Sepsis accounts for nearly 20% of global mortality, with antibiotic resistance worsening clinical outcomes. Rapid antibiotic administration and accurate pathogen identification remain crucial. It is well now known that extracellular vesicles (EVs) from human cells and bacterial membrane vesicles (bMVs) play a central role in the interaction between host and pathogen and represent promising biomarkers for early infections. This study investigated how antibiotic exposure alters EV responses in Staphylococcus aureus (SA)spiked blood and compared these findings with EV proteome profiles from bacteremia patients. In an in vitro model, whole blood from healthy donors was spiked with SA at a multiplicity of infection (MOI) of 0.001, treated with clinically relevant concentrations of piperacillin-tazobactam, vancomycin, or moxifloxacin, and plasma was subsequently isolated for EV analysis. EVs were isolated using the Miltenyi Pan EV Kit and analyzed by bead-based flow cytometry and high-resolution LC-MS/MS. In parallel, serum EVs from healthy controls (n = 6) and bacteremia patients (n = 12; 6 blood culture-positive and 6 culture-negative) were analyzed using the same workflow. Flow cytometry revealed increased levels of CMO⁺ CD45⁺ PanEV⁺ SA⁺ vesicles in SA-spiked samples, particularly following low-dose piperacillin-tazobactam and high-dose vancomycin treatment, despite minimal changes in vesicle size and total particle counts. Proteomic analysis of plasma EVs showed significant alterations in protein composition, including increased abundance of the SA-derived ribosomal protein rplU and host defense-associated proteins. Functional enrichment highlighted pathways related to neutrophil degranulation, vesicle-mediated transport, and antibacterial responses. In patient samples, serum EVs were enriched in acute-phase and immune-related proteins, including SERPINA1, SERPINA3, CRP, and SAA2, along with canonical EV markers such as CD81 and syntenin-1, irrespective of blood culture status. Antibiotic exposure and SA infection are associated with measurable changes in the human EV proteome, characterized by enrichment of immune and host defense-related proteins despite stable vesicle numbers. Similar EV-associated protein patterns were observed in both blood culture-positive and -negative patient samples, reflecting shared features of the systemic host response to infection and highlighting the potential of EV profiling to capture infection-associated biological signals.
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Sepsis accounts for nearly 20% of global mortality, with antibiotic resistance worsening clinical outcomes. Rapid antibiotic administration and accurate pathogen identification remain crucial. It is well now known that extracellular vesicles (EVs) from human cells and bacterial membrane vesicles (bMVs) play a central role in the interaction between host and pathogen and represent promising biomarkers for early infections. This study investigated how antibiotic exposure alters EV responses in Sta...
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