ABSTRACTThe oral microbiome is an important predictor of health and disease. We recently reported significant yet modest effects of HIV under highly active antiretroviral therapy (ART) on the oral microbiome (bacterial and fungal) in a large cohort of HIV-positive (HIV+) and matched HIV-negative (HIV−) individuals. As it was unclear whether ART added to or masked further effects of HIV on the oral microbiome, the present study aimed to analyze the effects of HIV and ART independently, which also included HIV−subjects on preexposure prophylaxis (PrEP) therapy. Cross-sectional analyses of the effect of HIV devoid of ART (HIV+ART−versus matched HIV−subjects) showed a significant effect on both the bacteriome and mycobiome (P< 0.024) after controlling for other clinical variables (permutational multivariate analysis of variance [PERMANOVA] of Bray-Curtis dissimilarity). Cross-sectional analyses evaluating the effects of ART (HIV+ART+versus HIV+ART−subjects) revealed a significant effect on the mycobiome (P< 0.007) but not the bacteriome. In parallel longitudinal analyses, ART (before versus after the initiation of ART) had a significant effect on the bacteriome, but not the mycobiome, of HIV+and HIV−PrEP subjects (P< 0.005 andP< 0.016, respectively). These analyses also revealed significant differences in the oral microbiome and several clinical variables between HIV−PrEP subjects (pre-PrEP) and the HIV-matched HIV−group (P< 0.001). At the species level, a small number of differences in both bacterial and fungal taxa were identified within the effects of HIV and/or ART. We conclude that the effects of HIV and ART on the oral microbiome are similar to those of the clinical variables but collectively are modest overall.