Infectious simian/human immunodeficiency virus with human immunodeficiency virus type 1 subtype C from an African isolate: rhesus macaque model

Abstract

Human immunodeficiency virus type 1 (HIV-1) subtype C is responsible for more than 56% of all infections in the HIV and AIDS pandemic. It is the predominant subtype in the rapidly expanding epidemic in southern Africa. To develop a relevant model that would facilitate studies of transmission, pathogenesis, and vaccine development for this subtype, we generated SHIV(MJ4), a simian/human immunodeficiency virus (SHIV) chimera based on HIV-1 subtype C. SHIV(MJ4) contains the majority of env, the entire second exon of tat, and a partial sequence of the second exon of rev, all derived from a CCR5-tropic, primary isolate envelope clone from southern Africa. SHIV(MJ4) replicated efficiently in human, rhesus, and pig-tailed macaque peripheral blood mononuclear cells (PBMCs) in vitro but not in CEMx174 cells. To assess in vivo infectivity, SHIV(MJ4) was intravenously inoculated into four rhesus macaques (Macaca mulatta). All four animals became infected as determined through virus isolation, PCR analysis, and viral loads of 10(7) to 10(8) copies of viral RNA per ml of plasma during the primary infection phase. We have established a CCR5-tropic SHIV(MJ4)/rhesus macaque model that may be useful in the studies of HIV-1 subtype C immunology and biology and may also facilitate the evaluation of vaccines to control the spread of HIV-1 subtype C in southern Africa and elsewhere.

FIG. 1

Mutations in SHIV-89.6PD-MC1, the plasmid used as the backbone for construction of SHIV_MJ4. A deletion in the HIV-1 env glycoprotein C-terminal region of SHIV-89.6P results in an in-frame fusion between HIV-1 and SIV env sequences and preserves the nef reading frame. The underlined amino acids represent HIV-1 sequences from SHIV-89.6 that have been deleted. ORF, open reading frame.

FIG. 2

Genomic organization of the SHIV_MJ4 chimera. Genetic construction of SHIVMJ4 was carried out by replacing the KpnI-BamHI fragment of SHIV-89.6PD-MC1 with a complementary fragment of the env gene from 96BWMOLE1 sample (MOLE 1 env). TM, transmembrane domain of gp41; LTR, long terminal repeat.

FIG. 3

Replication of cell-free SHIV_MJ4, SHIV-89.6, and SHIV-89.6PD-MC1 in human PBMCs. 293T cells were transfected with SHIV constructs and cocultured with PHA-stimulated human PBMCs. Supernatants from the cocultivation were then filtered through 0.45-μm filters and used to infect freshly activated PBMCs.

FIG. 4

Replication of SHIV_MJ4 and SHIV-89.6 in rhesus macaques (A) and pig-tailed macaques (B). The data shown represent a typical replication in PBMCs from one animal of either species.

FIG. 5

Western blot analysis of seroconversion of two inoculated rhesus macaques. A full-length HIV-1 clone expressing HIV-1 subtype C envelope was used as the source of antigen. The antibody response was analyzed by incubation with a 1:100 dilution of plasma obtained from the animals at various time points postinoculation.

FIG. 6

Comparison of the genetic composition and replication properties of SHIV_MJ4 and SHIV_CHN19, the only other HIV-1 subtype C-based SHIV chimera. LTR, long terminal repeat.