Analysis of Epstein-Barr virus (EBV) type and variant in spontaneous lymphoblastoid cells and Hu-SCID mouse tumours

Epstein-Barr virus (EBV) type and strain variations were examined using both lymphoblastoid cell lines (LCLs), spontaneously derived in vitro from peripheral blood mononuclear cells (PBMC) of 15 HIV-1-seropositive individuals; and SCID mouse tumours induced by inoculation of PBMC from 11 healthy human donors (Hu-SCID tumours). Polymerase chain reaction (PCR) analysis disclosed that all but one of the 26 EBV + samples harboured EBV nuclear antigen (EBNA) 2 and 3C type A virus. On the other hand, single strand conformation polymorphism (SSCP) analysis using Epstein-Barr encoded RNA (EBER) specific primers detected an AG876-like (type B) band pattern in 21 of the 26 EBV + samples. Three Hu-SCID tumours scored as B95.8-like (type A), and two showed neither a type A nor a type B SSCP migration pattern. Sequence analysis of the amplified EBER fragments confirmed the PCR-SSCP findings; moreover, additional mutations were present not only in the two EBV + samples with anomalous SSCP pattern, but also in two other samples with a standard SSCP profile. Thus, EBER analysis did not correlate with EBNA typing, and appeared to be unsuitable for EBV type assessment. Latent membrane protein (LMP) analysis disclosed, on the whole, sever size variants: as expected, the differences were due to the variable numbers of a 33-bp repeat in the amplified fragment, as assessed by direct sequencing. The broader variability detected by LMP analysis should prove more useful than typing for assessing the presence of single and/or mixed variants resulting from EBV reactivation and/or reinfection.