Simian immunodeficiency virus tropism and evolution in the central nervous system: A molecular clone derived from the brain of a pig-tailed macaque displays a macrophage-restricted phenotype linked to a single amino acid change in env

The simian immunodeficiency virus (SIV) isolate SIVsmmFGb, initially derived from a sooty mangabey, causes neuropathogenic infection in approximately 90% of infected pig-tailed macaques and provides an excellent model for investigating mechanisms of HIV/SIV neuropathogenesis. A molecular clone generated from the brain of an SIVsmmFGb-infected pig-tailed macaque has been characterized in vitro and displays a strictly macrophage-tropic phenotype not previously described in HIV/SIV literature. This clone, BPZm.12, replicates to high levels in macrophages; however, BZPm.12 does not replicate in peripheral blood mononuclear cells (PBMC) until at least 21 days post inoculation. Immunomagnetic separation was used to determine that the target cells for BPZm.12 replication after 21 days are CD3+ cells. Therefore, BPZm.12 undergoes a switch in replication phenotype from macrophage tropic to dual tropic after culture in PBMC.; Genetic analysis of the virus replicating in PBMC after 21 days post inoculation revealed a single nucleotide change in env. This nucleotide change results in the reversion of a serine residue to a highly conserved proline residue at position 629 in the ectodomain of the envelope glycoprotein gp41. A mutant clone of BPZm.12, replacing the serine with the conserved proline, was generated by site-directed mutagenesis. The mutant, BPZm.12-629P, replicates in both macrophages and PBMC, confirming that the single amino acid change causes the switch in tropism. Additionally, a mutant of the dual tropic molecular clone PBj6.6 was generated, substituting the conserved proline for serine. This mutant, PBj6.6-629S, replicates to high levels in macrophages, but does not replicate in PBMC at any time point. These results indicate that a single determinant in gp41 of an SIV clone changes its phenotype from macrophage tropic to dual tropic.; Sequence analysis of BPZm.l2 also revealed a premature stop codon causing the truncation of gp41 downstream of it transmembrane domain. The premature stop codon is conserved in the BPZm.12 virus stock and virus replicating after 21 days in PBMC culture. Sequence analysis of other env clones revealed that approximately 30% of clones derived from the brain of PZm contained premature stop codons, while no clones from the lymph node contained such mutations. These results support the hypothesis that viral evolution can and does occur within the central nervous system, separate from the evolution that occurs in the peripheral lymphoid system.