Background:  Rare individuals having repeated, unprotected sexual exposure to HIV-1 remain seronegative and appear to resist overt infection. We identified 4 of 81 exposed seronegative  volunteers and 2 of 34 low-risk controls demonstrating markedly reduced CD4⁺ T cell susceptibility to HIV-1 infection in vitro. To determine whether HIV-1 resistance in vitro might contribute to continued seronegativity in vivo, we investigated the mechanisms responsible for the observed defect in CD4⁺ T-cell susceptibility in these 6 subjects compared with 4 persons of normal susceptibility.

Methods:  Susceptibility to 3 R5 HIV-1 isolates in vitro was assessed by intracellular Gag expression in single-round infection and p24 antigen production at 3, 5, 7, 10, and 12 days post-infection. TCID₅₀ was defined for JR-CSF and LAI strains. Findings were fitted to a rate-equation model to estimate the basic reproductive number in vitro (R₀). Comparison of cohort susceptibility was performed by t-test. Data from subjects having in vitro HIV-1 resistance or susceptibility was assessed by Mann-Whitney test. Exclusion criteria were CCR5Δ32 homozygosity, CD8⁺ T-cell contamination > 5%, or viability < 60% in uninfected cultures run in parallel.

Results:  Testing of CD4⁺ T cells from 6 persons having reduced in vitro HIV-1 susceptibility revealed a reproducible 10- to 1000-fold decline in ability to sustain in vitro infection (median log pg p24/mL 2.35 vs 4.50 in susceptible volunteers). CD4⁺ T cells from these subjects demonstrated a 10-fold increase in the viral inoculum required to establish infection (median TCID₅₀/μL 26.8 vs 374.3, p = 0.008), and a 3.5-fold reduction in R₀ (median 1.58 vs 5.45, p = 0.008).

Conclusions:  The pronounced reduction in CD4⁺ T-cell susceptibility and corresponding decrease in R₀ observed in 4 exposed seronegative subjects and 2 low-risk controls may provide protection from overt HIV-1 infection in vivo in this subset of individuals. Because in vitro infectivity culture conditions do not directly simulate in vivo infection, the reported values of R₀ cannot be regarded as true reproductive numbers in vivo. However, the relative values of R₀ and TCID₅₀ may reflect significant biological differences between the 2 groups (normal and low HIV-1 susceptibility). Given the low rate of sexual transmission of HIV-1, the markedly diminished R₀ values, and increased viral inoculum required to establish infection that we report here may contribute to these volunteers’ continued seronegativity.

Keywords: exposed seronegative; cellular resistance; basic reproductive number