The detection of a 0-month Env that was resistant to one of the Mabs was unexpected given the early timing and high sensitivity of these Envs to patient plasma. Therefore, the frequency of this predicted glycan site in V1 during acute/early infection was investigated using 21 uncloned single genome amplified V1V4 sequences from a p24-positive, antibody-negative sample (1-Mar-03) and 31 from the 0-month sample (27-Mar-03), which was antibody positive [38] (see Table 1 ). These sequences were combined with the five cloned 0-month Envs from this study, and a highlighter plot was created using the HIV Database (Fig. S6) . Forty-four out of 57 sequences (77%) were identical throughout V1V4, and all of these contained the predicted glycan addition site in V1. Two variants (including EnvPL6.3), both from the antibody positive time point, carried an identical G to A mutation that abrogated the predicted V1 glycan addition site. Taken together, these observations provided strong evidence that the founder virus, like the 0-month Envs PB1.1 and PL4.1, carried the predicted glycan site in V1, and that the mutation in 0-month EnvPL6.3 arose shortly after transmission, but circulated only transiently.