Vaccine effect using a live attenuated nef-deficient simian immunodeficiency virus of African green monkeys in the absence of detectable vaccine virus replication in vivo

Abstract

Immunization of adult macaques with live attenuated simian immunodeficiency viruses (SIVs) lacking the nef genes has been shown to protect against challenge with full-length pathogenic SIV. To test live attenuated virus vaccines for the first time in a natural host we have constructed a mutant SIV from African green monkeys (SIVagm) with a deletion of 125 bp in the nef gene (SIVagm3 delta nef). This mutant showed moderately delayed in vitro replication in the T cell line MOLT-4/8 and in primary peripheral blood mononuclear cells from African green monkeys (Cercopithecus aetiops) and pig-tailed macaques (Macaca nemestrina) compared with cloned wild-type SIVagm3. In contrast, in vivo replication of SIVagm3 delta nef in African green monkeys was severely impaired or undetectable and did not induce seroconversion. After challenge with wild-type SIVagm3 the SIVagm3 delta nef preinoculated African green monkeys showed a memory antibody response that declined after week 2. In three of four African green monkeys the cell-associated virus load and in two of four African green monkeys the plasma virus load was dramatically decreased after the challenge compared with naive control animals. The remaining animal showed no evidence of productive challenge virus replication. This study demonstrates that a strong vaccine effect or protection in the SIVagm/African green monkey system is possible using a live attenuated vaccine in the absence of a productive infection and corresponding humoral immune response.

Figure 1

(A) Growth kinetics of SIVagm3Δnef and of SIVagm3 (nef open) in MOLT-4/8 cells. MOLT-4/8 cells were infected in duplicate at a multiplicity of infection of 4 × 10⁻⁴ either with SIVagm3Δnef, SIVagm3 (nef open), or exposed to medium alone. Supernatants were assayed for RT activity. (B) Growth kinetics of SIVagm3Δnef and of SIVagm3 on phytohemagglutinin blasts of an SIVagm-negative African green monkey or a pig-tailed macaque. PBMCs were infected with a multiplicity of infection of 3 × 10⁻⁴ either with SIVagm3Δnef, SIVagm3 (nef open), or exposed to medium alone. Supernatants were assayed for RT activity. M. nem, Macaca nemestrina.

Figure 2

Plasma virus loads in the (A) SIVagm3Δnef immunized (AGM 111, 112, 113, 115) and the (B) control (AGM 158, 159, 160, 190) animals. The RT activities of the plasma (fg RT per 100 ml of plasma) were determined using the very sensitive PERT assay as described above. Open area, before challenge with SIVagm3; shaded area, after challenge with SIVagm3. Only AGM 113 exhibited prechallenge RT activity at 16, 25, and 31 weeks.

Figure 3

Binding antibodies to whole SIVagm antigen in the (A) SIVagm3Δnef immunized (AGM 111, 112, 113, 115) and the (B) control (AGM 158, 159, 160, 190) animals. The binding antibodies were determined using a standard ELISA protocol. Open area, before challenge with SIVagm3; shaded area, after challenge with SIVagm3.