BackgroundH2O2 produced by vaginal lactobacilli is believed to protect against infection, and H2O2-producing lactobacilli inactivate pathogens in vitro in protein-free salt solution. However, cervicovaginal fluid (CVF) and semen have significant H2O2-blocking activity.MethodsWe measured the H2O2 concentration of CVF and the H2O2-blocking activity of CVF and semen using fluorescence and in vitro bacterial-exposure experiments.ResultsThe mean H2O2 measured in fully aerobic CVF was 23 ± 5 μM; however, 50 μM H2O2 in salt solution showed no in vitro inactivation of HSV-2, Neisseria gonorrhoeae, Hemophilus ducreyii, or any of six BV-associated bacteria. CVF reduced 1 mM added H2O2 to an undetectable level, while semen reduced 10 mM added H2O2 to undetectable. Moreover, the addition of just 1% CVF supernatant abolished in vitro pathogen-inactivation by H2O2-producing lactobacilli.ConclusionsGiven the H2O2-blocking activity of CVF and semen, it is implausible that H2O2-production by vaginal lactobacilli is a significant mechanism of protection in vivo.