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Epstein-Barr virus (EBV) isolates can be broadly classified as type 1 or type 2 on the basis of allelic polymorphism of the virus-encoded nuclear antigens EBNAs 2, 3a, 3b and 3c, and individually identified based on Mr values of their EBNA proteins (EBNA type). Here we have used this natural heterogeneity amongst isolates to re-examine the question of EBV persistence in vivo, asking in particular whether virus carriage in oropharyngeal epithelium and/or in B lymphoid tissues involves infection with a single or with multiple virus strains. Firstly, 76 healthy virus carriers were classified into serotype groups on the basis of preferential antibody reactivity to type 1 EBNAs (serotype 1) or to type 2 EBNAs (serotype 2); 60 of the 76 donors were serotype 1, four of the 76 donors were serotype 2 and 12 of the 76 donors were anti-EBNA 2, 3a, 3b, 3c antibody-negative and therefore could not be serotyped. Representative donors from each group were then selected for virus isolations from blood (by spontaneous in vitro transformation) and from throat washings (by cord blood cell transformation). All 13 serotype 1 donors tested and six of seven non-serotypeable donors gave a type 1 virus isolate, whereas all four serotype 2 donors and one of the seven non-serotypeable donors gave a type 2 isolate. Multiple transforming virus isolates from any one donor, whether from blood or throat washings, were all of the one strain characteristic of that particular donor; sequential isolations showed retention of the same strain over several years. Finally, throat washing samples from these same donors were examined for amplifiable EBV DNA in the polymerase chain reaction using EBV type-specific oligonucleotide primers and probes derived from the polymorphic EBNA 2 and EBNA 3c loci. The results were consistent with earlier virus isolation studies, each individual donor showing amplification either of type 1 or type 2 sequences. We conclude that multiple EBV infections must occur rarely, if at all, in healthy virus carriers; EBV persistence in vivo is characterized by dominance of a single transforming virus strain.

The Epstein--Barr virus carrier state: dominance of a single growth-transforming isolate in the blood and in the oropharynx of healthy virus carriers