Incomplete B-cell reconstitution in ART-treated people living with HIV is associated with EBV-linked lymphoma progression.

Background: Despite the widespread use of antiretroviral therapy (ART), people living with HIV continue to exhibit persistent immune alterations. Among the most affected cell populations are B lymphocytes, which show disruptions in differentiation, class-switch recombination, and the development of immunological memory. However, the relationship between these abnormalities, Epstein-Barr virus (EBV) coinfection, and clinical outcomes across different stages of HIV disease remains poorly understood. Methods: We analyzed 272 patients living with HIV from Hospital Universitario San Ignacio (Bogotá, Colombia), divided in four groups: clinical stage (S1, S2, S3 and lymphoma), clinical categories: AIDS-defining diseases, non-AIDS-defining diseases, and coinfections (CI), time on ART (<1 year, >1 year); and EBV coinfection. B lymphocyte subpopulations and immunoglobulin isotypes were quantified (next-generation flow cytometry). Cytokines (multiplex assays) and EBV viral load (quantitative PCR) were measured. Results: HIV progression was observed, associated with B cell compartment remodeling, characterized by depletion of naïve and memory B cells (smIgA1-2 and smIgG1-4) and an increase in immature/transitional cells and alterations in smIgM with isotype switching. These variations correlated negatively with clinical stage (ρ up to -0·43). The cytokine profile showed a persistent inflammatory signature (MIP-1β, G-CSF, FLT-3L, and IL-3). EBV coinfection intensified this phenotype, being associated with elevated levels of IL-6, IL-10, IL-15, TNF-α, and sCD40L, and with greater loss of memory cell subpopulations. Patients with AIDS-defining diseases and lymphomas exhibited the most profound alteration. Even after prolonged ART (>1 year), B cell reconstitution remained incomplete and biased toward immature phenotypes. Conclusions: This study shows that recovery of the B cell compartment under ART is incomplete and functionally unbalanced. Humoral memory loss, bias toward immature phenotypes, and persistent inflammation create a state of dysregulation that is exacerbated by EBV coinfection, especially in advanced clinical stages. These immunological defects provide a basis for the pathogenesis of non-AIDS-defining diseases and Epstein-Barr-associated lymphomas. Our findings support the need to integrate advanced immunological assessments, including standardized flow cytometry, as a complement to CD4+ count and viral load, in order to more accurately characterize immune competence and anticipate clinical risks in people living with HIV.