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Background Human immunodeficiency virus (HIV)-1 genetic diversity increases during infection and can help infer the time elapsed since infection. However, the effect of antiretroviral treatment (ART) on the inference remains unknown. Methods Participants with estimated duration of HIV-1 infection based on repeated testing were sourced from cohorts in Botswana (n = 1944). Full-length HIV genome sequencing was performed from proviral deoxyribonucleic acid. We optimized a machine learning model to classify infections as &amp;lt; or &amp;gt;1 year based on viral genetic diversity, demographic, and clinical data. Results The best predictive model included variables for genetic diversity of HIV-1 gag, pol, and env, viral load, age, sex, and ART status. Most participants were on ART. Balanced accuracy was 90.6% (95% confidence interval, 86.7%–94.1%). We tested the algorithm among newly diagnosed participants with or without documented negative HIV tests. Among those without records, those who self-reported a negative HIV test within &amp;lt;1 year were more frequently classified as recent than those who reported a test &amp;gt;1 year previously. There was no difference in classification between those self-reporting a negative HIV test &amp;lt;1 year, whether or not they had a record. Conclusions These results indicate that recency of HIV-1 infection can be inferred from viral sequence diversity even among patients on suppressive ART.

Human Immunodeficiency Virus (HIV) Genetic Diversity Informs Stage of HIV-1 Infection Among Patients Receiving Antiretroviral Therapy in Botswana