OBJECTIVE: We sought to investigate the effects of HIV infection on the vaginal microbiota and associations with treatment and demographic factors. We thus compared vaginal microbiome samples from HIV-infected (HIV+) and HIV-uninfected (HIV-) women collected at two Chicago area hospitals. DESIGN: We studied vaginal microbiome samples from 178 women analyzed longitudinally (n = 324 samples) and collected extensive data on clinical status and demographic factors. METHODS: We used 16S rRNA gene sequencing to characterize the bacterial lineages present, then UniFrac, Shannon diversity, and other measures to compare community structure with sample metadata. RESULTS: Differences in microbiota measures were modest in the comparison of HIV+ and HIV- samples, in contrast to several previous studies, consistent with effective antiretroviral therapy. Proportions of healthy Lactobacillus species were not higher in HIV- patients overall, but were significantly higher when analyzed within each hospital in isolation. Rates of bacterial vaginosis were higher among African-American women and HIV+ women. Bacterial vaginosis was associated with higher frequency of HIV+. Unexpectedly, African-American women were more likely to switch bacterial vaginosis status between sampling times; switching was not associated with HIV+ status. CONCLUSION: The influence of HIV infection on the vaginal microbiome was modest for this cohort of well suppressed urban American women, consistent with effective antiretroviral therapy. HIV+ was found to be associated with bacterial vaginosis. Although bacterial vaginosis has previously been associated with HIV transmission, most of the women studied here became HIV+ many years before our test for bacterial vaginosis, thus implicating additional mechanisms linking HIV infection and bacterial vaginosis.