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J Virol, July 1998, p. 5680-5698, Vol. 72, No. 7
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

A Comprehensive Panel of Near-Full-Length Clones and Reference Sequences for Non-Subtype B Isolates of Human Immunodeficiency Virus Type 1

Feng Gao,1 David L. Robertson,1,dagger Catherine D. Carruthers,1 Sandra G. Morrison,1 Bixi Jian,1 Yalu Chen,1 Françoise Barré-Sinoussi,2 Marc Girard,3 Alagarsamy Srinivasan,4 Alash'le G. Abimiku,5 George M. Shaw,1,6 Paul M. Sharp,7 and Beatrice H. Hahn1,*

Department of Medicine and Microbiology1 and Howard Hughes Medical Institute,6 University of Alabama at Birmingham, Birmingham, Alabama 35294; Unité de Biologie des Retrovirus, Institut Pasteur, Paris 75724,2 and Laboratory of Molecular Virology, Institut Pasteur, Paris 75015,3 France; Department of Microbiology and Immunology, Jefferson Cancer Institute, Thomas Jefferson University, Philadelphia, Pennsylvania 191074; Institute of Human Virology, Baltimore, Maryland 212015; and Division of Genetics, University of Nottingham, Queens Medical Center, Nottingham, United Kingdom7

Received 9 December 1997/Accepted 30 March 1998

Non-subtype B viruses cause the vast majority of new human immunodeficiency virus type 1 (HIV-1) infections worldwide and are thus the major focus of international vaccine efforts. Although their geographic dissemination is carefully monitored, their immunogenic and biological properties remain largely unknown, in part because well-characterized virological reference reagents are lacking. In particular, full-length clones and sequences are rare, since subtype classification is frequently based on small PCR-derived viral fragments. There are only five proviral clones available for viruses other than subtype B, and these represent only 3 of the 10 proposed (group M) sequence subtypes. This lack of reference sequences also confounds the identification and analysis of mosaic (recombinant) genomes, which appear to be arising with increasing frequency in areas where multiple sequence subtypes cocirculate. To generate a more representative panel of non-subtype B reference reagents, we have cloned (by long PCR or lambda phage techniques) and sequenced 10 near-full-length HIV-1 genomes (lacking less than 80 bp of long terminal repeat sequences) from primary isolates collected at major epicenters of the global AIDS pandemic. Detailed phylogenetic analyses identified six that represented nonrecombinant members of HIV-1 subtypes A (92UG037.1), C (92BR025.8), D (84ZR085.1 and 94UG114.1), F (93BR020.1), and H (90CF056.1), the last two comprising the first full-length examples of these subtypes. Four others were found to be complex mosaics of subtypes A and C (92RW009.6), A and G (92NG083.2 and 92NG003.1), and B and F (93BR029.4), again emphasizing the impact of intersubtype recombination on global HIV-1 diversification. Although a number of clones had frameshift mutations or translational stop codons in major open reading frames, all the genomes contained a complete set of genes and three had intact genomic organizations without inactivating mutations. Reconstruction of one of these (94UG114.1) yielded replication-competent virus that grew to high titers in normal donor peripheral blood mononuclear cell cultures. This panel of non-subtype B reference genomes should prove valuable for structure-function studies of genetically diverse viral gene products, the generation of subtype-specific immunological reagents, and the production of DNA- and protein-based subunit vaccines directed against a broader spectrum of viruses.

* Corresponding author. Mailing address: Department of Medicine and Microbiology, University of Alabama at Birmingham, 701 S. 19th St., LHRB 613, Birmingham, AL 35294. Phone: (205) 934-0412. Fax: (205) 934-1580. E-mail: bhahn{at}cordelia.dom.uab.edu.

dagger Present address: Laboratory of Structural and Genetic Information, CNRS-EP 91, Marseilles, France 13402.

J Virol, July 1998, p. 5680-5698, Vol. 72, No. 7
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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