Establishment of an animal model of MRSA osteomyelitis and evaluation of an in situ diagnostic using CY7-labeled anti-PBP2a mAb

Introduction: Methicillin-resistant Staphylococcus aureus (MRSA) osteomyelitis is a severe bone infection with high morbidity that poses a clinical challenge.Early diagnosis is crucial for effective treatment, as delayed detection increases the risk of sepsis, sequelae, and death.Experimental animal models are essential for understanding pathogenesis and testing new diagnostic and therapeutic strategies.In this context, developing an in situ diagnostic method using labeled anti-PBP2a monoclonal antibody (mAb) may not only provide new tools for early detection and monitoring, but also significantly reduce healthcare costs, improving patient outcomes and the overall healthcare system. Objectives:To establish an animal model of MRSA osteomyelitis and evaluate an in situ diagnostic method using CY7-labeled anti-PBP2a mAb.Methodology: Various concentrations (10, 10 4 , and 10 5 CFU) of a bioluminescent MRSA strain (SAP231) were inoculated into the femur-tibial joint of six 8-week-old female BALB/c mice after sedation and local anesthesia.Animals were monitored for 21 days to assess infection establishment and clinical signs.In a separate experiment, nine animals received 10 4 CFU of SAP231 via the same inoculation method.After 72 hours, different concentrations (100, 200, and 300 g) of CY7-labeled murine anti-PBP2a IgG were administered intravenously, followed by IVIS imaging and radiography at 2, 4, 6, and 24 hours postinjection.Images captured luciferase emission from SAP231 and Cy7 fluorescence.The animal ethics committee approved all procedures.Results: All animals in the first experiment emitted bioluminescent signals throughout 21 days, indicating 10 4 CFU inoculation as the most suitable for greater bioluminescent signal and animal survival.Bacterial signals were detected in the bone after excision, confirming a successful infection establishment.Additionally, animals showed clinical signs of osteomyelitis, like lameness.In the second experiment, all animals receiving anti-PBP2a IgG at any concentration exhibited fluorescence at the infection site, indicating antibody binding to MRSA and enabling infection detection. Conclusion:The animal model of MRSA osteomyelitis was successfully established, and the CY7labeled anti-PBP2a IgG antibody effectively detected the infection in situ.This is a promising approach, and further studies should be performed to validate and potentiate this new method for the early detection and monitoring of MRSA osteomyelitis.