The primary Epstein-Barr virus (EBV) infection occurs in the oropharynx, where the virus infects B cells and subsequently establishes latency in the memory B-cell compartment. EBV has previously been shown to induce changes in the cell surface expression of several chemokine receptors in cell lines and the transfection of EBNA2 or LMP1 into a B-cell-lymphoma-derived cell line decreased the expression of CXCR4. We show that in vitro EBV infection reduces the expression of CXCR4 on primary tonsil B cells already 43 hr after infection. Furthermore, EBV infection affects the chemotactic response to stromal cell-derived factor (SDF-1)alpha/CXCL12, the ligand for CXCR4, with a reduction of SDF-1alpha-induced migration. To clarify whether this reduced migration is EBV-specific or a consequence of cell activation, tonsillar B cells were either infected with EBV, activated with anti-CD40 and interleukin-4 (IL-4) or kept in medium. Activation by anti-CD40 and IL-4 decreased the CXCR4 expression but the CD40 + IL-4-stimulated cells showed no reduction of chemotactic efficacy. Our finding suggests that changing the SDF-1alpha response of the EBV-infected B cells may serve the viral strategy by directing the infected cells into the extrafollicular areas, rather than retaining them in the lymphoepithelium.