Epstein-Barr virus-infected B cells expanding in germinal centers of infectious mononucleosis patients do not participate in the germinal center reaction

Abstract

To assess the impact of the germinal center (GC) reaction on viral spread in Epstein-Barr virus (EBV) infection, we isolated EBV(+) GC B cells from the tonsils of two infectious mononucleosis patients, sequenced their rearranged V genes, and determined expression of the EBV latency genes EBV nuclear antigen 2 and latent membrane protein 1. Most EBV(+) GC B cells belonged to clones of cells harboring somatically mutated V gene rearrangements. Ongoing somatic hypermutation, the hallmark of the GC reaction, was seen only in uninfected GC B cell clones, not in EBV(+) B cell clones. Thus, in infectious mononucleosis, GC and/or memory B cells are directly infected by EBV and expand without somatic hypermutation, whereas the GC passage of EBV-infected naive B cells does not contribute detectably to the generation of infected memory B cells, the main reservoir of EBV during persistence. Most, if not all, EBV-infected cells in GCs exhibited an unusual EBV gene expression pattern in that they were positive for EBV nuclear antigen 2 but negative for latent membrane protein 1. Although the three main types of EBV-associated B cell lymphomas (Burkitt's, Hodgkin's, and posttransplant lymphomas) presumably are derived from GC B cells, EBV(+) GC B cells resembling these EBV(+) GC B cell lymphomas in terms of EBV gene expression and somatic hypermutation pattern could not be identified.

Figure 1

Phenotypic characterization of EBV-infected cells located in GCs and the IFR. Staining was performed on frozen tonsillar sections. Counterstaining was performed by using haemalaun (blue staining, A, B, and D) or Hoechst 33258 (blue fluorescent staining, C and E). (A–C) Sections of case 1 stained for EBER transcripts (dark blue-purple staining, A) and EBNA2 (dark brown staining, B) and LMP1 expression (red fluorescent staining, C). The border of GC1 to its mantle is indicated. (D) Section of case 2 stained for EBER transcripts (dark blue-purple staining). Approximately 10 EBER⁺ cells are located in GC2 (left) and GC3 (right). (E) Section of case 2 stained for LMP1 expression (red fluorescent staining) showing the interfollicular location of LMP1-expressing cells. Borders of GC3 and its mantle zone are indicated. A part of GC3 is visible in the lower right corner.