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Journal of Virology, January 2009, p. 384-395, Vol. 83, No. 1
0022-538X/09/$08.00+0     doi:10.1128/JVI.01854-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Deletions of the Hypervariable Region (HVR) in Open Reading Frame 1 of Hepatitis E Virus Do Not Abolish Virus Infectivity: Evidence for Attenuation of HVR Deletion Mutants In Vivo{triangledown}

R. S. Pudupakam,1 Y. W. Huang,1 T. Opriessnig,2 P. G. Halbur,2 F. W. Pierson,1 and X. J. Meng1*

Center for Molecular Medicine and Infectious Diseases, Department of Biomedical Sciences and Pathobiology, College of Veterinary Medicine, Virginia Polytechnic Institute and State University, Blacksburg, Virginia,1 Department of Veterinary Diagnostic and Production Animal Medicine, College of Veterinary Medicine, Iowa State University, Ames, Iowa2

Received 3 September 2008/ Accepted 11 October 2008

Hepatitis E virus (HEV) is an important human pathogen, although little is known about its biology and replication. Comparative sequence analysis revealed a hypervariable region (HVR) with extensive sequence variations in open reading frame 1 of HEV. To elucidate the role of the HVR in HEV replication, we first constructed two HVR deletion mutants, hHVRd1 and hHVRd2, with in-frame deletion of amino acids (aa) 711 to 777 and 747 to 761 in the HVR of a genotype 1 human HEV replicon. Evidence of HEV replication was detected in Huh7 cells transfected with RNA transcripts from mutant hHVRd2, as evidenced by expression of enhanced green fluorescent protein. To confirm the in vitro results, we constructed three avian HEV mutants with various HVR deletions: mutants aHVRd1, with deletion of aa 557 to 585 ({Delta}557-585); aHVRd2 ({Delta}612-641); and aHVRd3 ({Delta}557-641). Chickens intrahepatically inoculated with capped RNA transcripts from mutants aHVRd1 and aHVRd2 developed active viral infection, as evidenced by seroconversion, viremia, and fecal virus shedding, although mutant aHVRd3, with complete HVR deletion, was apparently attenuated in chickens. To further verify the results, we constructed four additional HVR deletion mutants using the genotype 3 swine HEV as the backbone. Mutants sHVRd2 ({Delta}722-781), sHVRd3 ({Delta}735-765), and sHVRd4 ({Delta}712-765) were shown to tolerate deletions and were infectious in pigs intrahepatically inoculated with capped RNA transcripts from the mutants, whereas mutant sHVRd1 ({Delta}712-790), with a nearly complete HVR deletion, exhibited an attenuation phenotype in infected pigs. The data from these studies indicate that deletions in HVR do not abolish HEV infectivity in vitro or in vivo, although evidence for attenuation was observed for HEV mutants with a larger or nearly complete HVR deletion.

* Corresponding author. Mailing address: Department of Biomedical Sciences and Pathobiology, Virginia Polytechnic Institute and State University, 1410 Price's Fork Road, Blacksburg, VA 24061-0342. Phone: (540) 231-6912. Fax: (540) 231-3426. E-mail: xjmeng{at}vt.edu

{triangledown} Published ahead of print on 22 October 2008.

Journal of Virology, January 2009, p. 384-395, Vol. 83, No. 1
0022-538X/09/$08.00+0     doi:10.1128/JVI.01854-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Kalia, M., Chandra, V., Rahman, S. A., Sehgal, D., Jameel, S. (2009). Heparan Sulfate Proteoglycans Are Required for Cellular Binding of the Hepatitis E Virus ORF2 Capsid Protein and for Viral Infection. J. Virol. 83: 12714-12724 [Abstract] [Full Text]  


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