Bovine leukemia virus (BLV), a close relative of human T cell leukemia virus types-1 and -2 (HTLV-1 and HTLV-2), is the etiologic agent responsible for enzootic bovine leukosis (EBL), which is the most common neoplastic disease of cattle. Infection by BLV may remain clinically silent at the aleukemic (AL) stage. However, in 30% of infected cattle the infection may manifest as persistent lymphocytosis (PL; a condition characterized by an increase in the number of B lymphocytes), and in around 1–5% of cases it may manifest as B cell lymphoma after a long period of latency
. Sheep experimentally inoculated with BLV develop B cell tumors at a higher frequency than naturally infected cattle, and the period of latency is shorter
In infected cattle with no evident tumor, BLV has been identified in B cells, CD2+ T cells, CD3+ T cells, CD4+ T cells, CD8+ T cells, γ/δ T cells, monocytes and granulocytes
[4–6]. By contrast, the increase in lymphocyte numbers observed in cows with PL is entirely attributable to the expansion of the CD5- and CD5+ B cell subpopulations, indicating that CD5- and CD5+ B cells are the only mononuclear cells within the peripheral blood that are significantly infected with BLV
. Furthermore, the most consistent tumor cell phenotypes isolated from cattle with EBL are CD5+, CD6–, B1 low+, B2+, major histocompatibility complex class II+, and either sIgM+ or sIgM–; this indicates the involvement of the CD5+ B cell sub population rather than the CD5-B cell sub population. However, the mechanism by which BLV induces uncontrolled CD5+ B cell proliferation is unknown. It is interesting to note that in sheep, transformed B cells show a CD5- phenotype
. Indeed, we previously showed that the extended survival of peripheral blood mononuclear cells (PBMCs) ex vivo was mainly due to the presence of BLV-expressing CD5– B cells, indicating that sheep CD5– B cells may be particularly susceptibility to the transforming effects of BLV
. This increase in the survival of BLV-expressing sheep PBMCs was also associated with an increase in the expression of mRNA for bcl-xl, but not that for bcl-2 or bax. However, the mechanism by which BLV protects ex vivo cultured cells against apoptosis is unknown.
After infecting cattle, BLV enters a period of latency, during which expression is blocked at the transcriptional level
[10–12]. BLV-infected cattle retain at least one copy of the full-length proviral genome throughout the course of the disease
, suggesting that the BLV provirus remains integrated within the cellular genome
, even in the absence of detectable BLV antibodies
. Therefore, diagnostic BLV polymerase chain reaction (PCR) techniques, which detect the integrated BLV proviral genome within the host genome, are now commonly used to detect BLV infection in addition to routine diagnostic tests such as agar gel immunodiffusion and enzyme-linked immunosorbent assays (ELISAs)
[13, 15–18]. Recently, we developed a new quantitative real-time PCR method using Coordination of Common Motifs (CoCoMo) primers to measure the proviral load of both known and novel BLV variants in BLV-infected animals
[14, 19]. The assay was highly effective in detecting BLV in cattle from a number of international locations. The BLV-CoCoMo-qPCR technique amplifies a single-copy host gene, the bovine leukocyte antigen (BoLA)-DRA gene, in parallel with viral genomic DNA, which effectively normalizes the level of viral genomic DNA. Thus, we were able to show that the proviral load correlates not only with the level of BLV propagation, as assessed by syncytium formation, but also with BLV disease progression.
While the primary cellular target of BLV is B cells, recent studies suggest that monocytes, granulocytes, CD2+ T cells, CD3+ T cells, CD4+ T cells, CD8+ T cells and γ/δ T cells are also targets
[4–6, 20]. However, because Mirsky et al.
, fractionated B cells into the CD5+ IgM+ B cells and CD5- IgM+ B cell subpopulations, but did not fractionate CD2+ T cells into the CD4+ and CD8+ T cell subpopulations. In contrast, Wu et al.
 isolated the CD4+ and CD8+ T cell subpopulations, but did not fractionate B cells into the CD5+ IgM+ B cells and CD5- IgM+ B cell subpopulations. It remains to be clarified the variations of the BLV proviral load among CD5+ IgM+ B cells, CD5- IgM+ B cells, CD4+ T cells, and CD8+ T cells in the same experiment. Therefore, to clarify whether these subpopulations are susceptible to BLV infection, we obtained PBMCs from cattle naturally infected with BLV and isolated CD5+ IgM+ B cells, CD5- IgM+ B cells, CD4+ T cells, and CD8+ T cells by flow cytometry or using magnetic beads. We then estimated the BLV proviral load using the BLV-CoCoMo-qPCR technique. The results show that CD5+ IgM+ B cells, CD5- IgM+ B cells, CD4+ T cells, and CD8+ T cells are all primary targets for BLV.