Background: Infection with human immunodeficiency virus type 1 (HIV) typically results from transmission of a small and genetically uniform viral population. Following transmission, the virus population becomes more diverse because of recombination and acquired mutations through genetic drift and selection. Viral intrahost genetic diversity remains a major obstacle to the cure of HIV; however, there is a disagreement whether intrahost viral genetic diversification associates positively or negatively with disease progression and progression markers. Viral load is a key progression marker and understanding its relationship to viral intrahost genetic diversity could help design future strategies for HIV monitoring and treatment.

Methods: We analyzed deep-sequenced viral genomes from 2,650 treatment-naive HIV-infected persons to measure the intrahost genetic diversity of 2,447 genomic codon positions as calculated by Shannon entropy. We tested for associations between viral load (VL) and amino acid (AA) entropy accounting for sex, age, race, duration of infection, and HIV population structure.

Results: We confirmed that the intrahost genetic diversity is highest in the env gene. Furthermore, we showed that mean Shannon entropy is significantly associated with VL, especially in infections of >24 months duration. We identified 16 significant associations between VL (p-value<2.0x10^-5) and Shannon entropy at AA positions which in our association analysis explained 13% of the variance in VL.

Conclusions: Our results elucidate that viral intrahost genetic diversity is associated with VL and could be used as a better disease progression marker than HIV consensus sequence variants, especially in infections of longer duration. We emphasize that viral intrahost diversity should be considered when studying viral genomes and infection outcomes.