This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Google Scholar
Right arrow Articles by Añez, G.
Right arrow Articles by Lai, C.-J.
PubMed
Right arrow PubMed Citation
Right arrow Articles by Añez, G.
Right arrow Articles by Lai, C.-J.

 Previous Article  |  Next Article 

Journal of Virology, October 2009, p. 10384-10394, Vol. 83, No. 20
0022-538X/09/$08.00+0     doi:10.1128/JVI.01083-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Passage of Dengue Virus Type 4 Vaccine Candidates in Fetal Rhesus Lung Cells Selects Heparin-Sensitive Variants That Result in Loss of Infectivity and Immunogenicity in Rhesus Macaques{triangledown}

Germán Añez,1 Ruhe Men,1 Kenneth H. Eckels,2 and Ching-Juh Lai1*

Molecular Viral Biology Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892-8005,1 Walter Reed Army Institute of Research, Silver Spring, Maryland 209102

Received 27 May 2009/ Accepted 20 July 2009

Three dengue virus type 4 (DENV-4) vaccine candidates containing deletions in the 3' noncoding region were prepared by passage in DBS-FRhL-2 (FRhL) cells. Unexpectedly, these vaccine candidates and parental DENV-4 similarly passaged in the same cells failed to elicit either viremia or a virus-neutralizing antibody response. Consensus sequence analysis revealed that each of the three viruses, as well as the parental DENV-4 when passaged in FRhL cells, rapidly acquired a single Glu327-Gly substitution in domain III (DIII) of the envelope protein (E). These variants appear to have accumulated in response to growth adaptation to FRhL cells as shown by growth analysis, and the mutation was not detected in the virus following passage in C6/36 cells, primary African green monkey kidney cells, or Vero cells. The Glu327-Gly substitution was predicted by molecular modeling to increase the net positive charge on the surface of E. The Glu327-Gly variant of the full-length DENV-4 selected after three passages in FRhL cells showed increased affinity for heparan sulfate compared to the unpassaged DENV-4, as measured by heparin binding and infectivity inhibition assays. Evidence indicates that the Glu327-Gly mutation in DIII of the DENV-4 E protein was responsible for reduced infectivity and immunogenicity in rhesus monkeys. Our results point out the importance of cell substrates for vaccine preparation since the virus may change during passages in certain cells through adaptive selection, and such mutations may affect cell tropism, virulence, and vaccine efficacy.

* Corresponding author. Mailing address: Molecular Viral Biology Section, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 50 South Drive, Bethesda, MD 20892-8005. Phone: (301) 594-2422. Fax: (301) 496-8312. E-mail: clai{at}niaid.nih.gov

{triangledown} Published ahead of print on 5 August 2009.

Journal of Virology, October 2009, p. 10384-10394, Vol. 83, No. 20
0022-538X/09/$08.00+0     doi:10.1128/JVI.01083-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»