Temporal analyses of virus replication, immune responses, and efficacy in rhesus macaques immunized with a live, attenuated simian immunodeficiency virus vaccine

Abstract

Despite evidence that live, attenuated simian immunodeficiency virus (SIV) vaccines can elicit potent protection against pathogenic SIV infection, detailed information on the replication kinetics of attenuated SIV in vivo is lacking. In this study, we measured SIV RNA in the plasma of 16 adult rhesus macaques immunized with a live, attenuated strain of SIV (SIVmac239Deltanef). To evaluate the relationship between replication of the vaccine virus and the onset of protection, four animals per group were challenged with pathogenic SIVmac251 at either 5, 10, 15, or 25 weeks after immunization. SIVmac239Deltanef replicated efficiently in the immunized macaques in the first few weeks after inoculation. SIV RNA was detected in the plasma of all animals by day 7 after inoculation, and peak levels of viremia (10(5) to 10(7) RNA copies/ml) occurred by 7 to 12 days. Following challenge, SIVmac251 was detected in all of the four animals challenged at 5 weeks, in two of four challenged at 10 weeks, in none of four challenged at 15 weeks, and one of four challenged at 25 weeks. One animal immunized with SIVmac239Deltanef and challenged at 10 weeks had evidence of disease progression in the absence of detectable SIVmac251. Although complete protection was not achieved at 5 weeks, a transient reduction in viremia (approximately 100-fold) occurred in the immunized macaques early after challenge compared to the nonimmunized controls. Two weeks after challenge, SIV RNA was also reduced in the lymph nodes of all immunized macaques compared with control animals. Taken together, these results indicate that host responses capable of reducing the viral load in plasma and lymph nodes were induced as early as 5 weeks after immunization with SIVmac239Deltanef, while more potent protection developed between 10 and 15 weeks. In further experiments, we found that resistance to SIVmac251 infection did not correlate with the presence of antibodies to SIV gp130 and p27 antigens and was achieved in the absence of significant neutralizing activity against the primary SIVmac251 challenge stock.

FIG. 1

SIV RNA in plasma following immunization with SIVmac239Δnef and challenge with SIVmac251. Viremia was determined by measuring total SIV RNA in plasma in a bDNA assay (sensitivity of 10⁴ RNA copies/ml). Immunization with SIVmac239Δnef () was performed on day 0. The animals were then challenged with SIVmac251 () at either 5 (A), 10 (B), 15 (C), or 25 (D) weeks. Levels of SIV RNA for the nonimmunized control animal in each challenge group are designated by open symbols.

FIG. 2

Comparison of SIV RNA in the plasma (▵) and lymph nodes (⧫) 2 weeks after challenge with SIVmac251. Lymph node biopsy samples were collected, and the tissues were divided into quadruplicate samples. SIV copy number was determined for each sample by bDNA assay, and the data were normalized per 10 mg of tissue. Individual data points, as well as the geometric mean, are presented for the quadruplicate samples. Paired plasma samples were analyzed for SIV RNA, and the data are expressed as SIV copies per ml. Data for controls depict the results obtained for the nonimmunized animals in each challenge group.

FIG. 3

Antibody responses to SIV gp130 (•) and p27 (▵) measured by ELISA following immunization with SIVmac239Δnef () and challenge with SIVmac251 () of animals in 5-week (A), 10-week (B), 15-week (C), and 25-week (D) challenge groups. Data for nonimmunized control animals (1480, 1492, 1504, and 1516) are shown on the far right.

FIG. 4

Antibody titers to SIV gp130 and p27 measured by ELISA on the day of challenge for each of the 16 macaques immunized with SIVmac239Δnef. The results are presented based on the outcome following challenge, irrespective of the challenge group (unprotected [n = 7] or protected [n = 9]).

FIG. 5

CD4⁺ T-cell counts for immunized and control animals in the 5-week (A), 10-week (B), 15-week (C), and 25-week (D) challenge groups. The results for nonimmunized control animals are depicted by open symbols. , day of challenge with SIVmac251.

FIG. 6

DNA PCR analysis of sequential PBMC and plasma samples from animal 1490. Sequences in nef were amplified by nested DNA PCR using primers that span the deleted region in SIVmac239Δnef as described in Materials and Methods. (A) Sequential PBMC samples obtained before and after challenge with SIVmac251 (70*, day of challenge); (B) plasma samples from sequential time points following coculture with CEMx174 cells. M, molecular size markers; (+), amplification of nef sequences from rhesus PBMC used to propagate the SIVmac239Δnef immunization stock; (−), no DNA control.