A Multivariable Plasma Extracellular Vesicle Surface Profile Associated with Post-COVID-19 Syndrome

Post-COVID-19 syndrome (PCS) is characterized by fatigue, neurological impairment and systemic symptoms. This heterogeneity of symptoms hinders biomarker development. Here, we profiled extracellular-vesicle (EV) surface markers in plasma and CSF from 61 participants with PCS (COVID ^post ), 80 recovered controls (COVID ^reco ), and 10 participants with non-SARS-CoV-2 post-viral syndromes. EVs were analysed by bead-based multiplex flow cytometry using tetraspanin-directed (TSPN) and phosphatidylserine-directed lactadherin (PS) detection. Amongst 37 targets covering tetraspanins and vasculature-, immunity- and stemness-associated markers, none met a 1% false-discovery-rate threshold. However, L1-regularized logistic regression under fully nested 5×5 cross-validation identified a distributed plasma EV profile, with mean out-of-fold areas under the receiver operating characteristic curve (AUCs) of 0.788 (95% CI 0.715–0.852) for TSPN and 0.716 (95% CI 0.636–0.792) for PS detection. Across the pooled COVID ^post and COVID ^reco population, EV classification scores covaried with clinical group differences, but did not track clinical severity within either cohort. These PCS-EV classification scores decreased at one-year follow-up in COVID ^post participants. Our findings identify an internally cross-validated multivariable EV surface profile associated with COVID ^post versus COVID ^reco status and support independent validation and exploration of EV-based biomarkers in post-viral fatigue syndromes.