Background:  We recently defined the properties of viruses present in 8 heterosexual donor-recipient pairs near the time of transmission. Viruses that established infection uniformly exhibited shorter length and fewer N-linked glycosylation (N-gly) sites in the gp120 V1-V4 region; limited sequence divergence as evidenced by short branch lengths in maximum likelihood analyses; and high sensitivity to neutralization relative to the quasi-species in the infecting partner. In contrast, these parameters were broadly variable in the donor quasi-species. Utilizing this sequence and neutralization data, we investigated whether the observed sequence-based differences predicted neutralization susceptibility.

Methods:  A Kendall’s rank correlation t non-parametric statistical test was applied to determine whether neutralization sensitivity exhibited a significant correlation with length and number of N-gly sites in V1-V4, as well as branch length, using donor and recipient Envs from 5 transmission pairs.

Results:  Branch length (a measure of sequence divergence) was highly correlated with neutralization by both autologous (p = 0.0002) and heterologous antibodies (p = 0.001). V1-V4 length (a measure of insertions/deletions) showed a significant negative correlation with autologous (p = 0.01), but not heterologous, neutralization sensitivity. In contrast, no significant correlation was observed between the number of N-gly sites in V1-V4 and autologous or heterologous neutralization. Consistent with the existence of multiple neutralization escape pathways, no relationship was observed between neutralizing antibody sensitivity and phylogenetic clustering on maximum likelihood trees.

Conclusions:  The fact that sequences closest to the ancestral node are the most sensitive to neutralization argues that there is a significant effect of base substitutions outside of the variable loop domains (which are stripped out of the maximum likelihood tree) on this biological property of Env. However, because sequences with compact variable loops are also more sensitive to neutralization by donor plasma, it is likely that acquisition of length in V1V2 and V4 is involved in autologous neutralization escape. The finding that the number of N-gly sites within the V1-V4 region is not correlated with neutralization sensitivity argues that changes in position, not accumulation of sites, is more important for neutralization escape.

Keywords: Transmission; Neutralization; variable loops