We have examined sequence variations in the EBNA-1 protein of EBV in normal peripheral blood lymphocytes (PBL) and Burkitt's lymphomas (BL). We find two EBNA-1 strains P (prototype) and V (variant) which differ by 15 amino acids. Each strain has two subtypes defined by the amino acid at position 487 (P-ala, P-thr, V-pro and V-leu). In PBLs from 32 normal individuals, up to three of these subtypes were found in each sample, but the V strain did not occur in the absence of P strain viruses, nor was the V-leu subtype ever observed in normal PBL. In BLs only a single subtype was observed in each tumor. The P-thr and V-leu subtypes were more frequently seen than the P-ala and V-pro subtypes, which occurred in only two and one of the 36 tumor samples respectively. The P-thr was the most commonly observed subtype in peripheral blood of both American and African lymphocytes as well as in African tumors. However, in 11 of 12 American tumors, the EBNA-1 subtype was V-leu. These data indicate that some EBNA-1 subtypes are more likely to lead to oncogenesis, and one subtype, V-leu, appears only to occur in tumors.