Abstract

HIV undergoes a high rate of mutation and recombination during reverse transcription, but it is not known whether these events occur independently or are linked mechanistically. Here we use a system of silent marker mutations in HIV and a single round of infection in primary T-lymphocytes, combined with a high-throughput sequencing and mathematical modelling approach to directly estimate the viral recombination and mutation rates. From >7 million nt of sequences from HIV infection, we observe 4801 recombination events and 859 substitution mutations ( approximately 1.51 and 0.12 events per 1000 nt respectively). We use experimental controls to account for PCR-induced and transfection-induced recombination and sequencing error. We find the single cycle virus-induced mutation rate is 4.6 x 10-5 mutations per nt after correction. By sorting our data into recombined and non-recombined sequences, we find a significantly higher mutation rate in recombined regions (p=0.003, Fisher’s exact). We use a permutation approach to eliminate a number of potential confounding factors and confirm that mutation occurs around the site of recombination, and is not simply co-located in the genome. By comparing mutation rates in recombined and non-recombined regions we find that recombination-associated mutations account for 15-20% of all mutations occurring during reverse transcription.