PCR with degenerate primers for highly conserved DNA polymerase gene of the herpes virus family show neither HHV-8 nor a variant in bone marrow stromal cells from multiple myeloma

The possibility has been raised that either a human herpesvirus-8 (HHV-8) variant or a novel, unidentified, gamma-herpesvirus related to HHV-8 is frequently associated with multiple myeloma (MM), which could explain the lack of antibodies to HHV-8 antigens and the discordant results from polymerase chain reaction (PCR) studies of HHV-8-specific sequences in MM patients. Thus, we used a sensitive PCR assay with degenerate primers targeting the highly conserved DNA polymerase gene of the herpesvirus family to examine the long-term cultures of bone marrow stromal cells (BMSCs) from 19 MM, 3 monoclonal gammopathies of undetermined significance and 6 control patients. Both the culture supernatant and the adherent stromal layer were examined from the 2nd until the 8th week of culture to assess the immunophenotype of the various cell types harvested for the molecular analysis. BMSCs consisted of a mixed population of fibroblast, macrophage, dendritic and endothelial cells. An amplified product of the expected size was obtained only in 3 MM cases, both in the adherent and nonadherent fractions. Direct sequencing and alignment of the nucleotide and amino acid sequences showed that the DNA sequences were 100% identical to Epstein-Barr virus (EBV) DNA. The PCR positivity was due to the presence of EBV-infected lymphoblastoid cells with plasmacytoid features, expressing the EBV-encoded latent membrane protein-1 and detectable either in the stromal cells or in the culture supernatant. Our data do not support a causal role of either HHV-8 or a novel herpesviral variant related to HHV-8 in MM.