Primary effusion lymphoma (PEL) is a unique form of malignant lymphoma associated with infection by the Kaposi's sarcoma-associated herpesvirus (KSHV)/human herpesvirus-8 (HHV-8). The majority of PELs also contain the EBV genome. Although viral infection is believed to play a critical role in the pathogenesis of PEL, it has been suggested that additional molecular lesions are required for the development of PEL. Alternative splicing of pre-mRNA is an important mechanism in the regulation of cellular and viral gene expression. Deregulation of pre-mRNA splicing may shift the gene expression balance and lead to the development of cancer. In order to investigate mRNA splicing in PELs, we examined mRNA splicing of three genes, DNA polymerase beta (pol beta), Bcl-x and CD45, in eight PEL cell lines. We found that the average variant percentage of pol beta in PEL cell lines is two times higher than in peripheral blood mononuclear cells (PBMC) and that the variant pattern of genes bcl-x and CD45 is quite different in PEL cell lines than in PBMC. In addition, we also found that the percentage of variant pol beta increased two-fold in PBMC following Epstein-Barr virus (EBV) infection. Therefore, viral infection may contribute to mRNA alternative splicing in PEL. In order to explore the mechanism by which viral infection affects mRNA splicing, we also examined the roles of genes KS-SM, SM and EBERs and viral copies in mRNA splicing. Our findings indicate that various factors acting as positive or negative regulators may be involved in mRNA alternative splicing caused by viral infection. In conclusion, mRNA splicing in PEL can be altered by viral infection and this alteration may contribute to the pathogenesis of PEL.