EBV Conference 2008 Guangzhou - Baylor College of Medicine

10 (RegID: 1448)
Makoto Ohashi
Institution: Department of Medicine, Brigham and Women's Hospital, Harvard Medical School
e-mail: mohashi@rics.bwh.harvard.edu
EXPERIMENTAL RHESUS MACAQUE INFECTION WITH A RHESUS
LYMPHOCRYPTOVIRUS MOLECULAR CLONE.
Makoto Ohashi, Angela Carville, Carol Quink and Fred Wang
Oralabstract:
Background: Natural and experimental infection of rhesus macaques with the EBV-related rhesus
lymphocryptovirus (rhLCV) reproduces the biology of EBV infection in humans. We established a
Bacterial Artificial Chromosome derived rhLCV clone in order to develop an experimental genetic system
for this animal model of EBV infection.
Methods: The rhLCV molecular clone 16.5 was derived by homologous recombination into the viral
episome genome, rescue in E. coli, replication in eukaryotic cells, and recovery of transforming virus by
infection of rhesus PBMC.
Results: Clone 16.5 rhLCV was competent for viral replication and B cell immortalization in tissue
culture. Southern blot and PCR analysis confirmed the presence of a signature lox-scar sequence at the
site of homologous recombination in the rhBARF1 ORF. A SHIV-immunosuppressed,
rhLCV-naïve macaque was orally inoculated to test whether the rhLCV clone 16.5 was capable of
successfully infecting and persisting in vivo. Acute viral infection of the peripheral blood was detected
by EBERs RT-PCR at day 21, and recovery of a spontaneous LCL from the peripheral blood confirmed
the signature scar sequence for the rhLCV molecular clone. Persistent virus infection was established,
and oral shedding of virus was detected.
Conclusion: We have established the first rhLCV molecular clone and infection of a rhesus macaque with
a genetically engineered rhLCV. The rhLCV clone 16.5 is capable of acutely invading the peripheral blood
after oral inoculation, establishing persistent infection in the peripheral blood, and reactivating for oral
viral shedding. The results suggest that BARF1 is not essential for acute, persistent, or reactivated
infection in an immunosuppressed host. These studies establish a new genetic system for studying the
role of viral genes for LCV infection of the natural host.
EBV Conference 2008 Guangzhou
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