We have characterized a new Epstein-Barr-virus(EBV)-like herpesvirus associated with lymphomas of SIV-infected cynomolgus (Macaca fascicularis) monkeys and propose that this virus is designated herpesvirus macaca fascicularis I (HVMFI). Genomic regions in HVMF1 of potential significance for tumor pathogenesis were analyzed by Southern blotting, PCR and sequencing, and compared with human EBV DNA. Virus from 7 SIV-associated lymphomas and one lymphoma-derived cell line were shown to share homology with the EBNA1- and EBNA2-coding regions of EBV, while some homology to EBV-LMP1 was detectable only at low-stringency hybridization. Homologous regions to the long internal repeat (IR1; BamHI W), the EBER1 and 2 and the latent origin of DNA replication (oriP) could also be demonstrated in HVMF1. These coding regions, except IR1, showed restriction-enzyme maps different from those of EBV. Sequencing of the EBNA5 homologous region of HVMF1 DNA, corresponding to exons W1 and W2, showed 65% homology to the EBV exons W1 and W2, and 80% to the whole region including the intron. Since EBNA5 has been reported to bind tumor-suppressor proteins p53 and Rb in vitro, the HVMF1 homology could be important for the lymphomagenic capacity of this monkey herpesvirus.