RESEARCH ARTICLE Comparison of Major and Minor Viral SNPs Identified through Single Template Sequencing and Pyrosequencing in Acute HIV-1 Infection

Massively parallel sequencing (MPS) technologies, such as 454-pyrosequencing, allow for the identification of variants in sequence populations at lower levels than consensus sequencing and most single-template Sanger sequencing experiments. We sought to deter-mine if the greater depth of population sampling attainable using MPS technology would allow detection of minor variants in HIV founder virus populations very early in infection in instances where Sanger sequencing detects only a single variant. We compared single nucleotide polymorphisms (SNPs) during acute HIV-1 infection from 32 subjects using both single template Sanger and 454-pyrosequencing. Pyrosequences from a median of 2400 viral templates per subject and encompassing 40 % of the HIV-1 genome, were compared to a median of five individually amplified near full-length viral genomes sequenced using Sanger technology. There was no difference in the consensus nucleotide sequences over the 3.6kb compared in 84 % of the subjects infected with single founders and 33 % of sub-jects infected with multiple founder variants: among the subjects with disagreements, mis-matches were found in less than 1 % of the sites evaluated (of a total of nearly 117,000 site