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Journal of Virology, May 1999, p. 4052-4061, Vol. 73, No. 5
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

In Vitro Infection of Human Peripheral Blood Mononuclear Cells by GB Virus C/Hepatitis G Virus

Marta Fogeda, Sonia Navas,dagger Julio Martín,Dagger Mercedes Casqueiro, Elena Rodríguez, Carlos Arocena, and Vicente Carreño*

Department of Hepatology, Fundación Jiménez Díaz, and Fundación para el Estudio de las Hepatitis Virales, Madrid, Spain

Received 30 October 1998/Accepted 12 February 1999

GB virus C (GBV-C), also known as hepatitis G virus, is a recently discovered flavivirus-like RNA agent with unclear pathogenic implications. To investigate whether human peripheral blood mononuclear cells (PBMC) are susceptible to in vitro GBV-C infection, we have incubated PBMC from four healthy blood donors with a human GBV-C RNA-positive serum. By means of (i) strand-specific reverse transcription-PCR, cloning, and sequencing; (ii) sucrose ultracentrifugation and RNase sensitivity assays; (iii) fluorescent in situ hybridization; and (iv) Western blot analysis, it has been demonstrated that GBV-C is able to infect in vitro cells and replicate for as long as 30 days under the conditions developed in our cell culture system. The concentration of GBV-C RNA increased during the second and third weeks of culture. The titers of the genomic strand were 10 times higher than the titers of the antigenomic strand. In addition, the same predominant GBV-C sequence was found in all PBMC cultures and in the in vivo-GBV-C-infected PBMC isolated from the donor of the inoculum. GBV-C-specific fluorescent in situ hybridization signals were confined to the cytoplasm of cells at different times during the culture period. Finally, evidence obtained by sucrose ultracentrifugation, RNase sensitivity assays, and Western blot analysis of the culture supernatants suggests that viral particles are released from in vitro-GBV-C-infected PBMC. In conclusion, our study has demonstrated, for the first time, GBV-C replication in human lymphoid cells under experimental in vitro infection conditions.

* Corresponding author. Mailing address: Department of Hepatology, Fundación Jiménez Díaz, Avda. Reyes Católicos, 2, 28040 Madrid, Spain. Phone: 34-91.543.19.64. Fax: 34-91.544.92.28. E-mail: vcarreno{at}uni.fjd.es.

dagger Present address: Department of Microbiology, University of Pennsylvania School of Medicine, Philadelphia, PA 19104-6076.

Dagger Present address: Department of Neurology, University of Pennsylvania, Philadelphia, PA 19104-6146.

Journal of Virology, May 1999, p. 4052-4061, Vol. 73, No. 5
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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