Broadly neutralizing antibodies have been associated with spontaneous clearance of the hepatitis C infection as well as viral persistence by immune escape. Further study of neutralizing antibody epitopes is needed to unravel pathways of resistance to virus neutralization, and to identify conserved regions for vaccine design. All reported broadly neutralizing antibody (BNAb) epitopes in the HCV Envelope (E2) glycoprotein were identified. The critical contact residues of these epitopes were mapped onto the linear E2 sequence. All publicly available E2 sequences were then downloaded and the contact residues within the BNAb epitopes were assessed for the level of conservation, as well as the frequency of occurrence of experimentally-proven resistance mutations. Epitopes were also compared between two sequence datasets obtained fromsamples collected atwell-defined time points from acute (180 days) infections, to identify any significant differences in residue usage. The contact residues for all BNAbswere containedwithin 3 linear regions of the E2 protein sequence. An analysis of 1749 full length E2 sequences frompublic databases showed that only 10 out of 29 experimentally-proven resistance mutations were present at a frequency >5%. Comparison of subtype 1a viral sequences obtained from samples collected during acute or chronic infection revealed significant differences at positions 610 and 655 with changes in residue (p < 0.05), and at position 422 (p < 0.001) with a significant difference in variability (entropy). The majority of experimentally-described escape variants do not occur frequently in nature. The observed differences between acute and chronically isolated sequences suggest constraints on residue usage early in infection. (C) 2017 Elsevier B. V. All rights reserved.