry. Aptamer conjugation was validated using gel electrophoresis and DLS. Targeting efficiency and antimicrobial activity were evaluated through fluorescence microscopy, bacterial viability assays and in vitro biofilm models. Biodistribution and joint retention were assessed using fluorescent aptamer-conjugated NPs following intra-articular injection in rat knee joints. Results: The optimized vancomycin-loaded NPs exhibited uniform morphology, high vancomycin loading capacity (7.9 ± 2.2%), and a sustained biphasic release profile with an early high-release phase followed by slower extended release. PK modeling indicated prolonged intra-articular drug retention with concentrations maintained above the minimum inhibitory concentration (MIC) for an extended period. Aptamer conjugation preserved nanoparticle stability and enhanced binding and retention within S. aureus biofilms in vitro , supporting improved biofilm localization. In-vivo fluorescence imaging demonstrated high local retention of aptamer-conjugated NPs, supporting localized joint retention with limited off-target tissue signal. Conclusions: S. aureus -specific aptamer conjugated nanoparticles (SA Apt-P(V)) represent a targeted intra-articular delivery platform with potential for localized PJI therapy. The combined in-vitro and in-vivo findings highlight their potential to improve local therapeutic concentration, minimize systemic exposure, and enhance infection control within prosthetic joints.

Musculoskeletal infection (MSKI) is a leading cause of implant failure following orthopedic surgery for trauma or elective procedures and it is associated with catastrophic outcomes for patients and healthcare systems worldwide. International Consensus Meetings (ICM) aim to define state‐of‐the‐art, influencing clinical standards of care and accelerating discoveries by setting research priorities. The 3rd ICM was held on May 8–10, 2025 in Istanbul (Turkey) and included a 2‐year‐long Delphi process that culminated with in‐person voting by 1205 delegates on 102 General and 30 Biofilm‐specific MSKI questions. Consistent with prior ICMs, a Research Priorities Workgroup was established after the voting to interpret the results and summarize the most important future directions. Here, the group reports on several critical research priorities that emerged, which should be addressed to advance the field. These include: (1) improving diagnostics through standardized patient sampling, advanced non‐invasive imaging technologies, and biofilm‐specific biomarkers; (2) developing clinically relevant in‐vitro and in‐vivo models to rigorously and reproducibly test antibiofilm strategies; (3) identifying high priority immunological research areas, including deciphering the role of T‐cell immunity in biofilm persistence, and if T cell targeting therapies can be harnessed to disrupt chronic biofilm‐associated infection; (4) clinically evaluating novel anti‐biofilm technologies on larger cohorts of patients; and (5) addressing translational barriers through the use of multi‐center data collection and large‐scale data tools to accelerate clinical application. These research priorities aim to enhance the prevention, diagnosis, and treatment of biofilm‐associated MSKI.

Periprosthetic joint infection (PJI) is a devastating complication of total joint arthroplasty, often necessitating two‐stage revision surgery with antibiotic‐loaded bone cement (ALBC) spacers, which cannot maintain therapeutic local antibiotic concentrations over time. This study evaluated the long‐term stability, antimicrobial efficacy, and biocompatibility of an alternative material, submicron gentamicin sulfate‐loaded ultrahigh molecular weight polyethylene (SM‐GS/UHMWPE), in vitro and in vivo by using a subcutaneous rat model. SM‐GS/UHMWPE implants containing 6 wt% and 10 wt% gentamicin sulfate (GS) were fabricated and tested in vitro and simultaneously in vivo by subcutaneous implantation in rats for 4, 8, and 26 weeks. Gentamicin stability was assessed using nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography‐mass spectrometry (LC–MS/MS). Antimicrobial efficacy against Staphylococcus aureus was determined via bacterial culture assays. Mechanical integrity was evaluated using tensile testing in vitro. Systemic toxicity was assessed through liver and kidney function markers, and histological analysis of kidney and skin tissues was performed. NMR and LC–MS/MS confirmed that gentamicin remained chemically stable over 26 weeks in vivo and in vitro. Antimicrobial testing showed sustained bacterial inhibition, with implants containing 10% GS achieving a 3.5‐log reduction in bacterial counts for 6 months. Mechanical testing demonstrated minimal changes in tensile properties over time. Serum biochemical markers remained within normal ranges, and histological evaluation showed no significant inflammation or tissue damage. Gentamicin elution profiles indicated sustained release, maintaining intraarticular levels above 100 times the minimum inhibitory concentration (MIC) of S. aureus for 6 months. SM‐GS/UHMWPE implants demonstrated prolonged antibiotic release while maintaining mechanical integrity, antimicrobial efficacy, and biocompatibility. These findings support the potential use of SM‐GS/UHMWPE as a next‐generation antibiotic‐eluting spacer for PJI treatment, offering an alternative to ALBC for sustained drug release and reduced systemic toxicity risks.

While Staphylococcus aureus is the predominant pathogen in periprosthetic joint infections (PJI), polymicrobial infections involving Gram-negative organisms, such as Escherichia coli, complicate clinical outcomes. Little is known regarding implant-associated polymicrobial interactions; consequently, current PJI treatments are not optimized for their treatment. This study explored the dynamics of S. aureus-E. coli dual-species biofilms, focusing on biofilm properties, antibiotic susceptibility, and molecular interactions. Co-culture experiments revealed that E. coli significantly suppressed S. aureus biofilm viability, observed for methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA). Microscopic analyses demonstrated enhanced E. coli attachment facilitated by S. aureus matrix proteins; however, over time, E. coli dominated the biofilm composition. In the presence of E. coli, MSSA biofilm exhibited improved gentamicin susceptibility while MRSA showed limited change, underscoring strain-specific interactions. Notably, E. coli biofilms exhibited enhanced resistance to gentamicin in dual-species settings. Gene expression profiling revealed molecular adaptation in S. aureus and E. coli, triggered by the differential regulation of stress, adhesion, virulence, and biofilm-associated genes within a dual-species implant-associated biofilm. The suppression of S. aureus by E. coli presents potential therapeutic avenues, and in vivo studies and mechanistic investigations are crucial for optimizing treatment strategies targeting polymicrobial PJIs.

Periprosthetic joint infection (PJI) is a debilitating condition which significantly affects the clinical outcome of arthroplasty. Staphylococcus spp., the predominant causative organism, resists antibiotic treatments and evades the host immune response by forming complex biofilms^[1]. Although extensive studies have characterized Staphylococcal biofilms in vitro, the growth dynamics and severity of biofilms in vivo are poorly understood^[2]. Our overarching goal is to devise antibacterial tools in the form of medical devices and long-acting drug delivery devices to prevent and treat PJI. To this end, we have developed antibiotic-eluting ultrahigh molecular weight polyethylene (UHMWPE), which has the potential to not only deliver antibiotics locally but to improve the functional recovery of PJI patients^[3,4,5].We established a subcutaneous implant infection model in the rat model using methicillin-sensitive S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) and analyzed implant capsule tissues across multiple time points (POD0, POD1, POD3, POD7) using RNA bulk sequencing. Differential gene expression, KEGG and GO enrichment, ssGSEA, and CIBERSORTx analyses were employed to identify diagnostic markers and characterize immune cell infiltration profiles. Additionally, immune profiling was conducted on a public dataset related to clinical orthopedic implant infections to determine overlaps in transcriptional pathways.The comparison and stratification of the risk and severity of infection across different preclinical models can guide clinical dosing and the design of antibacterial implants to effectively prevent or treat PJI. Here, we are going to present our material work in controlling the release rates of local antibiotics and synergistic non-antibiotic molecules^[6] and our work on bacterial^[7] and host characterization in vivo (including our transcriptomics above) and in vitro to determine clinical dosing guidance and administration timelines for these antibacterial implants.)

Little is known about the in-vivo dynamics of biofilms associated with medical-device infections and their interplay with systemic inflammation, local immune responses, and tissue healing processes. There may be an opportunity to tailor therapeutic strategies to target these dynamics to improve treatment outcomes. We investigated immune responses to a methicillin-susceptible (ATCC 12600) and a multi-drug resistant (L1101) S. aureus strain using a rat subcutaneous implant model, analyzing local and systemic inflammation through 19 gene expressions over 21 days. Our goals were to identify differences in the immune response due to infection and also with respect to the two strains. We observed that systemic inflammation, indicated by α-2-macroglobulin, was elevated in the initial stages (up to day 7). Local inflammatory cytokine levels (IL-6, TNF-α, IL-6, TNF-α, IL-1β, IL10, IL-17, IL12a, IL12b, IFNG) varied by strain, typically higher against the clinical strain. Infections generally hindered early macrophage (MCSF1) and T-cell (CD4, CD5, CD6, CD8A) recruitment, particularly in cases involving the clinical strain. Conversely, a better healing response was observed in the infection of the more susceptible ATCC 12600 strain (VEGF, CXCR1, CXCR2, MMP-1, MMP-3, MMP-13). These results are crucial for understanding immune responses to such infections, guiding therapeutic strategies.

Periprosthetic joint infection (PJI) is a severe complication of total joint arthroplasty (TJA), leading to high rates of revision surgeries, long‐term morbidity, and mortality. Conventional antibiotic treatments often suffer from limited bioavailability and systemic toxicity. This study explores a novel approach using vancomycin‐loaded poly(lactic‐co‐glycolic) acid (PLGA) microparticles (VMP) formulated via a microfluidic double emulsion method for controlled, localized drug delivery for managing PJI. The PLGA microparticles were synthesized to achieve high loading capacity and sustained vancomycin release, aiming to maintain therapeutic intra‐articular concentrations. In vitro characterization demonstrated optimal loading capacity (up to 28% w/w), morphology with a homogeneous particle size distribution (49–65 µm), and sustained release profiles over 8 weeks. In vivo efficacy was evaluated using a rat joint infection model, showing significant reductions in bacterial viability and enhanced bone healing compared to controls. Weight‐bearing recovery assessments showed that VMP‐treated rats regained functionality significantly earlier than controls (p < 0.05). Radiographic, histological, and immunofluorescent analyses confirmed reduced inflammation and improved bone integrity with VMP treatment. These findings suggest that microfluidic‐synthesized PLGA microparticles provide a promising strategy for localized, controlled release of antibiotics, potentially helping the management of PJI and improving postsurgical outcomes. Future research should explore the long‐term effects and scalability of clinical applications. This study lays the foundation for advancing controlled release systems in orthopedic postoperative care.

Radiation crosslinking could be used to obtain wear-resistant UHMWPE-based implants with suitable antibacterial activity against Staphylococcus aureus, which is one of the main pathogens involved in peri-prosthetic joint infections.