Analysis of evolutionary rate of HIV-1 subtype B using blood donor samples in Japan

There are few reports on HIV-1 intra-host evolutionary rate in asymptomatic treatment-naïve patients. Here, the HIV-1 intra-host evolutionary rate was estimated based on HIV-1 RNA sequences from plasma samples of blood donors in Japan. Blood donors were assumed to have received no treatment for and have no symptoms of HIV-1 infection because they were healthy, and declared no risky behaviors of HIV-1 infection on a self-reported questionnaire or interview followed by donation. HIV-1 RNA was obtained from 85 plasma samples from 36 blood donors who donated blood multiple times and were HIV-1-positive. The C2V3C3 region which encodes for a part of the envelope protein, and the V3 loop in the C2V3C3 region were analyzed by RT-PCR and direct sequencing, and the sequences were compared. The nucleotide substitution rate was calculated by linear regression. All HIV-1 samples analyzed were classified as subtype B. The mean nucleotide substitution rate in C2V3C3 was calculated to be 6.2 × 10^-3-1.8 × 10^-2/site/year (V3: 4.5 × 10^-3-2.3 × 10^-2/site/year). The mean non-synonymous substitution rate in C2V3C3 was calculated to be 5.2 × 10^-3-1.7 × 10^-2/site/year (V3: 4.5 × 10^-3-2.1 × 10^-2/site/year). The mean synonymous substitution rate in C2V3C3 was calculated to be 1.1 × 10^-4-2.3 × 10^-3/site/year (V3: 2.9 × 10^-3/site/year). Among HIV-1 subtype B RNA-positive blood donors in Japan, the nucleotide substitution rate in C2V3C3 was estimated to be higher than that of reported cases using HIV-1 samples mainly obtained from AIDS patients. Compared to AIDS patients, immune responses against HIV-1 are probably more effective in HIV-1 RNA-positive blood donors. Consequently, immune pressure presumably promotes mutation of the virus genome.